BioMed Research International: Genomics http://www.hindawi.com The latest articles from Hindawi Publishing Corporation © 2014 , Hindawi Publishing Corporation . All rights reserved. Dynamic Alu Methylation during Normal Development, Aging, and Tumorigenesis Wed, 27 Aug 2014 12:02:53 +0000 http://www.hindawi.com/journals/bmri/2014/784706/ DNA methylation primarily occurs on CpG dinucleotides and plays an important role in transcriptional regulations during tissue development and cell differentiation. Over 25% of CpG dinucleotides in the human genome reside within Alu elements, the most abundant human repeats. The methylation of Alu elements is an important mechanism to suppress Alu transcription and subsequent retrotransposition. Decades of studies revealed that Alu methylation is highly dynamic during early development and aging. Recently, many environmental factors were shown to have a great impact on Alu methylation. In addition, aberrant Alu methylation has been documented to be an early event in many tumors and Alu methylation levels have been associated with tumor aggressiveness. The assessment of the Alu methylation has become an important approach for early diagnosis and/or prognosis of cancer. This review focuses on the dynamic Alu methylation during development, aging, and tumor genesis. The cause and consequence of Alu methylation changes will be discussed. Yanting Luo, Xuemei Lu, and Hehuang Xie Copyright © 2014 Yanting Luo et al. All rights reserved. Integration Strategy Is a Key Step in Network-Based Analysis and Dramatically Affects Network Topological Properties and Inferring Outcomes Wed, 27 Aug 2014 08:49:41 +0000 http://www.hindawi.com/journals/bmri/2014/296349/ An increasing number of experiments have been designed to detect intracellular and intercellular molecular interactions. Based on these molecular interactions (especially protein interactions), molecular networks have been built for using in several typical applications, such as the discovery of new disease genes and the identification of drug targets and molecular complexes. Because the data are incomplete and a considerable number of false-positive interactions exist, protein interactions from different sources are commonly integrated in network analyses to build a stable molecular network. Although various types of integration strategies are being applied in current studies, the topological properties of the networks from these different integration strategies, especially typical applications based on these network integration strategies, have not been rigorously evaluated. In this paper, systematic analyses were performed to evaluate 11 frequently used methods using two types of integration strategies: empirical and machine learning methods. The topological properties of the networks of these different integration strategies were found to significantly differ. Moreover, these networks were found to dramatically affect the outcomes of typical applications, such as disease gene predictions, drug target detections, and molecular complex identifications. The analysis presented in this paper could provide an important basis for future network-based biological researches. Nana Jin, Deng Wu, Yonghui Gong, Xiaoman Bi, Hong Jiang, Kongning Li, and Qianghu Wang Copyright © 2014 Nana Jin et al. All rights reserved. Comparative Transcriptome Analysis of Leaves and Roots in Response to Sudden Increase in Salinity in Brassica napus by RNA-seq Thu, 07 Aug 2014 06:59:39 +0000 http://www.hindawi.com/journals/bmri/2014/467395/ Amphidiploid species in the Brassicaceae family, such as Brassica napus, are more tolerant to environmental stress than their diploid ancestors. A relatively salt tolerant B. napus line, N119, identified in our previous study, was used. N119 maintained lower Na+ content, and Na+/K+ and Na+/Ca2+ ratios in the leaves than a susceptible line. The transcriptome profiles of both the leaves and the roots 1 h and 12 h after stress were investigated. De novo assembly of individual transcriptome followed by sequence clustering yielded 161,537 nonredundant sequences. A total of 14,719 transcripts were differentially expressed in either organs at either time points. GO and KO enrichment analyses indicated that the same 49 GO terms and seven KO terms were, respectively, overrepresented in upregulated transcripts in both organs at 1 h after stress. Certain overrepresented GO term of genes upregulated at 1 h after stress in the leaves became overrepresented in genes downregulated at 12 h. A total of 582 transcription factors and 438 transporter genes were differentially regulated in both organs in response to salt shock. The transcriptome depicting gene network in the leaves and the roots regulated by salt shock provides valuable information on salt resistance genes for future application to crop improvement. Hui-Yee Yong, Zhongwei Zou, Eng-Piew Kok, Bih-Hua Kwan, Kingsley Chow, Shiori Nasu, Masami Nanzyo, Hiroyasu Kitashiba, and Takeshi Nishio Copyright © 2014 Hui-Yee Yong et al. All rights reserved. Improved Variant Calling Accuracy by Merging Replicates in Whole-Exome Sequencing Studies Mon, 04 Aug 2014 08:22:50 +0000 http://www.hindawi.com/journals/bmri/2014/319534/ In large scale population-based whole-exome sequencing (WES) studies, there are some samples occasionally sequenced two or more times due to a variety of reasons. To investigate how to efficiently utilize these duplicated sequencing data, we conducted comprehensive evaluation of variant calling strategies. 92 samples subjected to WES twice were selected from a large population study. These 92 duplicated samples were divided into two groups: group H consisting of the higher sequencing depth for each subject and group L consisting of the lower depth for each subject. The merged samples for each subject were put in a third group M. Using the GATK multisample toolkit, we compared variant calling accuracy among three strategies. Hierarchical clustering analysis indicated that the two replicates for each subject showed high homogeneity. The comparative analyses on the basis of heterozygous-homozygous ratio (Hete/Homo), transition-transversion ratio (Ti/Tv), and overlapping rate with the 1000 Genomes Project consistently showed that the data quality of the SNPs detected from the M group was more accurate than that of SNPs detected from the H and L groups. These results suggested that merging homogeneous duplicated exomes instead of using one of them could improve variant calling accuracy. Yanfeng Zhang, Bingshan Li, Chun Li, Qiuyin Cai, Wei Zheng, and Jirong Long Copyright © 2014 Yanfeng Zhang et al. All rights reserved. miRNA Transcriptome of Hypertrophic Skeletal Muscle with Overexpressed Myostatin Propeptide Thu, 24 Jul 2014 10:02:43 +0000 http://www.hindawi.com/journals/bmri/2014/328935/ MicroRNAs (miRNAs) play an imperative role in cell proliferation, differentiation, and cell metabolism through regulation of gene expression. Skeletal muscle hypertrophy that results from myostatin depression by its propeptide provides an interesting model to understand how miRNA transcriptome is involved in myostatin-based fiber hypertrophy. This study employed Solexa deep sequencing followed by Q-PCR methods to analyze miRNA transcriptome of skeletal muscle of myostatin propeptide transgenic mice in comparison with their littermate controls. A total of 461 mature known and 69 novel miRNAs were reported from this study. Fifty-seven miRNAs were expressed differentially between transgenic and littermate controls, of which most abundant miRNAs, miR-133a and 378a, were significantly differentially expressed. Expression profiling was validated on 8 known and 2 novel miRNAs. The miRNA targets prediction and pathway analysis showed that FST, SMAD3, TGFBR1, and AcvR1a genes play a vital role in skeletal muscle hypertrophy in the myostatin propeptide transgenic mice. It is predicted that miR-101 targeted to TGFBR1 and SMAD3, miR-425 to TGFBR2 and FST, and miR-199a to AcvR2a and TGF-β genes. In conclusion, the study offers initial miRNA profiling and methodology of miRNA targets prediction for myostatin-based hypertrophy. These differentially expressed miRNAs are proposed as candidate miRNAs for skeletal muscle hypertrophy. Ruheena Javed, Lu Jing, Jinzeng Yang, Xinyun Li, Jianhua Cao, and Shuhong Zhao Copyright © 2014 Ruheena Javed et al. All rights reserved. Informative Gene Selection and Direct Classification of Tumor Based on Chi-Square Test of Pairwise Gene Interactions Wed, 23 Jul 2014 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2014/589290/ In efforts to discover disease mechanisms and improve clinical diagnosis of tumors, it is useful to mine profiles for informative genes with definite biological meanings and to build robust classifiers with high precision. In this study, we developed a new method for tumor-gene selection, the Chi-square test-based integrated rank gene and direct classifier (χ2-IRG-DC). First, we obtained the weighted integrated rank of gene importance from chi-square tests of single and pairwise gene interactions. Then, we sequentially introduced the ranked genes and removed redundant genes by using leave-one-out cross-validation of the chi-square test-based Direct Classifier (χ2-DC) within the training set to obtain informative genes. Finally, we determined the accuracy of independent test data by utilizing the genes obtained above with χ2-DC. Furthermore, we analyzed the robustness of χ2-IRG-DC by comparing the generalization performance of different models, the efficiency of different feature-selection methods, and the accuracy of different classifiers. An independent test of ten multiclass tumor gene-expression datasets showed that χ2-IRG-DC could efficiently control overfitting and had higher generalization performance. The informative genes selected by χ2-IRG-DC could dramatically improve the independent test precision of other classifiers; meanwhile, the informative genes selected by other feature selection methods also had good performance in χ2-DC. Hongyan Zhang, Lanzhi Li, Chao Luo, Congwei Sun, Yuan Chen, Zhijun Dai, and Zheming Yuan Copyright © 2014 Hongyan Zhang et al. All rights reserved. miRNA Signature in Mouse Spermatogonial Stem Cells Revealed by High-Throughput Sequencing Thu, 17 Jul 2014 09:58:16 +0000 http://www.hindawi.com/journals/bmri/2014/154251/ Spermatogonial stem cells (SSCs) play fundamental roles in spermatogenesis. Although a handful of genes have been discovered as key regulators of SSC self-renewal and differentiation, the regulatory network responsible for SSC function remains unclear. In particular, small RNA signatures during mouse spermatogenesis are not yet systematically investigated. Here, using next generation sequencing, we compared small RNA signatures of in vitro expanded SSCs, testis-derived somatic cells (Sertoli cells), developing germ cells, mouse embryonic stem cells (ESCs), and mouse mesenchymal stem cells among mouse embryonic stem cells (ESCs) to address small RNA transition during mouse spermatogenesis. The results manifest that small RNA transition during mouse spermatogenesis displays overall declined expression profiles of miRNAs and endo-siRNAs, in parallel with elevated expression profiles of piRNAs, resulting in the normal biogenesis of sperms. Meanwhile, several novel miRNAs were preferentially expressed in mouse SSCs, and further investigation of their functional annotation will allow insights into the mechanisms involved in the regulation of SSC activities. We also demonstrated the similarity of miRNA signatures between SSCs and ESCs, thereby providing a new clue to understanding the molecular basis underlying the easy conversion of SSCs to ESCs. Tao Tan, Yanfeng Zhang, Weizhi Ji, and Ping Zheng Copyright © 2014 Tao Tan et al. All rights reserved. Large-Scale Genomic Analysis of Codon Usage in Dengue Virus and Evaluation of Its Phylogenetic Dependence Thu, 17 Jul 2014 06:47:59 +0000 http://www.hindawi.com/journals/bmri/2014/851425/ The increasing number of dengue virus (DENV) genome sequences available allows identifying the contributing factors to DENV evolution. In the present study, the codon usage in serotypes 1–4 (DENV1–4) has been explored for 3047 sequenced genomes using different statistics methods. The correlation analysis of total GC content (GC) with GC content at the three nucleotide positions of codons (GC1, GC2, and GC3) as well as the effective number of codons (ENC, ENCp) versus GC3 plots revealed mutational bias and purifying selection pressures as the major forces influencing the codon usage, but with distinct pressure on specific nucleotide position in the codon. The correspondence analysis (CA) and clustering analysis on relative synonymous codon usage (RSCU) within each serotype showed similar clustering patterns to the phylogenetic analysis of nucleotide sequences for DENV1–4. These clustering patterns are strongly related to the virus geographic origin. The phylogenetic dependence analysis also suggests that stabilizing selection acts on the codon usage bias. Our analysis of a large scale reveals new feature on DENV genomic evolution. Edgar E. Lara-Ramírez, Ma Isabel Salazar, María de Jesús López-López, Juan Santiago Salas-Benito, Alejandro Sánchez-Varela, and Xianwu Guo Copyright © 2014 Edgar E. Lara-Ramírez et al. All rights reserved. Advanced Heat Map and Clustering Analysis Using Heatmap3 Wed, 16 Jul 2014 15:52:46 +0000 http://www.hindawi.com/journals/bmri/2014/986048/ Heat maps and clustering are used frequently in expression analysis studies for data visualization and quality control. Simple clustering and heat maps can be produced from the “heatmap” function in R. However, the “heatmap” function lacks certain functionalities and customizability, preventing it from generating advanced heat maps and dendrograms. To tackle the limitations of the “heatmap” function, we have developed an R package “heatmap3” which significantly improves the original “heatmap” function by adding several more powerful and convenient features. The “heatmap3” package allows users to produce highly customizable state of the art heat maps and dendrograms. The “heatmap3” package is developed based on the “heatmap” function in R, and it is completely compatible with it. The new features of “heatmap3” include highly customizable legends and side annotation, a wider range of color selections, new labeling features which allow users to define multiple layers of phenotype variables, and automatically conducted association tests based on the phenotypes provided. Additional features such as different agglomeration methods for estimating distance between two samples are also added for clustering. Shilin Zhao, Yan Guo, Quanhu Sheng, and Yu Shyr Copyright © 2014 Shilin Zhao et al. All rights reserved. Metabolic Modeling of Common Escherichia coli Strains in Human Gut Microbiome Sun, 13 Jul 2014 12:10:37 +0000 http://www.hindawi.com/journals/bmri/2014/694967/ The recent high-throughput sequencing has enabled the composition of Escherichia coli strains in the human microbial community to be profiled en masse. However, there are two challenges to address: (1) exploring the genetic differences between E. coli strains in human gut and (2) dynamic responses of E. coli to diverse stress conditions. As a result, we investigated the E. coli strains in human gut microbiome using deep sequencing data and reconstructed genome-wide metabolic networks for the three most common E. coli strains, including E. coli HS, UTI89, and CFT073. The metabolic models show obvious strain-specific characteristics, both in network contents and in behaviors. We predicted optimal biomass production for three models on four different carbon sources (acetate, ethanol, glucose, and succinate) and found that these stress-associated genes were involved in host-microbial interactions and increased in human obesity. Besides, it shows that the growth rates are similar among the models, but the flux distributions are different, even in E. coli core reactions. The correlations between human diabetes-associated metabolic reactions in the E. coli models were also predicted. The study provides a systems perspective on E. coli strains in human gut microbiome and will be helpful in integrating diverse data sources in the following study. Yue-Dong Gao, Yuqi Zhao, and Jingfei Huang Copyright © 2014 Yue-Dong Gao et al. All rights reserved. Integrated Analysis Identifies Interaction Patterns between Small Molecules and Pathways Sun, 13 Jul 2014 10:50:55 +0000 http://www.hindawi.com/journals/bmri/2014/931825/ Previous studies have indicated that the downstream proteins in a key pathway can be potential drug targets and that the pathway can play an important role in the action of drugs. So pathways could be considered as targets of small molecules. A link map between small molecules and pathways was constructed using gene expression profile, pathways, and gene expression of cancer cell line intervened by small molecules and then we analysed the topological characteristics of the link map. Three link patterns were identified based on different drug discovery implications for breast, liver, and lung cancer. Furthermore, molecules that significantly targeted the same pathways tended to treat the same diseases. These results can provide a valuable reference for identifying drug candidates and targets in molecularly targeted therapy. Yan Li, Weiguo Li, Xin Chen, Hong Jiang, Jiatong Sun, Huan Chen, and Sali Lv Copyright © 2014 Yan Li et al. All rights reserved. Effect of Duplicate Genes on Mouse Genetic Robustness: An Update Thu, 10 Jul 2014 11:40:35 +0000 http://www.hindawi.com/journals/bmri/2014/758672/ In contrast to S. cerevisiae and C. elegans, analyses based on the current knockout (KO) mouse phenotypes led to the conclusion that duplicate genes had almost no role in mouse genetic robustness. It has been suggested that the bias of mouse KO database toward ancient duplicates may possibly cause this knockout duplicate puzzle, that is, a very similar proportion of essential genes () between duplicate genes and singletons. In this paper, we conducted an extensive and careful analysis for the mouse KO phenotype data and corroborated a strong effect of duplicate genes on mouse genetics robustness. Moreover, the effect of duplicate genes on mouse genetic robustness is duplication-age dependent, which holds after ruling out the potential confounding effect from coding-sequence conservation, protein-protein connectivity, functional bias, or the bias of duplicates generated by whole genome duplication (WGD). Our findings suggest that two factors, the sampling bias toward ancient duplicates and very ancient duplicates with a proportion of essential genes higher than that of singletons, have caused the mouse knockout duplicate puzzle; meanwhile, the effect of genetic buffering may be correlated with sequence conservation as well as protein-protein interactivity. Zhixi Su, Junqiang Wang, and Xun Gu Copyright © 2014 Zhixi Su et al. All rights reserved. Designing Peptide-Based HIV Vaccine for Chinese Sun, 06 Jul 2014 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2014/272950/ CD4+ T cells are central to the induction and maintenance of CD8+ T cell and antibody-producing B cell responses, and the latter are essential for the protection against disease in subjects with HIV infection. How to elicit HIV-specific CD4+ T cell responses in a given population using vaccines is one of the major areas of current HIV vaccine research. To design vaccine that targets specifically Chinese, we assembled a database that is comprised of sequences from 821 Chinese HIV isolates and 46 human leukocyte antigen (HLA) DR alleles identified in Chinese population. We then predicted 20 potential HIV epitopes using bioinformatics approaches. The combination of these 20 epitopes has a theoretical coverage of 98.1% of the population for both the prevalent HIV genotypes and also Chinese HLA-DR types. We suggest that testing this vaccine experimentally will facilitate the development of a CD4+ T cell vaccine especially catered for Chinese. Jiayi Shu, Xiaojuan Fan, Jie Ping, Xia Jin, and Pei Hao Copyright © 2014 Jiayi Shu et al. All rights reserved. Preliminary Characterization of Mitochondrial Genome of Melipona scutellaris, a Brazilian Stingless Bee Mon, 16 Jun 2014 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2014/927546/ Bees are manufacturers of relevant economical products and have a pollinator role fundamental to ecosystems. Traditionally, studies focused on the genus Melipona have been mostly based on behavioral, and social organization and ecological aspects. Only recently the evolutionary history of this genus has been assessed using molecular markers, including mitochondrial genes. Even though these studies have shed light on the evolutionary history of the Melipona genus, a more accurate picture may emerge when full nuclear and