Biochemistry Research International

miR-149 Inhibits Non-Small-Cell Lung Cancer Cells EMT by Targeting FOXM1

Yang Ke, Weiyong Zhao, Jie Xiong, and Rubo Cao — Thu, 16 May 2013 12:24:22 +0000

MicroRNAs (miRNAs) have been implied to play crucial roles for epithelial-to-mesenchymal transition (EMT) of non-small-cell lung cancer cells (NSCLC cells). Here we found that the expression of miR-149, downregulated in lung cancer, was inversely correlated with invasive capability and the EMT phenotype of NSCLC cells. miR-149 inhibited EMT in NSCLC cells. Furthermore, we demonstrated that miR-149 directly targeted Forkhead box M1 (FOXM1), and FOXM1 was involved in the EMT induced by TGF-β1 in A549 cells. Overexpression of FOXM1 restored EMT process inhibited by miR-149. Our work suggested that miR-149 might be an EMT suppressor in NSCLC cells.

C-Lobe of Lactoferrin: The Whole Story of the Half-Molecule

Sujata Sharma, Mau Sinha, Sanket Kaushik, Punit Kaur, and Tej P. Singh — Wed, 15 May 2013 13:54:28 +0000

Lactoferrin is an iron-binding diferric glycoprotein present in most of the exocrine secretions. The major role of lactoferrin, which is found abundantly in colostrum, is antimicrobial action for the defense of mammary gland and the neonates. Lactoferrin consists of two equal halves, designated as N-lobe and C-lobe, each of which contains one iron-binding site. While the N-lobe of lactoferrin has been extensively studied and is known for its enhanced antimicrobial effect, the C-lobe of lactoferrin mediates various therapeutic functions which are still being discovered. The potential of the C-lobe in the treatment of gastropathy, diabetes, and corneal wounds and injuries has been indicated. This review provides the details of the proteolytic preparation of C-lobe, and interspecies comparisons of its sequence and structure, as well as the scope of its therapeutic applications.

Effects of Spirulina on Cyclophosphamide-Induced Ovarian Toxicity in Rats: Biochemical and Histomorphometric Evaluation of the Ovary

Thu, 09 May 2013 11:56:55 +0000

Cyclophosphamide (Cyc) is known to cause ovotoxicity and infertility in women. Our aim is to investigate the possible ovotoxic effects of Cyc and possible antioxidant and protective effects of blue-green algae, Spirulina (Sp), in rat ovaries. Eighteen rats were given: group I (, control); group II (, CP), a single dose Cyc; group III (, Sp+Cyc), 7 days Sp+single dose Cyc. Tissue malondialdehyde (MDA) levels, superoxide dismutase (SOD), and catalase (CAT) activities are assessed biochemically. Normal and atretic primordial and primary follicle counts for all sections obtained for each ovary are calculated. Mean number of follicle counts for each group are compared. In Sp+Cyc group, tissue MDA levels were significantly lower than those in the CP and higher than those in the C group (+). Tissue SOD activity was significantly higher in Sp+Cyc group than that in the CP group and lower than that in the C group (+). No statistically significant difference was found between the ovarian CAT activities in any group. Histomorphometrically, there was also no significant difference between the mean numbers of normal and atretic small follicle counts. Our results suggest that single dose Cyc has adverse effects on oxidant status of the ovaries and Sp has protective effects in Cyc-induced ovotoxicity.

Erratum to “Overexpression of PGC-1 Increases Fatty Acid Oxidative Capacity of Human Skeletal Muscle Cells”

Tue, 26 Mar 2013 11:36:26 +0000

The Role of the Cullin-5 E3 Ubiquitin Ligase in the Regulation of Insulin Receptor Substrate-1

Christine Zhiwen Hu, Jaswinder K. Sethi, and Thilo Hagen — Sun, 09 Dec 2012 12:11:19 +0000

Background. SOCS proteins are known to negatively regulate insulin signaling by inhibiting insulin receptor substrate-1 (IRS1). IRS1 has been reported to be a substrate for ubiquitin-dependent proteasomal degradation. Given that SOCS proteins can function as substrate receptor subunits of Cullin-5 E3 ubiquitin ligases, we examined whether Cullin-5 dependent ubiquitination is involved in the regulation of basal IRS1 protein stability and signal-induced IRS1 degradation. Findings. Our results indicate that basal IRS1 stability varies between cell types. However, the Cullin-5 E3 ligase does not play a major role in mediating IRS1 ubiquitination under basal conditions. Protein kinase C activation triggered pronounced IRS1 destabilization. However, this effect was also independent of the function of Cullin-5 E3 ubiquitin ligases. Conclusions. In conclusion, SOCS proteins do not exert a negative regulatory effect on IRS1 by functioning as substrate receptors for Cullin-5-based E3 ubiquitin ligases both under basal conditions and when IRS1 degradation is induced by protein kinase C activation.

Cardiomyopathies: Classification, Clinical Characterization, and Functional Phenotypes

Danuta Szczesna-Cordary, Sachio Morimoto, Aldrin V. Gomes, and Jeffrey R. Moore — Tue, 04 Dec 2012 12:27:54 +0000

Ubiquitin Proteasome System in Stress and Disease

Dmitry Karpov, Michael H. Glickman, Shoshana Bar-Nun, and Philip Coffino — Mon, 26 Nov 2012 17:47:21 +0000

Biochemical Pathways in Cancer

Eun-Kyoung Yim Breuer, Mandi M. Murph, and Rolf J. Craven — Tue, 20 Nov 2012 15:33:23 +0000

A Critical Appraisal of Quantitative Studies of Protein Degradation in the Framework of Cellular Proteostasis

Beatriz Alvarez-Castelao, Carmen Ruiz-Rivas, and José G. Castaño — Sun, 21 Oct 2012 14:30:37 +0000

Protein homeostasis, proteostasis, is essential to understand cell function. Protein degradation is a crucial component of the proteostatic mechanisms of the cell. Experiments on protein degradation are nowadays present in many investigations in the field of molecular and cell biology. In the present paper, we focus on the different experimental approaches to study protein degradation and present a critical appraisal of the results derived from steady-state and kinetic experiments using detection of unlabelled and labelled protein methodologies with a proteostatic perspective. This perspective allows pinpointing the limitations in interpretation of results and the need of further experiments and/or controls to establish “definitive evidence” for the role of protein degradation in the proteostasis of a given protein or the entire proteome. We also provide a spreadsheet for simple calculations of mRNA and protein decays for mimicking different experimental conditions and a checklist for the analysis of experiments dealing with protein degradation studies that may be useful for researchers interested in the area of protein turnover.

Oxygen versus Reactive Oxygen in the Regulation of HIF-1: The Balance Tips

Thilo Hagen — Tue, 09 Oct 2012 08:38:36 +0000

Hypoxia inducible factor (HIF) is known as the master regulator of the cellular response to hypoxia and is of pivotal importance during development as well as in human disease, particularly in cancer. It is composed of a constitutively expressed β subunit (HIF-1β) and an oxygen-regulated α subunit (HIF-1α and HIF-2α), whose stability is tightly controlled by a family of oxygen- and iron-dependent prolyl hydroxylase enzymes. Whether or not mitochondria-derived reactive oxygen species (ROS) are involved in the regulation of Hypoxia Inducible Factor-1α has been a matter of contention for the last 10 years, with equally compelling evidence in favor and against their contribution. A number of recent papers appear to tip the balance against a role for ROS. Thus, it has been demonstrated that HIF prolyl hydroxylases are unlikely to be physiological targets of ROS and that the increase in ROS that is associated with downregulation of Thioredoxin Reductase in hypoxia does not affect HIF-1α stabilization. Finally, the protein CHCHD4, which modulates cellular HIF-1α concentrations by promoting mitochondrial electron transport chain activity, has been proposed to exert its regulatory effect by affecting cellular oxygen availability. These reports are consistent with the hypothesis that mitochondria play a critical role in the regulation of HIF-1α by controlling intracellular oxygen concentrations.

The Ubiquitin-Proteasome System in Huntington’s Disease: Are Proteasomes Impaired, Initiators of Disease, or Coming to the Rescue?

Sabine Schipper-Krom, Katrin Juenemann, and Eric A. J. Reits — Mon, 24 Sep 2012 12:03:04 +0000

Huntington’s disease is a progressive neurodegenerative disease, caused by a polyglutamine expansion in the huntingtin protein. A prominent hallmark of the disease is the presence of intracellular aggregates initiated by N-terminal huntingtin fragments containing the polyglutamine repeat, which recruit components of the ubiquitin-proteasome system. While it is commonly thought that proteasomes are irreversibly sequestered into these aggregates leading to impairment of the ubiquitin-proteasome system, the data on proteasomal impairment in Huntington’s disease is contradictory. In addition, it has been suggested that proteasomes are unable to actually cleave polyglutamine sequences in vitro, thereby releasing aggregation-prone polyglutamine peptides in cells. Here, we discuss how the proteasome is involved in the various stages of polyglutamine aggregation in Huntington’s disease, and how alterations in activity may improve clearance of mutant huntingtin fragments.