mitochondrial genomes of Melipona species become available. Here we present the assembly, annotation, and characterization of a draft mitochondrial genome of the Brazilian stingless bee Melipona scutellaris using Melipona bicolor as a reference organism. Using Illumina MiSeq data, we achieved the annotation of all protein coding genes, as well as the genes for the two ribosomal subunits (16S and 12S) and transfer RNA genes as well. Using the COI sequence as a DNA barcode, we found that M. cramptoni is the closest species to M. scutellaris. Manuella Souza Silverio, Vinícius de Rezende Rodovalho, Ana Maria Bonetti, Guilherme Corrêa de Oliveira, Sara Cuadros-Orellana, Carlos Ueira-Vieira, and Anderson Rodrigues dos Santos Copyright © 2014 Manuella Souza Silverio et al. All rights reserved. RNA-Seq Identifies Key Reproductive Gene Expression Alterations in Response to Cadmium Exposure Tue, 27 May 2014 05:56:06 +0000 http://www.hindawi.com/journals/bmri/2014/529271/ Cadmium is a common toxicant that is detrimental to many tissues. Although a number of transcriptional signatures have been revealed in different tissues after cadmium treatment, the genes involved in the cadmium caused male reproductive toxicity, and the underlying molecular mechanism remains unclear. Here we observed that the mice treated with different amount of cadmium in their rodent chow for six months exhibited reduced serum testosterone. We then performed RNA-seq to comprehensively investigate the mice testicular transcriptome to further elucidate the mechanism. Our results showed that hundreds of genes expression altered significantly in response to cadmium treatment. In particular, we found several transcriptional signatures closely related to the biological processes of regulation of hormone, gamete generation, and sexual reproduction, respectively. The expression of several testosterone synthetic key enzyme genes, such as Star, Cyp11a1, and Cyp17a1, were inhibited by the cadmium exposure. For better understanding of the cadmium-mediated transcriptional regulatory mechanism of the genes, we computationally analyzed the transcription factors binding sites and the mircoRNAs targets of the differentially expressed genes. Our findings suggest that the reproductive toxicity by cadmium exposure is implicated in multiple layers of deregulation of several biological processes and transcriptional regulation in mice. Hanyang Hu, Xing Lu, Xiang Cen, Xiaohua Chen, Feng Li, and Shan Zhong Copyright © 2014 Hanyang Hu et al. All rights reserved. Comprehensive Transcriptome Study to Develop Molecular Resources of the Copepod Calanus sinicus for Their Potential Ecological Applications Tue, 20 May 2014 09:35:05 +0000 http://www.hindawi.com/journals/bmri/2014/493825/ Calanus sinicus Brodsky (Copepoda, Crustacea) is a dominant zooplanktonic species widely distributed in the margin seas of the Northwest Pacific Ocean. In this study, we utilized an RNA-Seq-based approach to develop molecular resources for C. sinicus. Adult samples were sequenced using the Illumina HiSeq 2000 platform. The sequencing data generated 69,751 contigs from 58.9 million filtered reads. The assembled contigs had an average length of 928.8 bp. Gene annotation allowed the identification of 43,417 unigene hits against the NCBI database. Gene ontology (GO) and KEGG pathway mapping analysis revealed various functional genes related to diverse biological functions and processes. Transcripts potentially involved in stress response and lipid metabolism were identified among these genes. Furthermore, 4,871 microsatellites and 110,137 single nucleotide polymorphisms (SNPs) were identified in the C. sinicus transcriptome sequences. SNP validation by the melting temperature ()-shift method suggested that 16 primer pairs amplified target products and showed biallelic polymorphism among 30 individuals. The present work demonstrates the power of Illumina-based RNA-Seq for the rapid development of molecular resources in nonmodel species. The validated SNP set from our study is currently being utilized in an ongoing ecological analysis to support a future study of C. sinicus population genetics. Qing Yang, Fanyue Sun, Zhi Yang, and Hongjun Li Copyright © 2014 Qing Yang et al. All rights reserved. Stratification of Gene Coexpression Patterns and GO Function Mining for a RNA-Seq Data Series Mon, 19 May 2014 10:58:44 +0000 http://www.hindawi.com/journals/bmri/2014/969768/ RNA-Seq is emerging as an increasingly important tool in biological research, and it provides the most direct evidence of the relationship between the physiological state and molecular changes in cells. A large amount of RNA-Seq data across diverse experimental conditions have been generated and deposited in public databases. However, most developed approaches for coexpression analyses focus on the coexpression pattern mining of the transcriptome, thereby ignoring the magnitude of gene differences in one pattern. Furthermore, the functional relationships of genes in one pattern, and notably among patterns, were not always recognized. In this study, we developed an integrated strategy to identify differential coexpression patterns of genes and probed the functional mechanisms of the modules. Two real datasets were used to validate the method and allow comparisons with other methods. One of the datasets was selected to illustrate the flow of a typical analysis. In summary, we present an approach to robustly detect coexpression patterns in transcriptomes and to stratify patterns according to their relative differences. Furthermore, a global relationship between patterns and biological functions was constructed. In addition, a freely accessible web toolkit “coexpression pattern mining and GO functional analysis” (COGO) was developed. Hui Zhao, Fenglin Cao, Yonghui Gong, Huafeng Xu, Yiping Fei, Longyue Wu, Xiangmei Ye, Dongguang Yang, Xiuhua Liu, Xia Li, and Jin Zhou Copyright © 2014 Hui Zhao et al. All rights reserved. BLAT-Based Comparative Analysis for Transposable Elements: BLATCAT Sun, 18 May 2014 11:08:45 +0000 http://www.hindawi.com/journals/bmri/2014/730814/ The availability of several whole genome sequences makes comparative analyses possible. In primate genomes, the priority of transposable elements (TEs) is significantly increased because they account for ~45% of the primate genomes, they can regulate the gene expression level, and they are associated with genomic fluidity in their host genomes. Here, we developed the BLAST-like alignment tool (BLAT) based comparative analysis for transposable elements (BLATCAT) program. The BLATCAT program can compare specific regions of six representative primate genome sequences (human, chimpanzee, gorilla, orangutan, gibbon, and rhesus macaque) on the basis of BLAT and simultaneously carry out RepeatMasker and/or Censor functions, which are widely used Windows-based web-server functions to detect TEs. All results can be stored as a HTML file for manual inspection of a specific locus. BLATCAT will be very convenient and efficient for comparative analyses of TEs in various primate genomes. Sangbum Lee, Sumin Oh, Keunsoo Kang, and Kyudong Han Copyright © 2014 Sangbum Lee et al. All rights reserved. A Systematic Analysis of miRNA-mRNA Paired Variations Reveals Widespread miRNA Misregulation in Breast Cancer Sun, 18 May 2014 08:59:25 +0000 http://www.hindawi.com/journals/bmri/2014/291280/ MicroRNAs (miRNAs) are a class of small noncoding RNAs that can regulate gene expression by binding to target mRNAs and induce translation repression or RNA degradation. There have been many studies indicating that both miRNAs and mRNAs display aberrant expression in breast cancer. Previously, most researches into the molecular mechanism of breast cancer examined miRNA expression patterns and mRNA expression patterns separately. In this study, we systematically analysed miRNA-mRNA paired variations (MMPVs), which are miRNA-mRNA pairs whose pattern of regulation can vary in association with biopathological features, such as the oestrogen receptor (ER), TP53 and human epidermal growth factor receptor 2 (HER2) genes, survival time, and breast cancer subtypes. We demonstrated that the existence of MMPVs is general and widespread but that there is a general unbalance in the distribution of MMPVs among the different biopathological features. Furthermore, based on studying MMPVs that are related to multiple biopathological features, we propose a potential crosstalk mechanism between ER and HER2. Lei Zhong, Kuixi Zhu, Nana Jin, Deng Wu, Jianguo Zhang, Baoliang Guo, Zhaoqi Yan, and Qingyuan Zhang Copyright © 2014 Lei Zhong et al. All rights reserved. O18Quant: A Semiautomatic Strategy for Quantitative Analysis of High-Resolution 16O/18O Labeled Data Sun, 11 May 2014 07:37:17 +0000 http://www.hindawi.com/journals/bmri/2014/971857/ Proteolytic 18O-labeling has been widely used in quantitative proteomics since it can uniformly label all peptides from different kinds of proteins. There have been multiple algorithms and tools developed over the last few years to analyze high-resolution proteolytic 16O/18O labeled mass spectra. We have developed a software package, O18Quant, which addresses two major issues in the previously developed algorithms. First, O18Quant uses a robust linear model (RLM) for peptide-to-protein ratio estimation. RLM can minimize the effect of outliers instead of iteratively removing them which is a common practice in other approaches. Second, the existing algorithms lack applicable implementation. We address this by implementing O18Quant using C# under Microsoft.net framework and R. O18Quant automatically calculates the peptide/protein relative ratio and provides a friendly graphical user interface (GUI) which allows the user to manually validate the quantification results at scan, peptide, and protein levels. The intuitive GUI of O18Quant can greatly enhance the user’s visualization and understanding of the data analysis. O18Quant can be downloaded for free as part of the software suite ProteomicsTools. Yan Guo, Masaru Miyagi, Rong Zeng, and Quanhu Sheng Copyright © 2014 Yan Guo et al. All rights reserved. Construction and Characterization of a Bacterial Artificial Chromosome Library for the Hexaploid Wheat Line 92R137 Mon, 05 May 2014 14:07:20 +0000 http://www.hindawi.com/journals/bmri/2014/845806/ For map-based cloning of genes conferring important traits in the hexaploid wheat line 92R137, a bacterial artificial chromosome (BAC) library, including two sublibraries, was constructed using the genomic DNA of 92R137 digested with restriction enzymes HindIII and BamHI. The BAC library was composed of total 765,696 clones, of which 390,144 were from the HindIII digestion and 375,552 from the BamHI digestion. Through pulsed-field gel electrophoresis (PFGE) analysis of 453 clones randomly selected from the HindIII sublibrary and 573 clones from the BamHI sublibrary, the average insert sizes were estimated as 129 and 113 kb, respectively. Thus, the HindIII sublibrary was estimated to have a 3.01-fold coverage and the BamHI sublibrary a 2.53-fold coverage based on the estimated hexaploid wheat genome size of 16,700 Mb. The 765,696 clones were arrayed in 1,994 384-well plates. All clones were also arranged into plate pools and further arranged into 5-dimensional (5D) pools. The probability of identifying a clone corresponding to any wheat DNA sequence (such as gene Yr26 for stripe rust resistance) from the library was estimated to be more than 99.6%. Through polymerase chain reaction screening the 5D pools with Xwe173, a marker tightly linked to Yr26, six BAC clones were successfully obtained. These results demonstrate that the BAC library is a valuable genomic resource for positional cloning of Yr26 and other genes of interest. Qingdong Zeng, Fengping Yuan, Xin Xu, Xue Shi, Xiaojun Nie, Hua Zhuang, Xianming Chen, Zhonghua Wang, Xiaojie Wang, Lili Huang, Dejun Han, and Zhensheng Kang Copyright © 2014 Qingdong Zeng et al. All rights reserved. Antioxidant Defense Enzyme Genes and Asthma Susceptibility: Gender-Specific Effects and Heterogeneity in Gene-Gene Interactions between Pathogenetic Variants of the Disease Mon, 05 May 2014 09:38:59 +0000 http://www.hindawi.com/journals/bmri/2014/708903/ Oxidative stress resulting from an increased amount of reactive oxygen species and an imbalance between oxidants and antioxidants plays an important role in the pathogenesis of asthma. The present study tested the hypothesis that genetic susceptibility to allergic and nonallergic variants of asthma is determined by complex interactions between genes encoding antioxidant defense enzymes (ADE). We carried out a comprehensive analysis of the associations between adult asthma and 46 single nucleotide polymorphisms of 34 ADE genes and 12 other candidate genes of asthma in Russian population using set association analysis and multifactor dimensionality reduction approaches. We found for the first time epistatic interactions between ADE genes underlying asthma susceptibility and the genetic heterogeneity between allergic and nonallergic variants of the disease. We identified GSR (glutathione reductase) and PON2 (paraoxonase 2) as novel candidate genes for asthma susceptibility. We observed gender-specific effects of ADE genes on the risk of asthma. The results of the study demonstrate complexity and diversity of interactions between genes involved in oxidative stress underlying susceptibility to allergic and nonallergic asthma. Alexey V. Polonikov, Vladimir P. Ivanov, Alexey D. Bogomazov, Maxim B. Freidin, Thomas Illig, and Maria A. Solodilova Copyright © 2014 Alexey V. Polonikov et al. All rights reserved. RNA Sequencing Reveals Upregulation of RUNX1-RUNX1T1 Gene Signatures in Clear Cell Renal Cell Carcinoma Tue, 25 Mar 2014 11:39:40 +0000 http://www.hindawi.com/journals/bmri/2014/450621/ In the past few years, therapies targeted at the von Hippel-Lindau (VHL) and hypoxia-inducible factor (HIF) pathways, such as sunitinib and sorafenib, have been developed to treat clear cell renal cell carcinoma (ccRCC). However, the majority of patients will eventually show resistance to antiangiogenesis therapies. The purpose of our study was to identify novel pathways that could be potentially used as targets for new therapies. Whole transcriptome sequencing (RNA-Seq) was conducted on eight matched tumor and adjacent normal tissue samples. A novel RUNX1-RUNX1T1 pathway was identified which was upregulated in ccRCC through gene set enrichment analysis (GSEA). We also confirmed the findings based on previously published gene expression microarray data. Our data shows that upregulated of the RUNX1-RUNX1T1 gene set maybe an important factor contributing to the etiology of ccRCC. Zuquan Xiong, Hongjie Yu, Yan Ding, Chenchen Feng, Hanming Wei, Sha Tao, Dan Huang, Siqun Lilly Zheng, Jielin Sun, Jianfeng Xu, and Zujun Fang Copyright © 2014 Zuquan Xiong et al. All rights reserved. Analysis of Differentially Expressed Genes and Signaling Pathways Related to Intramuscular Fat Deposition in Skeletal Muscle of Sex-Linked Dwarf Chickens Mon, 17 Mar 2014 08:54:49 +0000 http://www.hindawi.com/journals/bmri/2014/724274/ Intramuscular fat (IMF) plays an important role in meat quality. However, the molecular mechanisms underlying IMF deposition in skeletal muscle have not been addressed for the sex-linked dwarf (SLD) chicken. In this study, potential candidate genes and signaling pathways related to IMF deposition in chicken leg muscle tissue were characterized using gene expression profiling of both 7-week-old SLD and normal chickens. A total of 173 differentially expressed genes (DEGs) were identified between the two breeds. Subsequently, 6 DEGs related to lipid metabolism or muscle development were verified in each breed based on gene ontology (GO) analysis. In addition, KEGG pathway analysis of DEGs indicated that some of them (GHR, SOCS3, and IGF2BP3) participate in adipocytokine and insulin signaling pathways. To investigate the role of the above signaling pathways in IMF deposition, the gene expression of pathway factors and other downstream genes were measured by using qRT-PCR and Western blot analyses. Collectively, the results identified potential candidate genes related to IMF deposition and suggested that IMF deposition in skeletal muscle of SLD chicken is regulated partially by pathways of adipocytokine and insulin and other downstream signaling pathways (TGF-β/SMAD3 and Wnt/catenin-β pathway). Yaqiong Ye, Shumao Lin, Heping Mu, Xiaohong Tang, Yangdan Ou, Jian Chen, Yongjiang Ma, and Yugu Li Copyright © 2014 Yaqiong Ye et al. All rights reserved. Phylogeny, Functional Annotation, and Protein Interaction Network Analyses of the Xenopus tropicalis Basic Helix-Loop-Helix Transcription Factors Sun, 10 Nov 2013 13:36:34 +0000 http://www.hindawi.com/journals/bmri/2013/145037/ The previous survey identified 70 basic helix-loop-helix (bHLH) proteins, but it was proved to be incomplete, and the functional information and regulatory networks of frog bHLH transcription factors were not fully known. Therefore, we conducted an updated genome-wide survey in the Xenopus tropicalis genome project databases and identified 105 bHLH sequences. Among the retrieved 105 sequences, phylogenetic analyses revealed that 103 bHLH proteins belonged to 43 families or subfamilies with 46, 26, 11, 3, 15, and 4 members in the corresponding supergroups. Next, gene ontology (GO) enrichment analyses showed 65 significant GO annotations of biological processes and molecular functions and KEGG pathways counted in frequency. To explore the functional pathways, regulatory gene networks, and/or related gene groups coding for Xenopus tropicalis bHLH proteins, the identified bHLH genes were put into the databases KOBAS and STRING to get the signaling information of pathways and protein interaction networks according to available public databases and known protein interactions. From the genome annotation and pathway analysis using KOBAS, we identified 16 pathways in the Xenopus tropicalis genome. From the STRING interaction analysis, 68 hub proteins were identified, and many hub proteins created a tight network or a functional module within the protein families. Wuyi Liu and Deyu Chen Copyright © 2013 Wuyi Liu and Deyu Chen. All rights reserved. A Genome-Wide Analysis of RNA Pseudoknots That Stimulate Efficient −1 Ribosomal Frameshifting or Readthrough in Animal Viruses Mon, 04 Nov 2013 09:46:30 +0000 http://www.hindawi.com/journals/bmri/2013/984028/ Programmed −1 ribosomal frameshifting (PRF) and stop codon readthrough are two translational recoding mechanisms utilized by some RNA viruses to express their structural and enzymatic proteins at a defined ratio. Efficient recoding usually requires an RNA pseudoknot located several nucleotides downstream from the recoding site. To assess the strategic importance of the recoding pseudoknots, we have carried out a large scale genome-wide analysis in which we used an in-house developed program to detect all possible H-type pseudoknots within the genomic mRNAs of 81 animal viruses. Pseudoknots are detected downstream from ~85% of the recoding sites, including many previously unknown pseudoknots. ~78% of the recoding pseudoknots are the most stable pseudoknot within the viral genomes. However, they are not as strong as some designed pseudoknots that exhibit roadblocking effect on the translating ribosome. Strong roadblocking pseudoknots are not detected within the viral genomes. These results indicate that the decoding pseudoknots have evolved to possess optimal stability for efficient recoding. We also found that the sequence at the gag-pol frameshift junction of HIV1 harbors potential elaborated pseudoknots encompassing the frameshift site. A novel mechanism is proposed for possible involvement of the elaborated pseudoknots in the HIV1 PRF event. Xiaolan Huang, Qiang Cheng, and Zhihua Du Copyright © 2013 Xiaolan Huang et al. All rights reserved. Ovarian and Breast Cancer Spheres Are Similar in Transcriptomic Features and Sensitive to Fenretinide Tue, 08 Oct 2013 11:25:56 +0000 http://www.hindawi.com/journals/bmri/2013/510905/ Cancer stem cells (CSCs) are resistant to chemotherapy and are ability to regenerate cancer cell populations, thus attracting much attention in cancer research. In this report, we first demonstrated that sphere cells from ovarian cancer cell line A2780 shared many features of CSCs, such as resistance to cisplatin and able to initiate tumors in an efficient manner. Then, we conducted cDNA microarray analysis on spheres from ovarian A2780 cells, and from breast MCF7 and SUM159 cells, and found that molecular pathways underlying spheres from these cancer cell lines were similar to a large extent, suggesting that similar mechanisms are involved in the genesis of CSCs in both ovarian and breast cancer types. In addition, we showed that spheres from these cancer types were highly sensitive to fenretinide, a stimulus of oxidative stress-mediated apoptosis in cancer cells. Thus, our results not only provide important insights into mechanisms underlying CSCs in ovarian and breast cancer, but also lead to the development of more sophisticated protocols of cancer therapy in near future. Haiwei Wang, Yuxing Zhang, and Yanzhi Du Copyright © 2013 Haiwei Wang et al. All rights reserved. A Comparison of Synonymous Codon Usage Bias Patterns in DNA and RNA Virus Genomes: Quantifying the Relative Importance of Mutational Pressure and Natural Selection Wed, 02 Oct 2013 14:20:52 +0000 http://www.hindawi.com/journals/bmri/2013/406342/ Codon usage bias patterns have been broadly explored for many viruses. However, the relative importance of mutation pressure and natural selection is still under debate. In the present study, I tried to resolve controversial issues on determining the principal factors of codon usage patterns for DNA and RNA viruses, respectively, by examining over 38000 ORFs. By utilizing variation partitioning technique, the results showed that 27% and 21% of total variation could be attributed to mutational pressure, while 5% and 6% of total variation could be explained by natural selection for DNA and RNA viruses, respectively, in codon usage patterns. Furthermore, the combined effect of mutational pressure and natural selection on influencing codon usage patterns of viruses is substantial (explaining 10% and 8% of total variation of codon usage patterns). With respect to GC variation, GC content is always negatively and significantly correlated with aromaticity. Interestingly, the signs for the significant correlations between GC, gene lengths, and hydrophobicity are completely opposite between DNA and RNA viruses, being positive for DNA viruses while being negative for RNA viruses. At last, GC12 versus G3s plot suggests that natural selection is more important than mutational pressure on influencing the GC content in the first and second codon positions. Youhua Chen Copyright © 2013 Youhua Chen. All rights reserved. Genome Instability at Common Fragile Sites: Searching for the Cause of Their Instability Thu, 05 Sep 2013 10:10:34 +0000 http://www.hindawi.com/journals/bmri/2013/730714/ Common fragile sites (CFS) are heritable nonrandomly distributed loci on human chromosomes that exhibit an increased frequency of chromosomal breakage under conditions of replication stress. They are considered the preferential targets for high genomic instability from the earliest stages of human cancer development, and increased chromosome instability at these loci has been observed following replication stress in a subset of human genetic diseases. Despite their biological and medical relevance, the molecular basis of CFS fragility in vivo has not been fully elucidated. At present, different models have been proposed to explain how instability at CFS arises and multiple factors seem to contribute to their instability. However, all these models involve DNA replication and suggest that replication fork stalling along CFS during DNA synthesis is a very frequent event. Consistent with this, the maintenance of CFS stability relies on the ATR-dependent checkpoint, together with a number of proteins promoting the recovery of stalled replication forks. In this review, we discuss mainly the possible causes that threaten the integrity of CFS in the light of new findings, paying particular attention to the role of the S-phase checkpoint. Annapaola Franchitto Copyright © 2013 Annapaola Franchitto. All rights reserved. Development of Pineapple Microsatellite Markers and Germplasm Genetic Diversity Analysis Mon, 19 Aug 2013 13:28:08 +0000 http://www.hindawi.com/journals/bmri/2013/317912/ Two methods were used to develop pineapple microsatellite markers. Genomic library-based SSR development: using selectively amplified microsatellite assay, 86 sequences were generated from pineapple genomic library. 91 (96.8%) of the 94 Simple Sequence Repeat (SSR) loci were dinucleotide repeats (39 AC/GT repeats and 52 GA/TC repeats, accounting for 42.9% and 57.1%, resp.), and the other three were mononucleotide repeats. Thirty-six pairs of SSR primers were designed; 24 of them generated clear bands of expected sizes, and 13 of them showed polymorphism. EST-based SSR development: 5659 pineapple EST sequences obtained from NCBI were analyzed; among 1397 nonredundant EST sequences, 843 were found containing 1110 SSR loci (217 of them contained more than one SSR locus). Frequency of SSRs in pineapple EST sequences is 1SSR/3.73 kb, and 44 types were found. Mononucleotide, dinucleotide, and trinucleotide repeats dominate, accounting for 95.6% in total. AG/CT and AGC/GCT were the dominant type of dinucleotide and trinucleotide repeats, accounting for 83.5% and 24.1%, respectively. Thirty pairs of primers were designed for each of randomly selected 30 sequences; 26 of them generated clear and reproducible bands, and 22 of them showed polymorphism. Eighteen pairs of primers obtained by the one or the other of the two methods above that showed polymorphism were selected to carry out germplasm genetic diversity analysis for 48 breeds of pineapple; similarity coefficients of these breeds were between 0.59 and 1.00, and they can be divided into four groups accordingly. Amplification products of five SSR markers were extracted and sequenced, corresponding repeat loci were found and locus mutations are mainly in copy number of repeats and base mutations in the flanking region. Suping Feng, Helin Tong, You Chen, Jingyi Wang, Yeyuan Chen, Guangming Sun, Junhu He, and Yaoting Wu Copyright © 2013 Suping Feng et al. All rights reserved. Novel One-Step Multiplex PCR-Based Method for HLA Typing and Preimplantational Genetic Diagnosis of -Thalassemia Thu, 04 Apr 2013 14:54:47 +0000 http://www.hindawi.com/journals/bmri/2013/585106/ Preimplantation genetic diagnosis (PGD) of single gene disorders, combined with HLA matching (PGD-HLA), has emerged as a tool for couples at risk of transmitting a genetic disease to select unaffected embryos of an HLA tissue type compatible with that of an existing affected child. Here, we present a novel one-step multiplex PCR to genotype a spectrum of STRs to simultaneously perform HLA typing and PGD for -thalassemia. This method is being routinely used for PGD-HLA cycles in our department, with a genotyping success rate of 100%. As an example, we present the first successful PGD-HLA typing in Spain, which resulted in the birth of a boy and subsequent successful HSC transplantation to his affected brother, who is doing well 4 years following transplantation. The advantage of our method is that it involves only a round of single PCR for multiple markers amplification (up to 10 markers within the HLA and 6 markers at the -globin loci). This strategy has allowed us to considerably reduce the optimization of the PCR method for each specific PGD-HLA family as well as the time to obtain molecular results in each cycle. Raquel M. Fernández, Ana Peciña, Maria Dolores Lozano-Arana, Juan Carlos García-Lozano, Salud Borrego, and Guillermo Antiñolo Copyright © 2013 Raquel M. Fernández et al. All rights reserved. Genotyping of CYP2C9 and VKORC1 in the Arabic Population of Al-Ahsa, Saudi Arabia Sun, 17 Mar 2013 14:18:07 +0000 http://www.hindawi.com/journals/bmri/2013/315980/ Polymorphisms in the genes encoding CYP2C9 enzyme and VKORC1 reductase significantly influence the dose variability of coumarinic oral anticoagulants (COAs). Substantial inter- and intraethnic variability exists in the frequencies of CYP2C9∗2 and ∗3 and VKORC1 –1639A alleles. However, the prevalence of CYP2C9 and VKORC1 genetic variants is less characterized in Arab populations. A total of 131 healthy adult subjects from the Al-Ahsa region of Saudi Arabia were genotyped for the CYP2C9∗2 and ∗3 and VKORC1 –1639G>A polymorphisms by PCR-RFLP method. The frequencies of the CYP2C9∗2 and ∗3 and VKORC1 –1639A alleles were 13.3%, 2.3%, and 42.4%, respectively, with no subjects carrying 2 defective alleles. The frequencies of the CYP2C9∗3 and VKORC1 –1639A alleles were significantly lower than those reported in different Arabian populations. None of the subjects with the VKORC1 –1639AA genotype were carriers of CYP2C9∗1/∗3 genotypes that lead to sensitivity to COAs therapy. The low frequency of the CYP2C9∗3 allele combined with the absence of subjects carrying 2 defective CYP2C9 alleles suggests that, in this specific population, pharmacogenetic COAs dosing may mostly rely upon VKORC1 genotyping. Abdullah M. Alzahrani, Georgia Ragia, Hamza Hanieh, and Vangelis G. Manolopoulos Copyright © 2013 Abdullah M. Alzahrani et al. All rights reserved. In Silico Expressed Sequence Tag Analysis in Identification of Probable Diabetic Genes as Virtual Therapeutic Targets Mon, 11 Feb 2013 08:32:26 +0000 http://www.hindawi.com/journals/bmri/2013/704818/ The expressed sequence tags (ESTs) are major entities for gene discovery, molecular transcripts, and single nucleotide polymorphism (SNPs) analysis as well as functional annotation of putative gene products. In our quest for identification of novel diabetic genes as virtual targets for type II diabetes, we searched various publicly available databases and found 7 reported genes. The in silico EST analysis of these reported genes produced 6 consensus contigs which illustrated some good matches to a number of chromosomes of the human genome. Again the conceptual translation of these contigs produced 3 protein sequences. The functional and structural annotations of these proteins revealed some important features which may lead to the discovery of novel therapeutic targets for the treatment of diabetes. Pabitra Mohan Behera, Deepak Kumar Behera, Aparajeya Panda, Anshuman Dixit, and Payodhar Padhi Copyright © 2013 Pabitra Mohan Behera et al. All rights reserved. The Short ITS2 Sequence Serves as an Efficient Taxonomic Sequence Tag in Comparison with the Full-Length ITS Thu, 17 Jan 2013 18:07:39 +0000 http://www.hindawi.com/journals/bmri/2013/741476/ An ideal DNA barcoding region should be short enough to be amplified from degraded DNA. In this paper, we discuss the possibility of using a short nuclear DNA sequence as a barcode to identify a wide range of medicinal plant species. First, the PCR and sequencing success rates of ITS and ITS2 were evaluated based entirely on materials from dry medicinal product and herbarium voucher specimens, including some samples collected back to 90 years ago. The results showed that ITS2 could recover 91% while ITS could recover only 23% efficiency of PCR and sequencing by using one pair of primer. Second, 12861 ITS and ITS2 plant sequences were used to compare the identification efficiency of the two regions. Four identification criteria (BLAST, inter- and intradivergence Wilcoxon signed rank tests, and TaxonDNA) were evaluated. Our results supported the hypothesis that ITS2 can be used as a minibarcode to effectively identify species in a wide variety of specimens and medicinal materials. Jianping Han, Yingjie Zhu, Xiaochen Chen, Baoshen Liao, Hui Yao, Jingyuan Song, Shilin Chen, and Fanyun Meng Copyright © 2013 Jianping Han et al. All rights reserved. Patterns of Synonymous Codon Usage on Human Metapneumovirus and Its Influencing Factors Mon, 05 Nov 2012 14:24:19 +0000 http://www.hindawi.com/journals/bmri/2012/460837/ Human metapneumovirus (HMPV) is an important agent of acute respiratory tract infection in children, while its pathogenicity and molecular evolution are lacking. Herein, we firstly report the synonymous codon usage patterns of HMPV genome. The relative synonymous codon usage (RSCU) values, effective number of codon (ENC) values, nucleotide contents, and correlation analysis were performed among 17 available whole genome of HMPV, including different genotypes. All preferred codons in HMPV are ended with A/U nucleotide and exhibited a great association with its high proportion of these two nucleotides in their genomes. Mutation pressure rather than natural selection is the main influence factor that determines the bias of synonymous codon usage in HMPV. The complementary pattern of codon usage bias between HMPV and human cell was observed, and this phenomenon suggests that host cells might be also act as an important factor to affect the codon usage bias. Moreover, the codon usage biases in each HMPV genotypes are separated into different clades, which suggest that phylogenetic distance might involve in codon usage bias formation as well. These analyses of synonymous codon usage bias in HMPV provide more information for better understanding its evolution and pathogenicity. Qiao Zhong, Weidong Xu, Yuanjian Wu, and Hongxing Xu Copyright © 2012 Qiao Zhong et al. All rights reserved. Technical Considerations for Reduced Representation Bisulfite Sequencing with Multiplexed Libraries Wed, 10 Oct 2012 14:09:43 +0000 http://www.hindawi.com/journals/bmri/2012/741542/ Reduced representation bisulfite sequencing (RRBS), which couples bisulfite conversion and next generation sequencing, is an innovative method that specifically enriches genomic regions with a high density of potential methylation sites and enables investigation of DNA methylation at single-nucleotide resolution. Recent advances in the Illumina DNA sample preparation protocol and sequencing technology have vastly improved sequencing throughput capacity. Although the new Illumina technology is now widely used, the unique challenges associated with multiplexed RRBS libraries on this platform have not been previously described. We have made modifications to the RRBS library preparation protocol to sequence multiplexed libraries on a single flow cell lane of the Illumina HiSeq 2000. Furthermore, our analysis incorporates a bioinformatics pipeline specifically designed to process bisulfite-converted sequencing reads and evaluate the output and quality of the sequencing data generated from the multiplexed libraries. We obtained an average of 42 million paired-end reads per sample for each flow-cell lane, with a high unique mapping efficiency to the reference human genome. Here we provide a roadmap of modifications, strategies, and trouble shooting approaches we implemented to optimize sequencing of multiplexed libraries on an a RRBS background. Aniruddha Chatterjee, Euan J. Rodger, Peter A. Stockwell, Robert J. Weeks, and Ian M. Morison Copyright © 2012 Aniruddha Chatterjee et al. All rights reserved. Lim Mineralization Protein 3 Induces the Osteogenic Differentiation of Human Amniotic Fluid Stromal Cells through Kruppel-Like Factor-4 Downregulation and Further Bone-Specific Gene Expression Tue, 02 Oct 2012 16:20:20 +0000 http://www.hindawi.com/journals/bmri/2012/813894/ Multipotent mesenchymal stem cells with extensive self-renewal properties can be easily isolated and rapidly expanded in culture from small volumes of amniotic fluid. These cells, namely, amniotic fluid-stromal cells (AFSCs), can be regarded as an attractive source for tissue engineering purposes, being phenotypically and genetically stable, plus overcoming all the safety and ethical issues related to the use of embryonic/fetal cells. LMP3 is a novel osteoinductive molecule acting upstream to the main osteogenic pathways. This study is aimed at delineating the basic molecular events underlying LMP3-induced osteogenesis, using AFSCs as a cellular model to focus on the molecular features underlying the multipotency/differentiation switch. For this purpose, AFSCs were isolated and characterized in vitro and transfected with a defective adenoviral vector expressing the human LMP3. LMP3 induced the successful osteogenic differentiation of AFSC by inducing the expression of osteogenic markers and osteospecific transcription factors. Moreover, LMP3 induced an early repression of the kruppel-like factor-4, implicated in MSC stemness maintenance. KLF4 repression was released upon LMP3 silencing, indicating that this event could be reasonably considered among the basic molecular events that govern the proliferation/differentiation switch during LMP3-induced osteogenic differentiation of AFSC. Marta Barba, Filomena Pirozzi, Nathalie Saulnier, Tiziana Vitali, Maria Teresa Natale, Giandomenico Logroscino, Paul D. Robbins, Andrea Gambotto, Giovanni Neri, Fabrizio Michetti, Enrico Pola, and Wanda Lattanzi Copyright © 2012 Marta Barba et al. All rights reserved. Genome-Wide Analysis of mir-548 Gene Family Reveals Evolutionary and Functional Implications Tue, 02 Oct 2012 12:14:30 +0000 http://www.hindawi.com/journals/bmri/2012/679563/ mir-548 is a larger, poorly conserved primate-specific miRNA gene family. 69 human mir-548 genes located in almost all human chromosomes whose widespread distribution pattern implicates the evolutionary origin from transposable elements. Higher level of nucleotide divergence was detected between these human miRNA genes, which mainly derived from divergence of multicopy pre-miRNAs and homologous miRNA genes. Products of  mir-548, miR-548-5p, and miR-548-3p showed inconsistent evolutionary patterns, which partly contributed to larger genetic distances between pre-miRNAs. “Seed shifting” events could be detected among miR-548 sequences due to various 5′ ends. The events led to shift of seed sequences and target mRNAs, even generated to new target mRNAs. Additionally, the phenomenon of miRNA:miRNA interaction in the miRNA gene family was found. The potential interaction between miRNAs may be contributed to dynamic miRNA expression profiles by complementarily binding events to form miRNA:miRNA duplex with 5′-/3′-overhangs. The miRNA gene family had important roles in multiple biological processes, including signaling pathways and some cancers. The potential abundant roles and functional implication further led to the larger and poorly conserved gene family with genetic variation based on transposable elements. The evolutionary pattern of the primate-specific gene family might contribute to dynamic expression profiles and regulatory network. Tingming Liang, Li Guo, and Chang Liu Copyright © 2012 Tingming Liang et al. All rights reserved. Artificial Chromosomes to Explore and to Exploit Biosynthetic Capabilities of Actinomycetes Tue, 07 Aug 2012 14:28:51 +0000 http://www.hindawi.com/journals/bmri/2012/462049/ Actinomycetes are an important source of biologically active compounds, like antibiotics, antitumor agents, and immunosuppressors. Genome sequencing is revealing that this class of microorganisms has larger genomes relative to other bacteria and uses a considerable fraction of its coding capacity (5–10%) for the production of mostly cryptic secondary metabolites. To access actinomycetes biosynthetic capabilities or to improve the pharmacokinetic properties and production yields of these chemically complex compounds, genetic manipulation of the producer strains can be performed. Heterologous expression in amenable hosts can be useful to exploit and to explore the genetic potential of actinomycetes and not cultivable but interesting bacteria. Artificial chromosomes that can be stably integrated into the Streptomyces genome were constructed and demonstrated to be effective for transferring entire biosynthetic gene clusters from intractable actinomycetes into more suitable hosts. In this paper, the construction of several shuttle Escherichia coli-Streptomyces artificial chromosomes is discussed together with old and new strategies applied to improve heterologous production of secondary metabolites. Rosa Alduina and Giuseppe Gallo Copyright © 2012 Rosa Alduina and Giuseppe Gallo. All rights reserved. Distribution of Genes and Repetitive Elements in the Diabrotica virgifera virgifera Genome Estimated Using BAC Sequencing Sun, 05 Aug 2012 09:36:08 +0000 http://www.hindawi.com/journals/bmri/2012/604076/ Feeding damage caused by the western corn rootworm, Diabrotica virgifera virgifera, is destructive to corn plants in North America and Europe where control remains challenging due to evolution of resistance to chemical and transgenic toxins. A BAC library, DvvBAC1, containing 109,486 clones with 104±34.5 kb inserts was created, which has an ~4.56X genome coverage based upon a 2.58 Gb (2.80 pg) flow cytometry-estimated haploid genome size. Paired end sequencing of 1037 BAC inserts produced 1.17 Mb of data (~0.05% genome coverage) and indicated ~9.4 and 16.0% of reads encode, respectively, endogenous genes and transposable elements (TEs). Sequencing genes within BAC full inserts demonstrated that TE densities are high within intergenic and intron regions and contribute to the increased gene size. Comparison of homologous genome regions cloned within different BAC clones indicated that TE movement may cause haplotype variation within the inbred strain. The data presented here indicate that the D. virgifera virgifera genome is large in size and contains a high proportion of repetitive sequence. These BAC sequencing