Potential Role of Peptidylarginine Deiminase Enzymes and Protein Citrullination in Cancer Pathogenesis

Sun, 16 Sep 2012 08:59:58 +0000

The peptidylarginine deiminases (PADs) are a family of posttranslational modification enzymes that catalyze the conversion of positively charged protein-bound arginine and methylarginine residues to the uncharged, nonstandard amino acid citrulline. This enzymatic activity is referred to as citrullination or, alternatively, deimination. Citrullination can significantly affect biochemical pathways by altering the structure and function of target proteins. Five mammalian PAD family members (PADs 1–4 and 6) have been described and show tissue-specific distribution. Recent reviews on PADs have focused on their role in autoimmune diseases. Here, we will discuss the potential role of PADs in tumor progression and tumor-associated inflammation. In the context of cancer, increasing clinical evidence suggests that PAD4 (and possibly PAD2) has important roles in tumor progression. The link between PADs and cancer is strengthened by recent findings showing that treatment of cell lines and mice with PAD inhibitors significantly suppresses tumor growth and, interestingly, inflammatory symptoms. At the molecular level, transcription factors, coregulators, and histones are functional targets for citrullination by PADs, and citrullination of these targets can affect gene expression in multiple tumor cell lines. Next generation isozyme-specific PAD inhibitors may have therapeutic potential to regulate both the inflammatory tumor microenvironment and tumor cell growth.

Divalent Metal Ion Transport across Large Biological Ion Channels and Their Effect on Conductance and Selectivity

Elena García-Giménez, Antonio Alcaraz, and Vicente M. Aguilella — Thu, 13 Sep 2012 15:30:57 +0000

Electrophysiological characterization of large protein channels, usually displaying multi-ionic transport and weak ion selectivity, is commonly performed at physiological conditions (moderate gradients of KCl solutions at decimolar concentrations buffered at neutral pH). We extend here the characterization of the OmpF porin, a wide channel of the outer membrane of E. coli, by studying the effect of salts of divalent cations on the transport properties of the channel. The regulation of divalent cations concentration is essential in cell metabolism and understanding their effects is of key importance, not only in the channels specifically designed to control their passage but also in other multiionic channels. In particular, in porin channels like OmpF, divalent cations modulate the efficiency of molecules having antimicrobial activity. Taking advantage of the fact that the OmpF channel atomic structure has been resolved both in water and in MgCl2 aqueous solutions, we analyze the single channel conductance and the channel selectivity inversion aiming to separate the role of the electrolyte itself, and the counterion accumulation induced by the protein channel charges and other factors (binding, steric effects, etc.) that being of minor importance in salts of monovalent cations become crucial in the case of divalent cations.

Cardiomyopathy-Related Mutations in Cardiac Troponin C, L29Q and G159D, Have Divergent Effects on Rat Cardiac Myofiber Contractile Dynamics

Sampath K. Gollapudi and Murali Chandra — Wed, 12 Sep 2012 15:38:28 +0000

Previous studies of cardiomyopathy-related mutations in cardiac troponin C (cTnC)—L29Q and G159D—have shown diverse findings. The link between such mutant effects and their divergent impact on cardiac phenotypes has remained elusive due to lack of studies on contractile dynamics. We hypothesized that a cTnC mutant-induced change in the thin filament will affect global myofilament mechanodynamics because of the interactions of thin filament kinetics with both Ca2+ binding and crossbridge (XB) cycling kinetics. We measured pCa-tension relationship and contractile dynamics in detergent-skinned rat cardiac papillary muscle fibers reconstituted with the recombinant wild-type rat cTnC (cTnCWT), cTnCL29Q, and cTnCG159D mutants. cTnCL29Q fibers demonstrated a significant decrease in Ca2+ sensitivity, but cTnCG159D fibers did not. Both mutants had no effect on Ca2+-activated maximal tension. The rate of XB recruitment dynamics increased in cTnCL29Q (26%) and cTnCG159D (25%) fibers. The rate of XB distortion dynamics increased in cTnCG159D fibers (15%). Thus, the cTnCL29Q mutant modulates the equilibrium between the non-cycling and cycling pool of XB by affecting the on/off kinetics of the regulatory units (Tropomyosin-Troponin); whereas, the cTnCG159D mutant increases XB cycling rate. Different effects on contractile dynamics may offer clue regarding how cTnCL29Q and cTnCG159D cause divergent effects on cardiac phenotypes.