methods that are applicable for characterization of genomes prior to sequencing may likely be valuable resources for genome annotation as well as scaffolding. Brad S. Coates, Analiza P. Alves, Haichuan Wang, Kimberly K. O. Walden, B. Wade French, Nicholas J. Miller, Craig A. Abel, Hugh M. Robertson, Thomas W. Sappington, and Blair D. Siegfried Copyright © 2012 Brad S. Coates et al. All rights reserved. Is BAC Transgenesis Obsolete? State of the Art in the Era of Designer Nucleases Mon, 30 Jul 2012 10:41:31 +0000 http://www.hindawi.com/journals/bmri/2012/308414/ DNA constructs based on bacterial artificial chromosomes (BACs) are frequently used to generate transgenic animals as they reduce the influence of position effects and allow predictable expression patterns for genes whose regulatory sequences are not fully identified. Despite these advantages BAC transgenics suffer from drawbacks such as complicated vector construction, low efficiency of transgenesis, and some remaining expression variegation. The recent development of transcription activator-like effector nucleases (TALENs) and zinc finger nucleases (ZFNs) has resulted in new transgenic techniques which do not have the drawbacks associated with BAC transgenesis. Initial reports indicate that such designer nucleases (DNs) allow the targeted insertion of transgenes into endogenous loci by direct injection of the targeting vector and mRNA/DNA encoding the predesigned nucleases into oocytes. This results in the transgene being inserted at a specific locus in the mouse genome, thus circumventing the drawbacks associated with BAC transgenesis. J. Beil, L. Fairbairn, P. Pelczar, and T. Buch Copyright © 2012 J. Beil et al. All rights reserved. BAC-Dkk3-EGFP Transgenic Mouse: An In Vivo Analytical Tool for Dkk3 Expression Sun, 15 Jul 2012 14:54:18 +0000 http://www.hindawi.com/journals/bmri/2012/973140/ Dickkopf (DKK) family proteins are secreted modulators of the Wnt signaling pathway and are capable of regulating the development of many organs and tissues. We previously identified Dkk3 to be a molecule predominantly expressed in the mouse embryonic retina. However, which cell expresses Dkk3 in the developing and mature mouse retina remains to be elucidated. To examine the precise expression of the Dkk3 protein, we generated BAC-Dkk3-EGFP transgenic mice that express EGFP integrated into the Dkk3 gene in a BAC plasmid. Expression analysis using the BAC-Dkk3-EGFP transgenic mice revealed that Dkk3 is expressed in retinal progenitor cells (RPCs) at embryonic stages and in Müller glial cells in the adult retina. Since Müller glial cells may play a potential role in retinal regeneration, BAC-Dkk3-EGFP mice could be useful for retinal regeneration studies. Yuki Muranishi and Takahisa Furukawa Copyright © 2012 Yuki Muranishi and Takahisa Furukawa. All rights reserved. Comparison of Next-Generation Sequencing Systems Thu, 05 Jul 2012 09:51:42 +0000 http://www.hindawi.com/journals/bmri/2012/251364/ With fast development and wide applications of next-generation sequencing (NGS) technologies, genomic sequence information is within reach to aid the achievement of goals to decode life mysteries, make better crops, detect pathogens, and improve life qualities. NGS systems are typically represented by SOLiD/Ion Torrent PGM from Life Sciences, Genome Analyzer/HiSeq 2000/MiSeq from Illumina, and GS FLX Titanium/GS Junior from Roche. Beijing Genomics Institute (BGI), which possesses the world’s biggest sequencing capacity, has multiple NGS systems including 137 HiSeq 2000, 27 SOLiD, one Ion Torrent PGM, one MiSeq, and one 454 sequencer. We have accumulated extensive experience in sample handling, sequencing, and bioinformatics analysis. In this paper, technologies of these systems are reviewed, and first-hand data from extensive experience is summarized and analyzed to discuss the advantages and specifics associated with each sequencing system. At last, applications of NGS are summarized. Lin Liu, Yinhu Li, Siliang Li, Ni Hu, Yimin He, Ray Pong, Danni Lin, Lihua Lu, and Maggie Law Copyright © 2012 Lin Liu et al. All rights reserved. Genomic Instability, Inflammation, and Cancer Mon, 02 Jul 2012 08:13:36 +0000 http://www.hindawi.com/journals/bmri/2012/308043/ Vassilis G. Gorgoulis Copyright © 2012 Vassilis G. Gorgoulis. All rights reserved. How Do Cytokines Trigger Genomic Instability? Sun, 17 Jun 2012 15:16:31 +0000 http://www.hindawi.com/journals/bmri/2012/536761/ Inflammation is a double-edged sword presenting a dual effect on cancer development, from one hand promoting tumor initiation and progression and from the other hand protecting against cancer through immunosurveillance mechanisms. Cytokines are crucial components of inflammation, participating in the interaction between the cells of tumor microenvironment. A comprehensive study of the role of cytokines in the context of the inflammation-tumorigenesis interplay helps us to shed light in the pathogenesis of cancer. In this paper we focus on the role of cytokines in the development of genomic instability, an evolving hallmark of cancer. Ioannis L. Aivaliotis, Ioannis S. Pateras, Marilena Papaioannou, Christina Glytsou, Konstantinos Kontzoglou, Elizabeth O. Johnson, and Vassilis Zoumpourlis Copyright © 2012 Ioannis L. Aivaliotis et al. All rights reserved. Oral Lichen Planus as a Preneoplastic Inflammatory Model Thu, 17 May 2012 09:51:45 +0000 http://www.hindawi.com/journals/bmri/2012/759626/ Oral lichen planus (OLP) is a chronic oral inflammatory disease of unknown etiology. According to reports, 1-2% of OLP patients develop oral squamous cell carcinoma (OSCC) in the long run. While World Health Organization (WHO) classifies OLP as “a potentially malignant disorder,” it is still a matter of debate which mechanisms drive OLP to such a condition. The current hypothesis connecting OLP and OSCC is that chronic inflammation results in crucial DNA damage which over time results in cancer development. Initial studies investigating the OLP and OSCC link were mainly retrospective clinical studies. Over the past years, several amount of information has accumulated, mainly from molecular studies on the OLP malignant potential. This article is a critical review of whether OLP has a malignant potential and, therefore, represents a model of preneoplastic inflammation. Eleni A. Georgakopoulou, Marina D. Achtari, Michael Achtaris, Periklis G. Foukas, and Athanassios Kotsinas Copyright © 2012 Eleni A. Georgakopoulou et al. All rights reserved. Inference of Tumor Phylogenies from Genomic Assays on Heterogeneous Samples Sun, 13 May 2012 15:40:17 +0000 http://www.hindawi.com/journals/bmri/2012/797812/ Tumorigenesis can in principle result from many combinations of mutations, but only a few roughly equivalent sequences of mutations, or “progression pathways,” seem to account for most human tumors. Phylogenetics provides a promising way to identify common progression pathways and markers of those pathways. This approach, however, can be confounded by the high heterogeneity within and between tumors, which makes it difficult to identify conserved progression stages or organize them into robust progression pathways. To tackle this problem, we previously developed methods for inferring progression stages from heterogeneous tumor profiles through computational unmixing. In this paper, we develop a novel pipeline for building trees of tumor evolution from the unmixed tumor data. The pipeline implements a statistical approach for identifying robust progression markers from unmixed tumor data and calling those markers in inferred cell states. The result is a set of phylogenetic characters and their assignments in progression states to which we apply maximum parsimony phylogenetic inference to infer tumor progression pathways. We demonstrate the full pipeline on simulated and real comparative genomic hybridization (CGH) data, validating its effectiveness and making novel predictions of major progression pathways and ancestral cell states in breast cancers. Ayshwarya Subramanian, Stanley Shackney, and Russell Schwartz Copyright © 2012 Ayshwarya Subramanian et al. All rights reserved. Detection of Herplex Simplex Virus-1 and -2 in Cardiac Myxomas Tue, 28 Feb 2012 08:03:00 +0000 http://www.hindawi.com/journals/bmri/2012/823949/ The etiology of sporadic cardiac myxomas remains elusive. The tendency for these lesions to recur following resection, their immunopathological characteristics, along with their histological and molecular profile, may implicate the presence of an infective agent in this type of tumor. In this study, we investigated the presence of herpes simplex virus (HSV) DNA in a cohort of cardiac myxomas in a tertiary referral centre. Twenty-nine formalin-fixed paraffin-embedded (FFPE) sporadic cardiac myxomas were obtained, 17 of which were shown to be informative. These were compared to 19 macroscopically and microscopically normal heart tissue specimens. The detection of HSV-1 and -2 genomic sequences was achieved with the use of a combined nested PCR-Restriction Fragment Length Polymorphism methodology. The presence of HSV-1 and/or -2 DNA was demonstrated in 6 of 17 (35%) informative sporadic cardiac myxomas, whereas no HSV DNA was detected in normal heart tissues (𝑃<0.01). The existence of HSV-1/2 DNA in sporadic cardiac myxomas, along with its absence from normal heart tissues, reinforces the possibility that HSV infection might be involved in the development of these lesions. Our findings raise the point of anti-HSV medication postsurgically with a potential benefit in reducing the rate of recurrences. Ioannis S. Pateras, Konstantinos Evangelou, Katerina Tsimaratou, Michalis Liontos, Stratigoula Sakellariou, Theodoros Barlogiannis, Petros Karakitsos, Apostolos Papalois, Athanassios Kotsinas, and Vassilis G. Gorgoulis Copyright © 2012 Ioannis S. Pateras et al. All rights reserved. Asbestos-Induced Cellular and Molecular Alteration of Immunocompetent Cells and Their Relationship with Chronic Inflammation and Carcinogenesis Mon, 06 Feb 2012 10:04:00 +0000 http://www.hindawi.com/journals/bmri/2012/492608/ Asbestos causes lung fibrosis known as asbestosis as well as cancers such as malignant mesothelioma and lung cancer. Asbestos is a mineral silicate containing iron, magnesium, and calcium with a core of SiO2. The immunological effect of silica, SiO2, involves the dysregulation of autoimmunity because of the complications of autoimmune diseases found in silicosis. Asbestos can therefore cause alteration of immunocompetent cells to result in a decline of tumor immunity. Additionally, due to its physical characteristics, asbestos fibers remain in the lung, regional lymph nodes, and the pleural cavity, particularly at the opening sites of lymphatic vessels. Asbestos can induce chronic inflammation in these areas due to the production of reactive oxygen/nitrogen species. As a consequence, immunocompetent cells can have their cellular and molecular features altered by chronic and recurrent encounters with asbestos fibers, and there may be modification by the surrounding inflammation, all of which eventually lead to decreased tumor immunity. In this paper, the brief results of our investigation regarding reduction of tumor immunity of immunocompetent cells exposed to asbestos in vitro are discussed, as are our findings concerned with an investigation of chronic inflammation and analyses of peripheral blood samples derived from patients with pleural plaque and mesothelioma that have been exposed to asbestos. Hidenori Matsuzaki, Megumi Maeda, Suni Lee, Yasumitsu Nishimura, Naoko Kumagai-Takei, Hiroaki Hayashi, Shoko Yamamoto, Tamayo Hatayama, Yoko Kojima, Rika Tabata, Takumi Kishimoto, Junichi Hiratsuka, and Takemi Otsuki Copyright © 2012 Hidenori Matsuzaki et al. All rights reserved. Role of Nitrative and Oxidative DNA Damage in Inflammation-Related Carcinogenesis Thu, 26 Jan 2012 11:07:55 +0000 http://www.hindawi.com/journals/bmri/2012/623019/ Chronic inflammation induced by biological, chemical, and physical factors has been found to be associated with the increased risk of cancer in various organs. We revealed that infectious agents including liver fluke, Helicobacter pylori, and human papilloma virus and noninfectious agents such as asbestos fiber induced iNOS-dependent formation of 8-nitroguanine and 8-oxo-7, 8-dihydro-2′-deoxyguanosine (8-oxodG) in cancer tissues and precancerous regions. Our results with the colocalization of phosphorylated ATM and γ-H2AX with 8-oxodG and 8-nitroguanine in inflammation-related cancer tissues suggest that DNA base damage leads to double-stranded breaks. It is interesting from the aspect of genetic instability. We also demonstrated IL-6-modulated iNOS expression via STAT3 and EGFR in Epstein-Barr-virus-associated nasopharyngeal carcinoma and found promoter hypermethylation in several tumor suppressor genes. Such epigenetic alteration may occur by controlling the DNA methylation through IL-6-mediated JAK/STAT3 pathways. Collectively, 8-nitroguanine would be a useful biomarker for predicting the risk of inflammation-related cancers. Mariko Murata, Raynoo Thanan, Ning Ma, and Shosuke Kawanishi Copyright © 2012 Mariko Murata et al. All rights reserved. CTLA-4 Gene Polymorphism and the Risk of Systemic Lupus Erythematosus in the Chinese Population Sun, 11 Sep 2011 13:26:41 +0000 http://www.hindawi.com/journals/bmri/2011/167395/ Several variants of CTLA-4 have been reported to be associated with susceptibility systemic lupus erythematosus (SLE); however, findings have been inconsistent across different populations. Using a case-control study design, we have investigated the role of CTLA-4 polymorphism at positions −1661 and −1722 on SLE susceptibility in our Chinese SLE population in central China's Hubei province. Samples were collected from 148 SLE patients and 170 healthy controls. Polymerase chain reaction restriction fragments length polymorphism (PCR-RFLP) was used to analyze the genotypes of the two sites. Statistically significant difference was observed in genotypes for −1722, but not for −1661. The frequency of the T allele on the −1722 SNP was significantly increased in SLE patients: 57.8% versus 40.6% in controls (𝑃<0.001, OR = 2.002). While the detected C allele frequency in the controls was significantly elevated in comparison to that in the SLE patients (59.4% versus 42.2%). On the contrary, no association was found between SLE and CTLA-4 polymorphism at position −1661. Ahmad Taha Khalaf, Ji-Quan Song, Ting-Ting Gao, Xiang-Ping Yu, and Tie-Chi Lei Copyright © 2011 Ahmad Taha Khalaf et al. All rights reserved. Variability in Estrogen-Metabolizing Genes and Their Association with Genomic Instability in Untreated Breast Cancer Patients and Healthy Women Tue, 14 Jun 2011 08:39:29 +0000 http://www.hindawi.com/journals/bmri/2011/571784/ In the present study, we investigated the relationship between polymorphisms in the estrogen-metabolizing genes CYP17, CYP1B1, CYP1A1, and COMT and genomic instability in the peripheral blood lymphocytes of 62 BC patients and 62 controls considering that increased or prolonged exposure to estrogen can damage the DNA molecule and increase the genomic instability process in breast tissue. Our data demonstrated increased genomic instability in BC patients and that individuals with higher frequencies of MN exhibited higher risk to BC when belonging Val/Met genotype of the COMT gene. We also observed that CYP17 and CYP1A1 polymorphisms can modify the risk to BC depending on the menopause status. We can conclude that the genetic background in estrogen metabolism pathway can modulate chromosome damage in healthy controls and patients and thereby influence the risk to BC. These findings suggest the importance to ally biomarkers of susceptibility and effects to estimate risk groups. Raquel Alves dos Santos, Ana Cláudia Teixeira, Mônica Beatriz Mayorano, Hélio Humberto Angotti Carrara, Jurandyr Moreira de Andrade, and Catarina Satie Takahashi Copyright © 2011 Raquel Alves dos Santos et al. All rights reserved. Next-Generation Sequencing Tue, 29 Mar 2011 10:47:33 +0000 http://www.hindawi.com/journals/bmri/2010/370710/ Momiao Xiong, Zhongming Zhao, Jonathan Arnold, and Fuli Yu Copyright © 2010 Momiao Xiong et al. All rights reserved. Proteomics Thu, 24 Mar 2011 16:09:18 +0000 http://www.hindawi.com/journals/bmri/2010/453045/ Benjamin A. Garcia, Helen J. Cooper, Pieter C. Dorrestein, Kai Tang, and Beatrix M. Ueberheide Copyright © 2010 Benjamin A. Garcia et al. All rights reserved. Construction, Characterization, and Preliminary BAC-End Sequence Analysis of a Bacterial Artificial Chromosome Library of the Tea Plant (Camellia sinensis) Thu, 23 Dec 2010 10:59:00 +0000 http://www.hindawi.com/journals/bmri/2011/476723/ We describe the construction and characterization of a publicly available BAC library for the tea plant, Camellia sinensis. Using modified methods, the library was constructed with the aim of developing public molecular resources to advance tea plant genomics research. The library consists of a total of 401,280 clones with an average insert size of 135 kb, providing an approximate coverage of 13.5 haploid genome equivalents. No empty vector clones were observed in a random sampling of 576 BAC clones. Further analysis of 182 BAC-end sequences from randomly selected clones revealed a GC content of 40.35% and low chloroplast and mitochondrial contamination. Repetitive sequence analyses indicated that LTR retrotransposons were the most predominant sequence class (86.93%–87.24%), followed by DNA retrotransposons (11.16%–11.69%). Additionally, we found 25 simple sequence repeats (SSRs) that could potentially be used as genetic markers. Jinke Lin, Dave Kudrna, and Rod A. Wing Copyright © 2011 Jinke Lin et al. All rights reserved. Diagnosis and Prognostication of Ductal Adenocarcinomas of the Pancreas Based on Genome-Wide DNA Methylation Profiling by Bacterial Artificial Chromosome Array-Based Methylated CpG Island Amplification Tue, 21 Dec 2010 09:04:17 +0000 http://www.hindawi.com/journals/bmri/2011/780836/ To establish diagnostic criteria for ductal adenocarcinomas of the pancreas (PCs), bacterial artificial chromosome (BAC) array-based methylated CpG island amplification was performed using 139 tissue samples. Twelve BAC clones, for which DNA methylation status was able to discriminate cancerous tissue (T) from noncancerous pancreatic tissue in the learning cohort with a specificity of 100%, were identified. Using criteria that combined the 12 BAC clones, T-samples were diagnosed as cancers with 100% sensitivity and specificity in both the learning and validation cohorts. DNA methylation status on 11 of the BAC clones, which was able to discriminate patients showing early relapse from those with no relapse in the learning cohort with 100% specificity, was correlated with the recurrence-free and overall survival rates in the validation cohort and was an independent prognostic factor by multivariate analysis. Genome-wide DNA methylation profiling may provide optimal diagnostic markers and prognostic indicators for patients with PCs. Masahiro Gotoh, Eri Arai, Saori Wakai-Ushijima, Nobuyoshi Hiraoka, Tomoo Kosuge, Fumie Hosoda, Tatsuhiro Shibata, Tadashi Kondo, Sana Yokoi, Issei Imoto, Johji Inazawa, and Yae Kanai Copyright © 2011 Masahiro Gotoh et al. All rights reserved. Development and Application of Bovine and Porcine Oligonucleotide Arrays with Protein-Based Annotation Tue, 14 Dec 2010 11:42:56 +0000 http://www.hindawi.com/journals/bmri/2010/453638/ The design of oligonucleotide sequences for the detection of gene expression in species with disparate volumes of genome and EST sequence information has been broadly studied. However, a congruous strategy has yet to emerge to allow the design of sensitive and specific gene expression detection probes. This study explores the use of a phylogenomic approach to align transcribed sequences to vertebrate protein sequences for the detection of gene families to design genomewide 70-mer oligonucleotide probe sequences for bovine and porcine. The bovine array contains 23,580 probes that target the transcripts of 16,341 genes, about 72% of the total number of bovine genes. The porcine array contains 19,980 probes targeting 15,204 genes, about 76% of the genes in the Ensembl annotation of the pig genome. An initial experiment using the bovine array demonstrates the specificity and sensitivity of the array. John R. Garbe, Christine G. Elsik, Eric Antoniou, James M. Reecy, Karl J. Clark, Anand Venkatraman, Jae-Woo Kim, Robert D. Schnabel, C. Michael Dickens, Russell D. Wolfinger, Scott C. Fahrenkrug, and Jeremy F. Taylor Copyright © 2010 John R. Garbe et al. All rights reserved. Isolation of BAC Clones Containing Conserved Genes from Libraries of Three Distantly Related Moths: A Useful Resource for Comparative Genomics of Lepidoptera Thu, 25 Nov 2010 13:59:38 +0000 http://www.hindawi.com/journals/bmri/2011/165894/ Lepidoptera, butterflies and moths, is the second largest animal order and includes numerous agricultural pests. To facilitate comparative genomics in Lepidoptera, we isolated BAC clones containing conserved and putative single-copy genes from libraries of three pests, Heliothis virescens, Ostrinia nubilalis, and Plutella xylostella, harboring the haploid chromosome number, 𝑛=31, which are not closely related with each other or with the silkworm, Bombyx mori, (𝑛=28), the sequenced model lepidopteran. A total of 108–184 clones representing 101–182 conserved genes were isolated for each species. For 79 genes, clones were isolated from more than two species, which will be useful as common markers for analysis using fluorescence in situ hybridization (FISH), as well as for comparison of genome sequence among multiple species. The PCR-based clone isolation method presented here is applicable to species which lack a sequenced genome but have a significant collection of cDNA or EST sequences. Yuji Yasukochi, Makiko Tanaka-Okuyama, Manabu Kamimura, Ryo Nakano, Yota Naito, Yukio Ishikawa, and Ken Sahara Copyright © 2011 Yuji Yasukochi et al. All rights reserved. Isolation of Specific Clones from Nonarrayed BAC Libraries through Homologous Recombination Wed, 13 Oct 2010 18:30:16 +0000 http://www.hindawi.com/journals/bmri/2011/560124/ We have developed a new approach to screen bacterial artificial chromosome (BAC) libraries by recombination selection. To test this method, we constructed an orangutan BAC library using an E. coli strain (DY380) with temperature inducible homologous recombination (HR) capability. We amplified one library segment, induced HR at C to make it recombination proficient, and prepared electrocompetent cells for transformation with a kanamycin cassette to target sequences in the orangutan genome through terminal recombineering homologies. Kanamycin-resistant colonies were tested for the presence of BACs containing the targeted genes by the use of a PCR-assay to confirm the presence of the kanamycin insertion. The results indicate that this is an effective approach for screening clones. The advantage of recombination screening is that it avoids the high costs associated with the preparation, screening, and archival storage of arrayed BAC libraries. In addition, the screening can be conceivably combined with genetic engineering to create knockout and reporter constructs for functional studies. Mikhail Nefedov, Lucia Carbone, Matthew Field, Jacquie Schein, and Pieter J. de Jong Copyright © 2011 Mikhail Nefedov et al. All rights reserved. Serine Protease Variants Encoded by Echis ocellatus Venom Gland cDNA: Cloning and Sequencing Analysis Wed, 29 Sep 2010 10:49:31 +0000 http://www.hindawi.com/journals/bmri/2010/134232/ Envenoming by Echis saw-scaled viper is the leading cause of death and morbidity in Africa due to snake bite. Despite its medical importance, there have been few investigations into the toxin composition of the venom of this viper. Here, we report the cloning of cDNA sequences encoding four groups or isoforms of the haemostasis-disruptive Serine protease proteins (SPs) from the venom glands of Echis ocellatus. All these SP sequences encoded the cysteine residues scaffold that form the 6-disulphide bonds responsible for the characteristic tertiary structure of venom serine proteases. All the Echis ocellatus EoSP groups showed varying degrees of sequence similarity to published viper venom SPs. However, these groups also showed marked intercluster sequence conservation across them which were significantly different from that of previously published viper SPs. Because viper venom SPs exhibit a high degree of sequence similarity and yet exert profoundly different effects on the mammalian haemostatic system, no attempt was made to assign functionality to the new Echis ocellatus EoSPs on the basis of sequence alone. The extraordinary level of interspecific and intergeneric sequence conservation exhibited by the Echis ocellatus EoSPs and analogous serine proteases from other viper species leads us to speculate that antibodies to representative molecules should neutralise (that we will exploit, by epidermal DNA immunization) the biological function of this important group of venom toxins in vipers that are distributed throughout Africa, the Middle East, and the Indian subcontinent. S. S. Hasson, R. A. Mothana, T. A. Sallam, M. S. Al-balushi, M. T. Rahman, and A. A. Al-Jabri Copyright © 2010 S. S. Hasson et al. All rights reserved. Construction and Characterization of a Bacterial Artificial Chromosome Library for the A-Genome of Cotton (G. arboreum L.) Mon, 23 Aug 2010 11:09:18 +0000 http://www.hindawi.com/journals/bmri/2011/457137/ A bacterial artificial chromosome (BAC) library for the A-genome of cotton has been constructed from the leaves of G. arboreum L cv. Jianglinzhongmian. It is used as elite A-genome germplasm resources in the present cotton breeding program and has been used to build a genetic reference map of cotton. The BAC library consists of 123,648 clones stored in 322 384-well plates. Statistical analysis of a set of 103 randomly selected BAC clones indicated that each clone has an average insert length of 100.2 kb per plasmid, with a range of 30 to 190 kb. Theoretically, this represents 7.2 haploid genome equivalents based on an A-genome size of 1697 Mb. The BAC library has been arranged in column pools and superpools allowing screening with various PCR-based markers. In the future, the A-genome cotton BAC library will serve as both a giant gene resource and a valuable tool for map-based gene isolation, physical mapping and comparative genome analysis. Yan Hu, Yamin Lu, Dan Ma, Wangzhen Guo, and Tianzhen Zhang Copyright © 2011 Yan Hu et al. All rights reserved. Global Egr1-miRNAs Binding Analysis in PMA-Induced K562 Cells Using ChIP-Seq Tue, 03 Aug 2010 12:36:58 +0000 http://www.hindawi.com/journals/bmri/2010/867517/ Although much is known about microRNAs' regulation in gene expression and their contributions in cell fate, to date, globally lineage-(cell-) specific identification of the binding events between a transcription factor and its targeting microRNA genes is still waiting for elucidation. In this paper, we performed a ChIP-Seq experiment to find the targeting microRNA genes of a transcription factor, Egr1, in human erythroleukemia cell line K562. We found Egr1 binding sites near the promoters of 124 distinct microRNA genes, accounting for about 42% of the miRNAs which have high-confidence predicted promoters (294). We also found EGR1 bind to another 63 pre-miRNAs. We chose 12 of the 187 microRNAs with Egr1 binding sites to perform ChIP-PCR assays and the positive binding signal from ChIP-PCR confirmed the ChIP-Seq results. Our experiments provide the first global binding profile between Egr1 and its targeting microRNA genes in PMA-treated K562 cells, which may facilitate the understanding of pathways controlling microRNA biology in this specific cell line. Wei Wang, Dequang Zhou, Xiaolong Shi, Chao Tang, Xueying Xie, Jing Tu, Qinyu Ge, and Zuhong Lu Copyright © 2010 Wei Wang et al. All rights reserved. Gene-Gene Interaction in Maternal and Perinatal Research Tue, 27 Jul 2010 12:01:19 +0000 http://www.hindawi.com/journals/bmri/2010/853612/ Janet S. Sinsheimer, Robert C. Elston, and Wenjiang J. Fu Copyright © 2010 Janet S. Sinsheimer et al. All rights reserved. Computational Method for Estimating DNA Copy Numbers in Normal Samples, Cancer Cell Lines, and Solid Tumors Using Array Comparative Genomic Hybridization Thu, 08 Jul 2010 14:02:01 +0000 http://www.hindawi.com/journals/bmri/2010/386870/ Genomic copy number variations are a typical feature of cancer. These variations may influence cancer outcomes as well as effectiveness of treatment. There are many computational methods developed to detect regions with deletions and amplifications without estimating actual copy numbers (CN) in these regions. We have developed a computational method capable of detecting regions with deletions and amplifications as well as estimating actual copy numbers in these regions. The method is based on determining how signal intensity from different probes is related to CN, taking into account changes in the total genome size, and incorporating into analysis contamination of the solid tumors with benign tissue. Hidden Markov Model is used to obtain the most likely CN solution. The method has been implemented for Affymetrix 500K GeneChip arrays and Agilent 244K oligonucleotide arrays. The results of CN analysis for normal cell lines, cancer cell lines, and tumor samples are presented. The method is capable of detecting copy number alterations in tumor samples with up to 80% contamination with benign tissue. Analysis of 178 cancer cell lines reveals multiple regions of common homozygous deletions and strong amplifications encompassing known tumor suppressor genes and oncogenes as well as novel cancer related genes. Victor Abkevich, Diana Iliev, Kirsten M. Timms, Thanh Tran, Mark Skolnick, Jerry S. Lanchbury, and Alexander Gutin Copyright © 2010 Victor Abkevich et al. All rights reserved. Uncovering the Complexity of Transcriptomes with RNA-Seq Sun, 27 Jun 2010 11:54:26 +0000 http://www.hindawi.com/journals/bmri/2010/853916/ In recent years, the introduction of massively parallel sequencing platforms for Next Generation Sequencing (NGS) protocols, able to simultaneously sequence hundred thousand DNA fragments, dramatically changed the landscape of the genetics studies. RNA-Seq for transcriptome studies, Chip-Seq for DNA-proteins interaction, CNV-Seq for large genome nucleotide variations are only some of the intriguing new applications supported by these innovative platforms. Among them RNA-Seq is perhaps the most complex NGS application. Expression levels of specific genes, differential splicing, allele-specific expression of transcripts can be accurately determined by RNA-Seq experiments to address many biological-related issues. All these attributes are not readily achievable from previously widespread hybridization-based or tag sequence-based approaches. However, the unprecedented level of sensitivity and the large amount of available data produced by NGS platforms provide clear advantages as well as new challenges and issues. This technology brings the great power to make several new biological observations and discoveries, it also requires a considerable effort in the development of new bioinformatics tools to deal with these massive data files. The paper aims to give a survey of the RNA-Seq methodology, particularly focusing on the challenges that this application presents both from a biological and a bioinformatics point of view. Valerio Costa, Claudia Angelini, Italia De Feis, and Alfredo Ciccodicola Copyright © 2010 Valerio Costa et al. All rights reserved. Identification of microRNAs Involved in the Host Response to Enterovirus 71 Infection by a Deep Sequencing Approach Thu, 10 Jun 2010 09:51:23 +0000 http://www.hindawi.com/journals/bmri/2010/425939/ Role of microRNA (miRNA) has been highlighted in pathogen-host interactions recently. To identify cellular miRNAs involved in the host response to enterovirus 71 (EV71) infection, we performed a comprehensive miRNA profiling in EV71-infected Hep2 cells through deep sequencing. 64 miRNAs were found whose expression levels changed for more than 2-fold in response to EV71 infection. Gene ontology analysis revealed that many of these mRNAs play roles in neurological process, immune response, and cell death pathways, which are known to be associated with the extreme virulence of EV71. To our knowledge, this is the first study on host miRNAs expression alteration response to EV71 infection. Our findings supported the hypothesis that certain miRNAs might be essential in the host-pathogen interactions. Lunbiao Cui, Xiling Guo, Yuhua Qi, Xian Qi, Yiyue Ge, Zhiyang Shi, Tao Wu, Jun Shan, Yunfeng Shan, Zheng Zhu, and Hua Wang Copyright © 2010 Lunbiao Cui et al. All rights reserved. Detection of Fetomaternal Genotype Associations in Early-Onset Disorders: Evaluation of Different Methods and Their Application to Childhood Leukemia Wed, 09 Jun 2010 12:03:01 +0000 http://www.hindawi.com/journals/bmri/2010/369534/ Several designs and analytical approaches have been proposed to dissect offspring from maternal genetic contributions to early-onset diseases. However, lack of parental controls halts the direct verification of the assumption of mating symmetry (MS) required to assess maternally-mediated effects. In this study, we used simulations to investigate the performance of existing methods under mating asymmetry (MA) when parents of controls are missing. Our results show that the log-linear, likelihood-based framework using a case-triad/case-control hybrid design provides valid tests for maternal genetic effects even under MA. Using this approach, we examined fetomaternal associations between 29 SNPs in 12 cell-cycle genes and childhood pre-B acute lymphoblastic leukemia (ALL). We identified putative fetomaternal effects at loci CDKN2A rs36228834 (𝑃=.017) and CDKN2B rs36229158 (𝑃=.022) that modulate the risk of childhood ALL. These data further corroborate the importance of the mother's genotype on the susceptibility to early-onset diseases. Jasmine Healy, Mathieu Bourgey, Chantal Richer, Daniel Sinnett, and Marie-Helene Roy-Gagnon Copyright © 2010 Jasmine Healy et al. All rights reserved. High-Throughput Sequencing of MicroRNAs in Adenovirus Type 3 Infected Human Laryngeal Epithelial Cells Wed, 09 Jun 2010 10:06:08 +0000 http://www.hindawi.com/journals/bmri/2010/915980/ Adenovirus infection can cause various illnesses depending on the infecting serotype, such as gastroenteritis, conjunctivitis, cystitis, and rash illness, but the infection mechanism is still unknown. MicroRNAs (miRNA) have been reported to play essential roles in cell proliferation, cell differentiation, and pathogenesis of human diseases including viral infections. We analyzed the miRNA expression profiles from adenovirus type 3 (AD3) infected Human laryngeal epithelial (Hep2) cells using a SOLiD deep sequencing. 492 precursor miRNAs were identified in the AD3 infected Hep2 cells, and 540 precursor miRNAs were identified in the control. A total of 44 miRNAs demonstrated high expression and 36 miRNAs showed lower expression in the AD3 infected cells than control. The biogenesis of miRNAs has been analyzed, and some of the SOLiD results were confirmed by Quantitative PCR analysis. The present studies may provide a useful clue for the biological function research into AD3 infection. Yuhua Qi, Jing Tu, Lunbiao Cui, Xiling Guo, Zhiyang Shi, Shuchun Li, Wenting Shi, Yunfeng Shan, Yiyue Ge, Jun Shan, Hua Wang, and Zuhong Lu Copyright © 2010 Yuhua Qi et al. All rights reserved. Features of Recent Codon Evolution: A Comparative Polymorphism-Fixation Study Mon, 07 Jun 2010 09:08:07 +0000 http://www.hindawi.com/journals/bmri/2010/202918/ Features of amino-acid and codon changes can provide us important insights on protein evolution. So far, investigators have often examined mutation patterns at either interspecies fixed substitution or intraspecies nucleotide polymorphism level, but not both. Here, we performed a unique analysis of a combined set of intra-species polymorphisms and inter-species substitutions in human codons. Strong difference in mutational pattern was found at codon positions 1, 2, and 3 between the polymorphism and fixation data. Fixation had strong bias towards increasing the rarest codons but decreasing the most frequently used codons, suggesting that codon equilibrium has not been reached yet. We detected strong CpG effect on CG-containing codons and subsequent suppression by fixation. Finally, we detected the signature of purifying selection against A∣U dinucleotides at synonymous dicodon boundaries. Overall, fixation process could effectively and quickly correct the volatile changes introduced by polymorphisms so that codon changes could be gradual and directional and that codon composition could be kept relatively stable during evolution. Zhongming Zhao and Cizhong Jiang Copyright © 2010 Zhongming Zhao and Cizhong Jiang. All rights reserved. Proteomics of Plant Pathogenic Fungi Thu, 27 May 2010 08:51:24 +0000 http://www.hindawi.com/journals/bmri/2010/932527/ Plant pathogenic fungi cause important yield losses in crops. In order to develop efficient and environmental friendly crop protection strategies, molecular studies of the fungal biological cycle, virulence factors, and interaction with its host are necessary. For that reason, several approaches have been performed using both classical genetic, cell biology, and biochemistry and the modern, holistic, and high-throughput, omic techniques. This work briefly overviews the tools available for studying Plant Pathogenic Fungi and is amply focused on MS-based Proteomics analysis, based on original papers published up to December 2009. At a methodological level, different steps in a proteomic workflow experiment are discussed. Separate sections are devoted to fungal descriptive (intracellular, subcellular, extracellular) and differential expression proteomics and interactomics. From the work published we can conclude that Proteomics, in combination with other techniques, constitutes a powerful tool for providing important information about pathogenicity and virulence factors, thus opening up new possibilities for crop disease diagnosis and crop protection. Raquel González-Fernández, Elena Prats, and Jesús V. Jorrín-Novo Copyright © 2010 Raquel González-Fernández et al. All rights reserved. The Relationship between Birthweight and Longitudinal Changes of Blood Pressure Is Modulated by Beta-Adrenergic Receptor Genes: The Bogalusa Heart Study Tue, 11 May 2010 15:37:35 +0000 http://www.hindawi.com/journals/bmri/2010/543514/ This study examines the genetic influence of -adrenergic receptor gene polymorphisms (-AR Arg16Gly and -AR Trp64Arg) on the relationship of birthweight to longitudinal changes of blood pressure (BP) from childhood to adulthood in 224 black and 515 white adults, aged 21–47 years, enrolled in the Bogalusa Heart Study. Blacks showed significantly lower birthweight and frequencies of -AR Gly16 and -AR Trp64 alleles and higher BP levels and age-related trends than whites. In multivariable regression analyses using race-adjusted BP and birthweight, low birthweight was associated with greater increase in age-related trend of systolic BP (standardized regression coefficient , ) and diastolic BP (, ) in the combined sample of blacks and whites, adjusting for the first BP measurement in childhood, sex, age, and gestational age. Adjustment for the current body mass index strengthened the birthweight-BP association. Importantly, the strength of the association, measured as regression coefficients, was modulated by the combination of -AR and -AR genotypes for systolic ( for interaction) and diastolic BP age-related trend ( for interaction), with blacks and whites showing a similar trend in the interaction. These findings indicate that the intrauterine programming of BP regulation later in life depends on -AR genotypes. Wei Chen, Sathanur R. Srinivasan, D. Michael Hallman, and Gerald S. Berenson Copyright © 2010 Wei Chen et al. All rights reserved. Maternal-Zygotic Epistasis and the Evolution of Genetic Diseases Mon, 10 May 2010 09:56:34 +0000 http://www.hindawi.com/journals/bmri/2010/478732/ Many birth defects and genetic diseases are expressed in individuals that do not carry the disease causing alleles. Genetic diseases observed in offspring can be caused by gene expression in mothers and by interactions between gene expression in mothers and offspring. It is not clear whether the underlying pattern of gene expression (maternal versus offspring) affects the incidence of genetic disease. Here we develop a 2-locus population genetic model with epistatic interactions between a maternal gene and a zygotic gene to address this question. We show that maternal effect genes that affect disease susceptibility in offspring persist longer and at higher frequencies in a population than offspring genes with the same effects. We find that specific forms of maternal-zygotic epistasis can maintain disease causing alleles at high frequencies over a range of plausible values. Our findings suggest that the strength and form of epistasis and the underlying pattern of gene expression may greatly influence the prevalence of human genetic diseases. Nicholas K. Priest and Michael J. Wade Copyright © 2010 Nicholas K. Priest and Michael J. Wade. All rights reserved. Evaluation of Hepcidin Isoforms in Hemodialysis Patients by a Proteomic Approach Based on SELDI-TOF MS Thu, 15 Apr 2010 09:31:37 +0000 http://www.hindawi.com/journals/bmri/2010/329646/ The hepatic iron regulator hormone hepcidin consists, in its mature form, of 25 amino acids, but two other isoforms, hepcidin-20 and hepcidin-22, have been reported, whose biological meaning remains poorly understood. We evaluated hepcidin isoforms in sera from 57 control and 54 chronic haemodialysis patients using a quantitative proteomic approach based on SELDI-TOF-MS. Patients had elevated serum levels of both hepcidin-25 and hepcidin-20 as compared to controls (geometric means: 7.52 versus 4.69 nM, and 4.06 versus 1.76 nM, resp., for both). The clearance effects of a single dialysis session by different dialysis techniques and membranes were also investigated, showing an average reduction by % for hepcidin-25 and % for hepcidin-20 but only minor differences among the different dialysis modalities. Measurement of hepcidin isoforms through MS-based techniques can be a useful tool for better understanding of their biological role in hemodialysis patients and other clinical conditions. Natascia Campostrini, Annalisa Castagna, Federica Zaninotto, Valeria Bedogna, Nicola Tessitore, Albino Poli, Nicola Martinelli, Antonio Lupo, Oliviero Olivieri, and Domenico Girelli Copyright © 2010 Natascia Campostrini et al. All rights reserved. Evidence for Maternal-Fetal Genotype Incompatibility as a Risk Factor for Schizophrenia Tue, 06 Apr 2010 10:16:22 +0000 http://www.hindawi.com/journals/bmri/2010/576318/ Prenatal/obstetric complications are implicated in schizophrenia susceptibility. Some complications may arise from maternal-fetal genotype incompatibility, a term used to describe maternal-fetal genotype combinations that produce an adverse prenatal environment. A review of maternal-fetal genotype incompatibility studies suggests that schizophrenia susceptibility is increased by maternal-fetal genotype combinations at the RHD and HLA-B loci. Maternal-fetal genotype combinations at these loci are hypothesized to have an effect on the maternal immune system during pregnancy which can affect fetal neurodevelopment and increase schizophrenia susceptibility. This article reviews maternal-fetal genotype incompatibility studies and schizophrenia and discusses the hypothesized biological role of these ‘‘incompatibility genes’’. It concludes that research is needed to further elucidate the role of RHD and HLA-B maternal-fetal genotype incompatibility in schizophrenia and to identify other genes that produce an adverse prenatal environment through a maternal-fetal genotype incompatibility mechanism. Efforts to develop more sophisticated study designs and data analysis techniques for modeling maternal-fetal genotype incompatibility effects are warranted. Christina G. S. Palmer Copyright © 2010 Christina G. S. Palmer. All rights reserved. Genetic Risk for Recurrent Urinary Tract Infections in Humans: A Systematic Review Tue, 30 Mar 2010 09:24:39 +0000 http://www.hindawi.com/journals/bmri/2010/321082/ Urinary tract infections (UTIs) are a frequent cause of morbidity in children and adults and affect up to 10% of children; its recurrence rate is estimated at 30–40%. UTI may occur in up to 50% of all women in their lifetimes and frequently require medication. Recent advances have suggested that a deregulation of candidate genes in humans may predispose patients to recurrent UTI. The identification of a genetic component of UTI recurrences will make it possible to diagnose at-risk adults and to predict genetic recurrences in their offspring. Six out of 14 genes investigated in humans may be associated with susceptibility to recurrent UTI in humans. In particular, the HSPA1B, CXCR1 & 2, TLR2, TLR4, TGF-𝛽1 genes seem to be associated with an alteration of the host response to UTIs at various levels. M. Zaffanello, G. Malerba, L. Cataldi, F. Antoniazzi, M. Franchini, E. Monti, and V. Fanos Copyright © 2010 M. Zaffanello et al. All rights reserved. Proteomics Strategy for Identifying Candidate Bioactive Proteins in Complex Mixtures: Application to the Platelet Releasate Wed, 24 Mar 2010 14:09:15 +0000 http://www.hindawi.com/journals/bmri/2010/107859/ Proteomic approaches have proven powerful at identifying large numbers of proteins, but there are fewer reports of functional characterization of proteins in biological tissues. Here, we describe an experimental approach that fractionates proteins released from human platelets, linking bioassay activity to identity. We used consecutive orthogonal separation platforms to ensure sensitive detection: (a) ion-exchange of intact proteins, (b) SDS-PAGE separation of ion-exchange fractions and (c) HPLC separation of tryptic digests coupled to electrospray tandem mass spectrometry. Migration of THP-1 monocytes in response to complete or fractionated platelet releasate was assessed and located to just one of the forty-nine ion-exchange fractions. Over 300 proteins were identified in the releasate, with a wide range of annotated biophysical and biochemical properties, in particular platelet activation, adhesion, and wound healing. The presence of PEDF and involucrin, two proteins not previously reported in platelet releasate, was confirmed by western blotting. Proteins identified within the fraction with monocyte promigratory activity and not in other inactive fractions included vimentin, PEDF, and TIMP-1. We conclude that this analytical platform is effective for the characterization of complex bioactive samples. Roisin O'Connor, Lorna M. Cryan, Kieran Wynne, Andreas de Stefani, Desmond Fitzgerald, Colm O'Brien, and Gerard Cagney Copyright © 2010 Roisin O'Connor et al. All rights reserved. Enhanced MALDI-TOF MS Analysis of Phosphopeptides Using an Optimized DHAP/DAHC Matrix Sun, 21 Mar 2010 07:25:00 +0000 http://www.hindawi.com/journals/bmri/2010/759690/ Selecting an appropriate matrix solution is one of the most effective means of increasing the ionization efficiency of phosphopeptides in matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). In this study, we systematically assessed matrix combinations of 2, 6-dihydroxyacetophenone (DHAP) and diammonium hydrogen citrate (DAHC), and demonstrated that the low ratio DHAP/DAHC matrix was more effective in enhancing the ionization of phosphopeptides. Low femtomole level of phosphopeptides from the tryptic digests of -casein and -casein was readily detected by MALDI-TOF-MS in both positive and negative ion mode without desalination or phosphopeptide enrichment. Compared with the DHB/PA matrix, the optimized DHAP/DAHC matrix yielded superior sample homogeneity and higher phosphopeptide measurement sensitivity, particularly when multiple phosphorylated peptides were assessed. Finally, the DHAP/DAHC matrix was applied to identify phosphorylation sites from -casein and -casein and to characterize two phosphorylation sites from the human histone H1 treated with Cyclin-Dependent Kinase-1 (CDK1) by MALDI-TOF/TOF MS. Junjie Hou, Zhensheng Xie, Peng Xue, Ziyou Cui, Xiulan Chen, Jing Li, Tanxi Cai, Peng Wu, and Fuquan Yang Copyright © 2010 Junjie Hou et al. All rights reserved. Association of Combined Maternal-Fetal TNF- Gene G308A Genotypes with Preterm Delivery: A Gene-Gene Interaction Study Tue, 09 Mar 2010 16:04:42 +0000 http://www.hindawi.com/journals/bmri/2010/396184/ Preterm delivery (PTD) is a complicated perinatal adverse event. We were interested in association of G308A polymorphism in tumor necrosis factor- (TNF-) gene with PTD; so we conducted a genetic epidemiology study in Anqing City, Anhui Province, China. Case families and control families were all collected between July 1999 and June 2002. To control potential population stratification as we could, all eligible subjects were ethnic Han Chinese. 250 case families and 247 control families were included in data analysis. A hybrid design which combines case-parent triads and control parents was employed, to test maternal-fetal genotype (MFG) incompatibility. The method is based on a log-linear modeling approach. In summary, we found that when the mother's or child's genotype was G/A, there was a reduced risk of PTD; however when the mother's or child's genotype was genotype A/A, there was a relatively higher risk of PTD. Combined maternal-fetal genotype GA/GA showed the most reduced risk of PTD. Comparison of the LRTs showed that the model with maternal-fetal genotype effects fits significantly better than the model with only maternal and fetal genotype main effects (log-likelihood = −719.4, , significant at 0.05 level). That means that the combined maternal-fetal genotype incompatibility was significantly associated with PTD. The model with maternal-fetal genotype effects can be considered a gene-gene interaction model. We claim that both maternal effects and fetal effects should be considered together while investigating genetic factors of certain perinatal diseases. Mingbin Liang, Xun Wang, Jin Li, Fan Yang, Zhian Fang, Lihua Wang, Yonghua Hu, and Dafang Chen Copyright © 2010 Mingbin Liang et al. All rights reserved. Gene Expression Profiling of Placentas Affected by Pre-Eclampsia Wed, 03 Mar 2010 11:18:42 +0000 http://www.hindawi.com/journals/bmri/2010/787545/ Several studies point to the placenta as the primary cause of pre-eclampsia. Our objective was to identify placental genes that may contribute to the development of pre-eclampsia. RNA was purified from tissue biopsies from eleven pre-eclamptic placentas and eighteen normal controls. Messenger RNA expression from pooled samples was analysed by microarrays. Verification of the expression of selected genes was performed using real-time PCR. A surprisingly low number of genes (21 out of 15,000) were identified as differentially expressed. Among these were genes not previously associated with pre-eclampsia as bradykinin B1 receptor and a 14-3-3 protein, but also genes that have already been connected with pre-eclampsia, for example, inhibin beta A subunit and leptin. A low number of genes were repeatedly identified as differentially expressed, because they may represent the endpoint of a cascade of events effectuated throughout gestation. They were associated with transcriptional regulation and vasoregulative pathways, along with a number of hypothetical proteins and gene sequences with unknown functions. Anne Mette Hoegh, Rehannah Borup, Finn Cilius Nielsen, Steen Sørensen, and Thomas V. F. Hviid Copyright © 2010 Anne Mette Hoegh et al. All rights reserved. Two-Dimensional Liquid Chromatography Technique Coupled with Mass Spectrometry Analysis to Compare the Proteomic Response to Cadmium Stress in Plants Tue, 23 Feb 2010 15:47:23 +0000 http://www.hindawi.com/journals/bmri/2010/567510/ Plants are useful in studies of metal toxicity, because their physiological responses to different metals are correlated with the metal exposure dose and chemical state. Moreover a network of proteins and biochemical cascades that may lead to a controlled homeostasis of metals has been identified in many plant species. This paper focuses on the global protein variations that occur in a Populus nigra spp. clone (Poli) that has an exceptional tolerance to the presence of cadmium. Protein separation was based on a two-dimensional liquid chromatography technique. A subset of 20 out of 126 peaks were identified as being regulated differently under cadmium stress and were fingerprinted by MALDI-TOF. Proteins that were more abundant in the treated samples were located in the chloroplast and in the mitochondrion, suggesting the importance of these organelles in the response and adaptation to metal stress. Giovanna Visioli, Marta Marmiroli, and Nelson Marmiroli Copyright © 2010 Giovanna Visioli et al. All rights reserved. Insights into the Biology of IRES Elements through Riboproteomic Approaches Tue, 02 Feb 2010 13:42:01 +0000 http://www.hindawi.com/journals/bmri/2010/458927/ Translation initiation is a highly regulated process that exerts a strong influence on the posttranscriptional control of gene expression. Two alternative mechanisms govern translation initiation in eukaryotic mRNAs, the cap-dependent initiation mechanism operating in most mRNAs, and the internal ribosome entry site (IRES)-dependent mechanism, first discovered in picornaviruses. IRES elements are highly structured RNA sequences that, in most instances, require specific proteins for recruitment of the translation machinery. Some of these proteins are eukaryotic initiation factors. In addition, RNA-binding proteins (RBPs) play a key role in internal initiation control. RBPs are pivotal regulators of gene expression in response to numerous stresses, including virus infection. This review discusses recent advances on riboproteomic approaches to identify IRES transacting factors (ITAFs) and the relationship between RNA-protein interaction and IRES activity, highlighting the most relevant features on picornavirus and hepatitis C virus IRESs. Almudena Pacheco and Encarnacion Martinez-Salas Copyright © 2010 Almudena Pacheco and Encarnacion Martinez-Salas. All rights reserved. Improved Label-Free LC-MS Analysis by Wavelet-Based Noise Rejection Thu, 28 Jan 2010 16:26:36 +0000 http://www.hindawi.com/journals/bmri/2010/131505/ Label-free LC-MS analysis allows determining the differential expression level of proteins in multiple samples, without the use of stable isotopes. This technique is based on the direct comparison of multiple runs, obtained by continuous detection in MS mode. Only differentially expressed peptides are selected for further fragmentation, thus avoiding the bias toward abundant peptides typical of data-dependent tandem MS. The computational framework includes detection, alignment, normalization and matching of peaks across multiple sets, and several software packages are available to address these processing steps. Yet, more care should be taken to improve the quality of the LC-MS maps entering the pipeline, as this parameter severely affects the results of all downstream analyses. In this paper we show how the inclusion of a preprocessing step of background subtraction in a common laboratory pipeline can lead to an enhanced inclusion list of peptides selected for fragmentation and consequently to better protein identification. Salvatore Cappadona, Paolo Nanni, Marco Benevento, Fredrik Levander, Piera Versura, Aldo Roda, Sergio Cerutti, and Linda Pattini Copyright © 2010 Salvatore Cappadona et al. All rights reserved. Statistical Analysis of Variation in the Human Plasma Proteome Thu, 14 Jan 2010 16:12:07 +0000 http://www.hindawi.com/journals/bmri/2010/258494/ Quantifying the variation in the human plasma proteome is an essential prerequisite for disease-specific biomarker detection. We report here on the longitudinal and individual variation in human plasma characterized by two-dimensional difference gel electrophoresis (2-D DIGE) using plasma samples from eleven healthy subjects collected three times over a two week period. Fixed-effects modeling was used to remove dye and gel variability. Mixed-effects modeling was then used to quantitate the sources of proteomic variation. The subject-to-subject variation represented the largest variance component, while the time-within-subject variation was comparable to the experimental variation found in a previous technical variability study where one human plasma sample was processed eight times in parallel and each was then analyzed by 2-D DIGE in triplicate. Here, 21 protein spots had larger than 50% CV, suggesting that these proteins may not be appropriate as biomarkers and should be carefully scrutinized in future studies. Seventy-eight protein spots showing differential protein levels between different individuals or individual collections were identified by mass spectrometry and further characterized using hierarchical clustering. The results present a first step toward understanding the complexity of longitudinal and individual variation in the human plasma proteome, and provide a baseline for improved biomarker discovery. Todd H. Corzett, Imola K. Fodor, Megan W. Choi, Vicki L. Walsworth, Kenneth W. Turteltaub, Sandra L. McCutchen-Maloney, and Brett A. Chromy Copyright © 2010 Todd H. Corzett et al. All rights reserved. Gene-Gene Interactions in the Folate Metabolic Pathway and the Risk of Conotruncal Heart Defects Tue, 12 Jan 2010 15:33:08 +0000 http://www.hindawi.com/journals/bmri/2010/630940/ Conotruncal and related heart defects (CTRD) are common, complex malformations. Although there are few established risk factors, there is evidence that genetic variation in the folate metabolic pathway influences CTRD risk. This study was undertaken to assess the association between inherited (i.e., case) and maternal gene-gene interactions in this pathway and the risk of CTRD. Case-parent triads (), ascertained from the Children's Hospital of Philadelphia, were genotyped for ten functional variants of nine folate metabolic genes. Analyses of inherited genotypes were consistent with the previously reported association between MTHFR A1298C and CTRD (adjusted ), but provided no evidence that CTRD was associated with inherited gene-gene interactions. Analyses of the maternal genotypes provided evidence of a MTHFR C677T/CBS 844ins68 interaction and CTRD risk (unadjusted ). This association is consistent with the effects of this genotype combination on folate-homocysteine biochemistry but remains to be confirmed in independent study populations. Philip J. Lupo, Elizabeth Goldmuntz, and Laura E. Mitchell Copyright © 2010 Philip J. Lupo et al. All rights reserved. Proteomic Studies of Cholangiocarcinoma and Hepatocellular Carcinoma Cell Secretomes Sun, 27 Dec 2009 16:08:39 +0000 http://www.hindawi.com/journals/bmri/2010/437143/ Cholangiocarcinoma (CCA) and hepatocellular carcinoma (HCC) occur with relatively high incidence in Thailand. The secretome, proteins secreted from cancer cells, are potentially useful as biomarkers of the diseases. Proteomic analysis was performed on the secreted proteins of cholangiocarcinoma (HuCCA-1) and hepatocellular carcinoma (HCC-S102, HepG2, SK-Hep-1, and Alexander) cell lines. The secretomes of the five cancer cell lines were analyzed by SDS-PAGE combined with LC/MS/MS. Sixty-eight proteins were found to be expressed only in HuCCA-1. Examples include neutrophil gelatinase-associated lipocalin (lipocalin 2), laminin 5 beta 3, cathepsin D precursor, desmoplakin, annexin IV variant, and annexin A5. Immunoblotting was used to confirm the presence of lipocalin 2 in conditioned media and cell lysate of 5 cell lines. The results showed that lipocalin 2 was a secreted protein which is expressed only in the conditioned media of the cholangiocarcinoma cell line. Study of lipocalin 2 expression in different types of cancer and normal tissues from cholangiocarcinoma patients showed that lipocalin 2 was expressed only in the cancer tissues. We suggest that lipocalin 2 may be a potential biomarker for cholangiocarcinoma. Chantragan Srisomsap, Phannee Sawangareetrakul, Pantipa Subhasitanont, Daranee Chokchaichamnankit, Khajeelak Chiablaem, Vaharabhongsa Bhudhisawasdi, Sopit Wongkham, and Jisnuson Svasti Copyright © 2010 Chantragan Srisomsap et al. All rights reserved. Correctness of Protein Identifications of Bacillus subtilis Proteome with the Indication on Potential False Positive Peptides Supported by Predictions of Their Retention Times Wed, 23 Dec 2009 09:37:52 +0000 http://www.hindawi.com/journals/bmri/2010/718142/ The predictive capability of the retention time prediction model based on quantitative structure-retention relationships (QSRR) was tested. QSRR model was derived with the use of set of peptides identified with the highest scores and originated from 8 known proteins annotated as model ones. The predictive ability of the QSRR model was verified with the use of a Bacillus subtilis proteome digest after separation and identification of the peptides by LC-ESI-MS/MS. That ability was tested with three sets of testing peptides assigned to the proteins identified with different levels of confidence. First, the set of peptides identified with the highest scores achieved in the search were considered. Hence, proteins identified on the basis of more than one peptide were taken into account. Furthermore, proteins identified on the basis of just one peptide were also considered and, depending on the possessed scores, both above and below the assumed threshold, were analyzed in two separated sets. The QSRR approach was applied as the additional constraint in proteomic research verifying results of MS/MS ion search and confirming the correctness of the peptides identifications along with the indication of the potential false positives. Katarzyna Macur, Tomasz Bączek, Roman Kaliszan, Caterina Temporini, Federica Corana, Gabriella Massolini, Jolanta Grzenkowicz-Wydra, and Michał Obuchowski Copyright © 2010 Katarzyna Macur et al. All rights reserved. Challenges for Biomarker Discovery in Body Fluids Using SELDI-TOF-MS Sun, 06 Dec 2009 10:30:57 +0000 http://www.hindawi.com/journals/bmri/2010/906082/ Protein profiling using SELDI-TOF-MS has gained over the past few years an increasing interest in the field of biomarker discovery. The technology presents great potential if some parameters, such as sample handling, SELDI settings, and data analysis, are strictly controlled. Practical considerations to set up a robust and sensitive strategy for biomarker discovery are presented. This paper also reviews biological fluids generally available including a description of their peculiar properties and the preanalytical challenges inherent to sample collection and storage. Finally, some new insights for biomarker identification and validation challenges are provided. Muriel De Bock, Dominique de Seny, Marie-Alice Meuwis, Jean-Paul Chapelle, Edouard Louis, Michel Malaise, Marie-Paule Merville, and Marianne Fillet Copyright © 2010 Muriel De Bock et al. All rights reserved. The Mysterious Unfoldome: Structureless, Underappreciated, Yet Vital Part of Any Given Proteome Wed, 25 Nov 2009 11:20:35 +0000 http://www.hindawi.com/journals/bmri/2010/568068/ Contrarily to the general believe, many biologically active proteins lack stable tertiary and/or secondary structure under physiological conditions in vitro. These intrinsically disordered proteins (IDPs) are highly abundant in nature and many of them are associated with various human diseases. The functional repertoire of IDPs complements the functions of ordered proteins. Since IDPs constitute a significant portion of any given proteome, they can be combined in an unfoldome; which is a portion of the proteome including all IDPs (also known as natively unfolded proteins, therefore, unfoldome), and describing their functions, structures, interactions, evolution, and so forth. Amino acid sequence and compositions of IDPs are very different from those of ordered proteins, making possible reliable identification of IDPs at the proteome level by various computational means. Furthermore, IDPs possess a number of unique structural properties and are characterized by a peculiar conformational behavior, including their high stability against low pH and high temperature and their structural indifference toward the unfolding by strong denaturants. These peculiarities were shown to be useful for elaboration of the experimental techniques for the large-scale identification of IDPs in various organisms. Some of the computational and experimental tools for the unfoldome discovery are discussed in this review. Vladimir N. Uversky Copyright © 2010 Vladimir N. Uversky. All rights reserved. Mass Spectrometry-Based Label-Free Quantitative Proteomics Tue, 10 Nov 2009 11:57:32 +0000 http://www.hindawi.com/journals/bmri/2010/840518/ In order to study the differential protein expression in complex biological samples, strategies for rapid, highly reproducible and accurate quantification are necessary. Isotope labeling and fluorescent labeling techniques have been widely used in quantitative proteomics research. However, researchers are increasingly turning to label-free shotgun proteomics techniques for faster, cleaner, and simpler results. Mass spectrometry-based label-free quantitative proteomics falls into two general categories. In the first are the measurements of changes in chromatographic ion intensity such as peptide peak areas or peak heights. The second is based on the spectral counting of identified proteins. In this paper, we will discuss the technologies of these label-free quantitative methods, statistics, available computational software, and their applications in complex proteomics studies. Wenhong Zhu, Jeffrey W. Smith, and Chun-Ming Huang Copyright © 2010 Wenhong Zhu et al. All rights reserved. Quantitative Proteomics Analysis of Maternal Plasma in Down Syndrome Pregnancies Using Isobaric Tagging Reagent (iTRAQ) Thu, 05 Nov 2009 10:36:45 +0000 http://www.hindawi.com/journals/bmri/2010/952047/ Currently no specific biomarkers exist for the screening of pregnancies at risk for down syndrome (DS). Since a quantitative proteomic approach with isobaric labelling (iTRAQ) has recently been suggested to be highly suitable for the discovery of novel plasma biomarkers, we have now used this method to examine for potential quantitative changes in the plasma proteome of the pregnancies bearing DS fetuses in comparison to normal healthy babies. In our study, we used plasma from six women with DS pregnancies and six with uncomplicated pregnancies care were taken to match cases and controls for gestational and maternal age, as these could be a confounder. In our quantitative proteomics analysis we were able to detect 178 proteins using iTRAQ labelling in conjunction with 4800 MALDI TOF/TOF. Amongst these we observed changes in 𝛽HCG, a known screening marker for DS, indicating that our assay was functional. We found a number of elevated proteins Ig lambda chain C region, serum amyloid P-component, amyloid beta A4, and under expressed proteins like gamma-actin and titin in DS pregnancies. These proteins are also found in the sera of patients with Alzheimer disease, which share similar pathologies of DS. Our study therefore indicates that the iTRAQ labelling approach may be indeed useful for the detection of novel biomarkers. Varaprasad Kolla, Paul Jenö, Suzette Moes, Sevgi Tercanli, Olav Lapaire, Mahesh Choolani, and Sinuhe Hahn Copyright © 2010 Varaprasad Kolla et al. All rights reserved. A Proteomic Approach for Plasma Biomarker Discovery with iTRAQ Labelling and OFFGEL Fractionation Sun, 01 Nov 2009 16:20:43 +0000 http://www.hindawi.com/journals/bmri/2010/927917/ Human blood plasma contains a plethora of proteins, encompassing not only proteins that have plasma-based functionalities, but also possibly every other form of low concentrated human proteins. As it circulates through the tissues, the plasma picks up proteins that are released from their origin due to physiological events such as tissue remodeling and cell death. Specific disease processes or tumors are often characterized by plasma “signatures,” which may become obvious via changes in the plasma proteome profile, for example, through over expression of proteins. However, the wide dynamic range of proteins present in plasma makes their analysis very challenging, because high-abundance proteins tend to mask those of lower abundance. In the present study, we used a strategy combining iTRAQ as a reagent which improved the peptide ionization and peptide OFFGEL fractionation that has already been shown, in our previous research, to improve the proteome coverage of cellular extracts. Two prefractioning methods were compared: immunodepletion and a bead-based library of combinatorial hexapeptide technology. Our data suggested that both methods were complementary, with regard to the number of identified proteins. iTRAQ labelling, in association with OFFGEL fractionation, allowed more than 300 different proteins to be characterized from 400 𝜇g of plasma proteins. Emilie Ernoult, Anthony Bourreau, Erick Gamelin, and Catherine Guette Copyright © 2010 Emilie Ernoult et al. All rights reserved. Differential Proteome Analysis of the Preeclamptic Placenta Using Optimized Protein Extraction Sun, 13 Sep 2009 10:27:18 +0000 http://www.hindawi.com/journals/bmri/2010/458748/ The human placenta is a difficult tissue to work with using proteomic technology since it contains large amounts of lipids and glycogen. Both lipids and glycogen are known to interfere with the first step in the two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), the isoelectric focusing. In order to gain the best possible protein separation on 2D-PAGE, an optimized sample preparation protocol for placental proteins was developed. Two different buffers, urea/CHAPS and Hepes, were used for solubilization in combination with six different precipitation methods. The removal of glycogen from the samples by centrifugation was crucial for the final proteome maps. Solubilization with urea/CHAPS in combination with dichloromethane/methanol or acidified acetone proved to be the best precipitation procedures. When applied to clinical placenta samples apolipoprotein A1 was found to be accumulated in the preeclamptic placenta, where it may either have a nutritional effect or act as a modifier of signal transduction. Magnus Centlow, Stefan R. Hansson, and Charlotte Welinder Copyright © 2010 Magnus Centlow et al. All rights reserved. L1 Antisense Promoter Drives Tissue-Specific Transcription of Human Genes Tue, 09 May 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/071753/abs/ Transcription of transposable elements interspersed in the genome is controlled by complex interactions between their regulatory elements and host factors. However, the same regulatory elements may be occasionally used for the transcription of host genes. One such example is the human L1 retrotransposon, which contains an antisense promoter (ASP) driving transcription into adjacent genes yielding chimeric transcripts. We have characterized 49 chimeric mRNAs corresponding to sense and antisense strands of human genes. Here we show that L1 ASP is capable of functioning as an alternative promoter, giving rise to a chimeric transcript whose coding region is identical to the ORF of mRNA of the following genes: KIAA1797, CLCN5, and SLCO1A2. Furthermore, in these cases the activity of L1 ASP is tissue-specific and may expand the expression pattern of the respective gene. The activity of L1 ASP is tissue-specific also in cases where L1 ASP produces antisense RNAs complementary to COL11A1 and BOLL mRNAs. Simultaneous assessment of the activity of L1 ASPs in multiple loci revealed the presence of L1 ASP-derived transcripts in all human tissues examined. We also demonstrate that L1 ASP can act as a promoter in vivo and predict that it has a heterogeneous transcription initiation site. Our data suggest that L1 ASP-driven transcription may increase the transcriptional flexibility of several human genes. Kert Mätlik, Kaja Redik, and Mart Speek Copyright © 2006 Kert Mätlik et al. All rights reserved. The Genomic Distribution of L1 Elements: The Role of Insertion Bias and Natural Selection Sun, 07 May 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/075327/abs/ LINE-1 (L1) retrotransposons constitute the most successful family of retroelements in mammals and account for as much as 20% of mammalian DNA. L1 elements can be found in all genomic regions but they are far more abundant in AT-rich, gene-poor, and low-recombining regions of the genome. In addition, the sex chromosomes and some genes seem disproportionately enriched in L1 elements. Insertion bias and selective processes can both account for this biased distribution of L1 elements. L1 elements do not appear to insert randomly in the genome and this insertion bias can at least partially explain the genomic distribution of L1. The contrasted distribution of L1 and Alu elements suggests that postinsertional processes play a major role in shaping L1 distribution. The most likely mechanism is the loss of recently integrated L1 elements that are deleterious (negative selection) either because of disruption of gene function or their ability to mediate ectopic recombination. By comparison, the retention of L1 elements because of some positive effect is limited to a small fraction of the genome. Understanding the respective importance of insertion bias and selection will require a better knowledge of insertion mechanisms and the dynamics of L1 inserts in populations. Todd Graham and Stephane Boissinot Copyright © 2006 Todd Graham and Stephane Boissinot. All rights reserved. LINE-1 Retrotransposition: Impact on Genome Stability and Diversity and Human Disease Wed, 19 Apr 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/037049/abs/ Nina Luning Prak and Abdelali Haoudi Copyright © 2006 Nina Luning Prak and Abdelali Haoudi. All rights reserved. The Potential Regulation of L1 Mobility by RNA Interference Mon, 20 Mar 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/032713/abs/ The hypothesis that RNA interference constrains L1 mobility seems inherently reasonable: L1 mobility can be dangerous and L1 RNA, the presumed target of RNAi, serves as a critical retrotransposition intermediate. Despite its plausibility, proof for this hypothesis has been difficult to obtain. Studies attempting to link the L1 retrotransposition frequency to alterations in RNAi activity have been hampered by the long times required to measure retrotransposition frequency, the pleiotropic and toxic effects of altering RNAi over similar time periods, and the possibility that other cellular machinery may contribute to the regulation of L1s. Another problem is that the commonly used L1 reporter cassette may serve as a substrate for RNAi. Here we review the L1-RNAi hypothesis and describe a genetic assay with a modified reporter cassette that detects approximately 4 times more L1 insertions than the conventional retrotransposition assay. Shane R. Horman, Petr Svoboda, and Eline T. Luning Prak Copyright © 2006 Shane R. Horman et al. All rights reserved. Do LINEs Have a Role in X-Chromosome Inactivation? Thu, 16 Mar 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/059746/abs/ There is longstanding evidence that X-chromosome inactivation (XCI) travels less successfully in autosomal than in X-chromosomal chromatin. The interspersed repeat elements LINE1s (L1s) have been suggested as candidates for “boosters” which promote the spread of XCI in the X-chromosome. The present paper reviews the current evidence concerning the possible role of L1s in XCI. Recent evidence, accruing from the human genome sequencing project and other sources, confirms that mammalian X-chromosomes are indeed rich in L1s, except in regions where there are many genes escaping XCI. The density of L1s is the highest in the evolutionarily oldest regions. Recent work on X; autosome translocations in human and mouse suggested failure of stabilization of XCI in autosomal material, so that genes are reactivated, but resistance of autosomal genes to the original silencing is not excluded. The accumulation of L1s on the X-chromosome may have resulted from reduced recombination or late replication. Whether L1s are part of the mechanism of XCI or a result of it remains enigmatic. Mary F. Lyon Copyright © 2006 Mary F. Lyon. All rights reserved. DNA Damage and L1 Retrotransposition Wed, 15 Mar 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/037285/abs/ Barbara McClintock was the first to suggest that transposons are a source of genome instability and that genotoxic stress assisted in their mobilization. The generation of double-stranded DNA breaks (DSBs) is a severe form of genotoxic stress that threatens the integrity of the genome, activates cell cycle checkpoints, and, in some cases, causes cell death. Applying McClintock's stress hypothesis to humans, are L1 retrotransposons, the most active autonomous mobile elements in the modern day human genome, mobilized by DSBs? Here, evidence that transposable elements, particularly retrotransposons, are mobilized by genotoxic stress is reviewed. In the setting of DSB formation, L1 mobility may be affected by changes in the substrate for L1 integration, the DNA repair machinery, or the L1 element itself. The review concludes with a discussion of the potential consequences of L1 mobilization in the setting of genotoxic stress. Evan A. Farkash and Eline T. Luning Prak Copyright © 2006 Evan A. Farkash and Eline T. Luning Prak. All rights reserved. LINE-1 Endonuclease-Dependent Retrotranspositional Events Causing Human Genetic Disease: Mutation Detection Bias and Multiple Mechanisms of Target Gene Disruption Thu, 09 Mar 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/056182/abs/ LINE-1 (L1) elements are the most abundant autonomous non-LTR retrotransposons in the human genome. Having recently performed a meta-analysis of L1 endonuclease-mediated retrotranspositional events causing human genetic disease, we have extended this study by focusing on two key issues, namely, mutation detection bias and the multiplicity of mechanisms of target gene disruption. Our analysis suggests that whereas an ascertainment bias may have generally militated against the detection of autosomal L1-mediated insertions, autosomal L1 direct insertions could have been disproportionately overlooked owing to their unusually large size. Our analysis has also indicated that the mechanisms underlying the functional disruption of target genes by L1-mediated retrotranspositional events are likely to be dependent on several different factors such as the type of insertion (L1 direct, L1 trans-driven Alu, or SVA), the precise locations of the inserted sequences within the target gene regions, the length of the inserted sequences, and possibly also their orientation. Jian-Min Chen, Claude Férec, and David N. Cooper Copyright © 2006 Jian-Min Chen et al. All rights reserved. LINE-1 Hypomethylation in a Choline-Deficiency-Induced Liver Cancer in Rats: Dependence on Feeding Period Mon, 06 Mar 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/017142/abs/ Chronic feeding of methyl-donor (methionine, choline, folic acid, and vitamin B12) deficient diet induces hepatocellular carcinoma formation in rats. Previous studies have shown that promoter CpG islands in various cancer-related genes are aberrantly methylated in this model. Moreover, the global genome in methyl-donor-deficient diet fed rats contains a lesser amount of 5-methylcytosine than control livers. It is speculated that more than 90% of all 5-methylcytosines lie within the CpG islands of the transposons, including the long/short interspersed nucleotide elements (LINE and SINE). It is considered that the 5-methylcytosines in LINE-1 limit the ability of retrotransposons to be activated and transcribed; therefore, the extent of hypomethylation of LINE-1 could be a surrogate marker for aberrant methylation in other tumor-related genes as well as genome instability. Additionally, LINE-1 methylation status has been shown to be a good indicator of genome-wide methylation. In this study, we determined cytosine methylation status in the LINE-1 repetitive sequences of rats fed a choline-deficient (CD) diet for various durations and compared these with rats fed a choline-sufficient (CS) diet. The methylation status of LINE-1 was assessed by the combined bisulfite restriction analysis (COBRA) method, where the amount of bisulfite-modified and RsaI-cleaved DNA was quantified using gel electrophoresis. Progressive hypomethylation was observed in LINE-1 of CD livers as a function of feeding time; that is, the amount of cytosine in total cytosine (methylated and unmethylated) increased from 11.1% (1 week) to 19.3% (56 weeks), whereas in the control CS livers, it increased from 9.2% to 12.9%. Hypomethylation in tumor tissues was slightly higher (6%) than the nontumorous surrounding tissue. The present result also indicates that age is a factor influencing the extent of cytosine methylation. Kiyoshi Asada, Yashige Kotake, Rumiko Asada, Deborah Saunders, Robert H. Broyles, Rheal A. Towner, Hiroshi Fukui, and Robert A. Floyd Copyright © 2006 Kiyoshi Asada et al. All rights reserved. The ORF1 Protein Encoded by LINE-1: Structure and Function During L1 Retrotransposition Thu, 16 Feb 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/045621/abs/ LINE-1 or L1 is an autonomous non-LTR retrotransposon in mammals. Retrotransposition requires the function of the two L1-encoded polypeptides, ORF1p and ORF2p. Early recognition of regions of homology between the predicted amino acid sequence of ORF2 and known endonuclease and reverse transcriptase enzymes led to testable hypotheses regarding the function of ORF2p in retrotransposition. As predicted, ORF2p has been demonstrated to have both endonuclease and reverse transcriptase activities. In contrast, no homologs of known function have contributed to our understanding of the function of ORF1p during retrotransposition. Nevertheless, significant advances have been made such that we now know that ORF1p is a high-affinity RNA-binding protein that forms a ribonucleoprotein particle together with L1 RNA. Furthermore, ORF1p is a nucleic acid chaperone and this nucleic acid chaperone activity is required for L1 retrotransposition. Sandra L. Martin Copyright © 2006 Sandra L. Martin. All rights reserved. Do Small RNAs Interfere With LINE-1? Thu, 02 Feb 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/029049/abs/ Long interspersed elements (LINE-1 or L1) are the most active transposable elements in the human genome. Due to their high copy number and ability to sponsor retrotransposition of nonautonomous RNA sequences, unchecked L1 activity can negatively impact the genome by a number of means. Substantial evidence in lower eukaryotes demonstrates that the RNA interference (RNAi) machinery plays a major role in containing transposon activity. Despite extensive analysis in other eukaryotes, no experimental evidence has been presented that L1-derived siRNAs exist, or that the RNAi plays a significant role in restricting L1 activity in the human genome. This review will present evidence showing a direct role for RNAi in suppressing the movement of transposable elements in other eukaryotes, as well as speculate on the role RNAi might play in protecting the human genome from LINE-1 activity. Harris S. Soifer Copyright © 2006 Harris S. Soifer. All rights reserved. L1 Retrotransposons in Human Cancers Thu, 02 Feb 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/083672/abs/ Retrotransposons like L1 are silenced in somatic cells by a variety of mechanisms acting at different levels. Protective mechanisms include DNA methylation and packaging into inactive chromatin to suppress transcription and prevent recombination, potentially supported by cytidine deaminase editing of RNA. Furthermore, DNA strand breaks arising during attempted retrotranspositions ought to activate cellular checkpoints, and L1 activation outside immunoprivileged sites may elicit immune responses. A number of observations indicate that L1 sequences nevertheless become reactivated in human cancer. Prominently, methylation of L1 sequences is diminished in many cancer types and full-length L1 RNAs become detectable, although strong expression is restricted to germ cell cancers. L1 elements have been found to be enriched at sites of illegitimate recombination in many cancers. In theory, lack of L1 repression in cancer might cause transcriptional deregulation, insertional mutations, DNA breaks, and an increased frequency of recombinations, contributing to genome disorganization, expression changes, and chromosomal instability. There is however little evidence that such effects occur at a gross scale in human cancers. Rather, as a rule, L1 repression is only partly alleviated. Unfortunately, many techniques commonly used to investigate genetic and epigenetic alterations in cancer cells are not well suited to detect subtle effects elicited by partial reactivation of retroelements like L1 which are present as abundant, but heterogeneous copies. Therefore, effects of L1 sequences exerted on the local chromatin structure, on the transcriptional regulation of individual genes, and on chromosome fragility need to be more closely investigated in normal and cancer cells. Wolfgang A. Schulz Copyright © 2006 Wolfgang A. Schulz. All rights reserved. Links Between Repeated Sequences Wed, 18 Jan 2006 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2006/013569/abs/ L1 and Alu elements are long and short interspersed retrotransposable elements (LINEs and SINEs) in humans, respectively. Proteins encoded in the autonomous L1 mediate retrotransposition of the nonautonomous Alu and cellular mRNAs. Alu is the only active SINE in the human genome and is derived from 7SL RNA of signal recognition particle. In the other eukaryotic genomes, various tRNA- and 5S rRNA-derived SINEs are found. Some of the tRNA- and 5S rRNA-derived SINEs have partner LINEs of which 3' sequences are similar to those of the SINEs. One of the tRNA-derived SINEs is shown to be mobilized by its partner LINE. Many copies of tRNA and 5S rRNA pseudogenes are present in the human genome. These pseudogenes may have been generated via the retrotransposition process using L1 proteins. Although there are no sequence similarities between L1 and Alu, L1 functionally links with Alu and even cellular genes, impacting on our genome shaping. Sachiko Matsutani Copyright © 2006 Sachiko Matsutani. All rights reserved. Definition of Soybean Genomic Regions That Control Seed Phytoestrogen Amounts Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2004/286806/abs/ Soybean seeds contain large amounts of isoflavones or phytoestrogens such as genistein, daidzein, and glycitein that display biological effects when ingested by humans and animals. In seeds, the total amount, and amount of each type, of isoflavone varies by 5 fold between cultivars and locations. Isoflavone content and quality are one key to the biological effects of soy foods, dietary supplements, and nutraceuticals. Previously we had identified 6 loci (QTL) controlling isoflavone content using 150 DNA markers. This study aimed to identify and delimit loci underlying heritable variation in isoflavone content with additional DNA markers. We used a recombinant inbred line (RIL) population (n=100) derived from the cross of “Essex” by “Forrest,” two cultivars that contrast for isoflavone content. Seed isoflavone content of each RIL was determined by HPLC and compared against 240 polymorphic microsatellite markers by one-way analysis of variance. Two QTL that underlie seed isoflavone content were newly discovered. The additional markers confirmed and refined the positions of the six QTL already reported. The first new region anchored by the marker BARC-Satt063 was significantly associated with genistein (P=0.009, R2=29.5%) and daidzein (P=0.007, R2=17.0%). The region is located on linkage group B2 and derived the beneficial allele from Essex. The second new region defined by the marker BARC-Satt129 was significantly associated with total glycitein (P=0.0005, R2=32.0%). The region is located on linkage group D1a+Q and also derived the beneficial allele from Essex. Jointly the eight loci can explain the heritable variation in isoflavone content. The loci may be used to stabilize seed isoflavone content by selection and to isolate the underlying genes. My A. Kassem, K. Meksem, M. J. Iqbal, V. N. Njiti, W. J. Banz, T. A. Winters, A. Wood, and D. A. Lightfoot Copyright © 2004 All rights reserved. Optimization of Rolling-Circle Amplified Protein Microarrays for Multiplexed Protein Profiling Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/291092/abs/ Protein microarray-based approaches are increasingly being used in research and clinical applications to either profile the expression of proteins or screen molecular interactions. The development of high-throughput, sensitive, convenient, and cost-effective formats for detecting proteins is a necessity for the effective advancement of understanding disease processes. In this paper, we describe the generation of highly multiplexed, antibody-based, specific, and sensitive protein microarrays coupled with rolling-circle signal amplification (RCA) technology. A total of 150 cytokines were simultaneously detected in an RCA sandwich immunoassay format. Greater than half of these proteins have detection sensitivities in the pg/ml range. The validation of antibody microarray with human serum indicated that RCA-based protein microarrays are a powerful tool for high-throughput analysis of protein expression and molecular diagnostics. Weiping Shao, Zhimin Zhou, Isabelle Laroche, Hong Lu, Qiuling Zong, Dhavalkumar D. Patel, Stephen Kingsmore, and Steven P. Piccoli Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. Diagnosis of Ovarian Cancer Using Decision Tree Classification of Mass Spectral Data Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/231929/abs/ Recent reports from our laboratory and others support the SELDI ProteinChip technology as a potential clinical diagnostic tool when combined with n-dimensional analyses algorithms. The objective of this study was to determine if the commercially available classification algorithm biomarker patterns software (BPS), which is based on a classification and regression tree (CART), would be effective in discriminating ovarian cancer from benign diseases and healthy controls. Serum protein mass spectrum profiles from 139 patients with either ovarian cancer, benign pelvic diseases, or healthy women were analyzed using the BPS software. A decision tree, using five protein peaks resulted in an accuracy of 81.5% in the cross-validation analysis and 80%in a blinded set of samples in differentiating the ovarian cancer from the control groups. The potential, advantages, and drawbacks of the BPS system as a bioinformatic tool for the analysis of the SELDI high-dimensional proteomic data are discussed. Antonia Vlahou, John O. Schorge, Betsy W. Gregory, and Robert L. Coleman Copyright © 2003 All rights reserved. Proteomics in Health and Disease Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/232091/abs/ George L. Wright Jr and O. John Semmes Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. Protein and Chemical Microarrays—Powerful Tools for Proteomics Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/256494/abs/ In the last few years, protein and chemical microarrays have emerged as two important tools in the field of proteomics. Specific proteins, antibodies, small molecule compounds, peptides, and carbohydrates can now be immobilized on solid surfaces to form high-density microarrays. Depending on their chemical nature, immobilization of these molecules on solid support is accomplished by in situ synthesis, nonspecific adsorption, specific binding, nonspecific chemical ligation, or chemoselective ligation. These arrays of molecules can then be probed with complex analytes such as serum, total cell extracts, and whole blood. Interactions between the analytes and the immobilized array of molecules are evaluated with a number of different detection systems. In this paper, various components, methods, and applications of the protein and chemical microarray systems are reviewed. Qingchai Xu and Kit S. Lam Copyright © 2003 All rights reserved. Proteomics in Vaccinology and Immunobiology: An Informatics Perspective of the Immunone Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/167607/abs/ The postgenomic era, as manifest, inter alia, by proteomics, offers unparalleled opportunities for the efficient discovery of safe, efficacious, and novel subunit vaccines targeting a tranche of modern major diseases. A negative corollary of this opportunity is the risk of becoming overwhelmed by this embarrassment of riches. Informatics techniques, working to address issues of both data management and through prediction to shortcut the experimental process, can be of enormous benefit in leveraging the proteomic revolution. In this disquisition, we evaluate proteomic approaches to the discovery of subunit vaccines, focussing on viral, bacterial, fungal, and parasite systems. We also adumbrate the impact that proteomic analysis of host-pathogen interactions can have. Finally, we review relevant methods to the prediction of immunome, with special emphasis on quantitative methods, and the subcellular localization of proteins within bacteria. Irini A. Doytchinova and Paul Taylor Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. Bioinformatics Resources for In Silico Proteome Analysis Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/184738/abs/ In the growing field of proteomics, tools for the in silico analysis of proteins and even of whole proteomes are of crucial importance to make best use of the accumulating amount of data. To utilise this data for healthcare and drug development, first the characteristics of proteomes of entire species—mainly the human—have to be understood, before secondly differentiation between individuals can be surveyed. Specialised databases about nucleic acid sequences, protein sequences, protein tertiary structure, genome analysis, and proteome analysis represent useful resources for analysis, characterisation, and classification of protein sequences. Different from most proteomics tools focusing on similarity searches, structure analysis and prediction, detection of specific regions, alignments, data mining, 2D PAGE analysis, or protein modelling, respectively, comprehensive databases like the proteome analysis database benefit from the information stored in different databases and make use of different protein analysis tools to provide computational analysis of whole proteomes. Manuela Pruess and Rolf Apweiler Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. Genomic Regions That Underlie Soybean Seed Isoflavone Content Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2001/192037/abs/ Soy products contain isoflavones (genistein, daidzein, and glycitein) that display biological effects when ingested by humans and animals, these effects are species, dose and age dependent. Therefore, the content and quality of isoflavones in soybeans is a key to their biological effect. Our objective was to identify loci that underlie isoflavone content in soybean seeds. The study involved 100 recombinant inbred lines (RIL) from the cross of ‘Essex’ by ‘Forrest,’ two cultivars that contrast for isoflavone content. Isoflavone content of seeds from each RIL was determined by high performance liquid chromatography (HPLC). The distribution of isoflavone content was continuous and unimodal. The heritability estimates on a line mean basis were 79% for daidzein, 22% for genistein, and 88% for glycitein. Isoflavone content of soybean seeds was compared against 150 polymorphic DNA markers in a one-way analysis of variance. Four genomic regions were found to be significantly associated with the isoflavone content of soybean seeds across both locations and years. Molecular linkage group B1 contained a major QTL underlying glycitein content (P=0.0001,R 2=50.2%), linkage group N contained a QTL for glycitein (P=0.0033,R 2=11.1%) and a QTL for daidzein (P=0.0023,R 2=10.3%) and linkage group A1 contained a QTL for daidzein (P=0.0081,R 2=9.6%). Selection for these chromosomal regions in a marker assisted selection program will allow for the manipulation of amounts and profiles of isoflavones (genistein, daidzein, and glycitein) content of soybean seeds. In addition, tightly linked markers can be used in map based cloning of genes associated with isoflavone content. K. Meksem, V. N. Njiti, W. J. Banz, M. J. Iqbal, My. M. Kassem, D. L. Hyten, J. Yuang, T. A. Winters, and D. A. Lightfoot Copyright © 2001 Hindawi Publishing Corporation. All rights reserved. Use of Immunomatrix Methods to Improve Protein-Protein Interaction Detection Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/340967/abs/ Immunoprecipitation (IP) and coimmunoprecipitation (co-IP) are key techniques for studying protein-protein interactions. These methods utilize immobilized protein A or protein G to isolate antibody-bound target antigens. The main disadvantage of traditional immunoprecipitation and coimmunoprecipitation is that the conditions used to elute the precipitated antigen also release the antibody, contaminating the antigen and destroying the antibody support. To overcome these problems, we describe two methods to generate a reusable antibody support by cross-linking the antibody to immobilized protein A or protein G, or by coupling it directly to the resin. Our studies have demonstrated that the immobilization efficiency for the antibody coupling method was similar for several species of antibody. Furthermore, we illustrate that using both methods of antibody immobilization yields IP and co-IP results similar to traditional protocols but eliminate the antibody heavy and light chains contamination. M. Walid Qoronfleh, Ling Ren, Daryl Emery, Maria Perr, and Barbara Kaboord Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. Selective Enrichment of Membrane Proteins by Partition Phase Separation for Proteomic Studies Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/505318/abs/ The human proteome project will demand faster, easier, and more reliable methods to isolate and purify protein targets. Membrane proteins are the most valuable group of proteins since they are the target for 70–80% of all drugs. Perbio Science has developed a protocol for the quick, easy, and reproducible isolation of integral membrane proteins from eukaryotic cells. This procedure utilizes a proprietary formulation to facilitate cell membrane disruption in a mild, nondenaturing environment and efficiently solubilizes membrane proteins. The technique utilizes a two-phase partitioning system that enables the class separation of hydrophobic and hydrophilic proteins. A variety of protein markers were used to investigate the partitioning efficiency of the membrane protein extraction reagents (Mem-PER) (Mem-PER is a registered trademark of Pierce Biotechnology, Inc) system. These included membrane proteins with one or more transmembrane spanning domains as well as peripheral and cytosolic proteins. Based on densitometry analyses of our Western blots, we obtained excellent solubilization of membrane proteins with less than 10% contamination of the hydrophobic fraction with hydrophilic proteins. Compared to other methodologies for membrane protein solubilization that use time-consuming protocols or expensive and cumbersome instrumentation, the Mem-PER reagents system for eukaryotic membrane protein extraction offers an easy, efficient, and reproducible method to isolate membrane proteins from mammalian and yeast cells. M. Walid Qoronfleh, Betsy Benton, Ray Ignacio, and Barbara Kaboord Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. An Automated Peak Identification/Calibration Procedure for High-Dimensional Protein Measures From Mass Spectrometers Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/957834/abs/ Discovery of “signature” protein profiles that distinguish disease states (eg, malignant, benign, and normal) is a key step towards translating recent advancements in proteomic technologies into clinical utilities. Protein data generated from mass spectrometers are, however, large in size and have complex features due to complexities in both biological specimens and interfering biochemical/physical processes of the measurement procedure. Making sense out of such high-dimensional complex data is challenging and necessitates the use of a systematic data analytic strategy. We propose here a data processing strategy for two major issues in the analysis of such mass-spectrometry-generated proteomic data: (1) separation of protein “signals” from background “noise” in protein intensity measurements and (2) calibration of protein mass/charge measurements across samples. We illustrate the two issues and the utility of the proposed strategy using data from a prostate cancer biomarker discovery project as an example. Yutaka Yasui, Dale McLerran, Bao-Ling Adam, Marcy Winget, Mark Thornquist, and Ziding Feng Copyright © 2003 All rights reserved. SELDI ProteinChip® Array Technology: Protein-Based Predictive Medicine and Drug Discovery Applications Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/874759/abs/ Predictive medicine, utilizing the ProteinChip® Array technology, will develop through the implementation of novel biomarkers and multimarker patterns for detecting disease, determining patient prognosis, monitoring drug effects such as efficacy or toxicity, and for defining treatment options. These biomarkers may also serve as novel protein drug candidates or protein drug targets. In addition, the technology can be used for discovering small molecule drugs or for defining their mode of action utilizing protein-based assays. In this review, we describe the following applications of the ProteinChip Array technology: (1) discovery and identification of novel inhibitors of HIV-1 replication, (2) serum and tissue proteome analysis for the discovery and development of novel multimarker clinical assays for prostate, breast, ovarian, and other cancers, and (3) biomarker and drug discovery applications for neurological disorders. Guru Reddy and Enrique A. Dalmasso Copyright © 2003 Hindawi Publishing Corporation. All rights reserved. The Human Genome Revolution or Evolution? Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2001/865404/abs/ Claude Besmond and Marc Fellous Copyright © 2001 Hindawi Publishing Corporation. All rights reserved. Postgenomics: Proteomics and Bioinformatics in Cancer Research Mon, 01 Jan 1900 00:00:00 +0000 http://www.hindawi.com/journals/bmri/2003/705194/abs/ Now that the human genome is completed, the characterization of the proteins encoded by the sequence remains a challenging task. The study of the complete protein complement of the genome, the “proteome,” referred to as proteomics, will be essential if new therapeutic drugs and new disease biomarkers for early diagnosis are to be developed. Research efforts are already underway to develop the technology necessary to compare the specific protein profiles of diseased versus nondiseased states. These technologies provide a wealth of information and rapidly generate large quantities of data. Processing the large amounts of data will lead to useful predictive mathematical descriptions of biological systems which will permit rapid identification of novel therapeutic targets and identification of metabolic disorders. Here, we present an overview of the current status and future research approaches in defining the cancer cell's proteome in combination with different bioinformatics and computational biology tools toward a better understanding of health and disease. Halima Bensmail and Abdelali Haoudi Copyright © 2003 Hindawi Publishing Corporation. All rights reserved.