Ubiquitin-Mediated Regulation of Endocytosis by Proteins of the Arrestin Family

Tue, 04 Sep 2012 17:39:58 +0000

In metazoans, proteins of the arrestin family are key players of G-protein-coupled receptors (GPCRS) signaling and trafficking. Following stimulation, activated receptors are phosphorylated, thus allowing the binding of arrestins and hence an “arrest” of receptor signaling. Arrestins act by uncoupling receptors from G proteins and contribute to the recruitment of endocytic proteins, such as clathrin, to direct receptor trafficking into the endocytic pathway. Arrestins also serve as adaptor proteins by promoting the recruitment of ubiquitin ligases and participate in the agonist-induced ubiquitylation of receptors, known to have impact on their subcellular localization and stability. Recently, the arrestin family has expanded following the discovery of arrestin-related proteins in other eukaryotes such as yeasts or fungi. Surprisingly, most of these proteins are also involved in the ubiquitylation and endocytosis of plasma membrane proteins, thus suggesting that the role of arrestins as ubiquitin ligase adaptors is at the core of these proteins' functions. Importantly, arrestins are themselves ubiquitylated, and this modification is crucial for their function. In this paper, we discuss recent data on the intricate connections between arrestins and the ubiquitin pathway in the control of endocytosis.

Effect of Vitamins A, C, and E Supplementation in the Treatment of Metabolic Syndrome in Albino Rats

L. S. Bilbis, S. A. Muhammad, Y. Saidu, and Y. Adamu — Thu, 16 Aug 2012 17:04:49 +0000

Obesity and metabolic syndrome increase the risk of cardiovascular morbidity and mortality. Oxidative stress seems to be involved in the path physiology of cardiovascular complications of metabolic syndrome. In this study we investigated the effects of vitamins A, C, and E in the management of metabolic syndrome traits condition in albino rats fed with high salt diet. The rats were placed on 8% NaCl diet for 5 weeks and then supplemented with these vitamins for additional 4 weeks in the presence of salt diet. Supplementation with vitamins significantly (𝑃<0.01 ) decreased blood pressure of the rats as compared with the control. Supplementation also significantly (𝑃<0.05) reduced serum total cholesterol, triglyceride, low-density lipoprotein cholesterol, and very-low-density lipoprotein cholesterol and increased high-density lipoprotein cholesterol, and total antioxidant status as compared with untreated group. The percentage protection of the supplemented groups against atherogenesis indicated 55.50±3.75%. Percentage weight gain indicated significant positive correlation with triglyceride, insulin resistance, and malondialdehyde while total antioxidant status and nitric oxide showed significant negative correlation. Salt diet significantly (𝑃<0.05) induced features of metabolic syndrome. The result, therefore, indicated strong relationship between obesity and metabolic syndrome and underscores the role of these vitamins in the management of metabolic syndrome.

Mitochondria and Cytoprotection

Catherine Brenner, Renée Ventura-Clapier, and Etienne Jacotot — Thu, 16 Aug 2012 07:45:42 +0000

Genetic and Biochemical Alterations in Non-Small Cell Lung Cancer

Jackie L. Johnson, Smitha Pillai, and Srikumar P. Chellappan — Wed, 15 Aug 2012 10:04:27 +0000

Despite significant advances in the detection and treatment of lung cancer, it causes the highest number of cancer-related mortality. Recent advances in the detection of genetic alterations in patient samples along with physiologically relevant animal models has yielded a new understanding of the molecular etiology of lung cancer. This has facilitated the development of potent and specific targeted therapies, based on the genetic and biochemical alterations present in the tumor, especially non-small-cell lung cancer (NSCLC). It is now clear that heterogeneous cell signaling pathways are disrupted to promote NSCLC, including mutations in critical growth regulatory proteins (K-Ras, EGFR, B-RAF, MEK-1, HER2, MET, EML-4-ALK, KIF5B-RET, and NKX2.1) and inactivation of growth inhibitory pathways (TP53, PTEN, p16, and LKB-1). How these pathways differ between smokers and non-smokers is also important for clinical treatment strategies and development of targeted therapies. This paper describes these molecular targets in NSCLC, and describes the biological significance of each mutation and their potential to act as a therapeutic target.

Role of p16INK4A in Replicative Senescence and DNA Damage-Induced Premature Senescence in p53-Deficient Human Cells

Razmik Mirzayans, Bonnie Andrais, Gavin Hansen, and David Murray — Mon, 13 Aug 2012 11:26:05 +0000

The p16INK4A (hereafter p16) tumor suppressor is encoded by the INK4A/ARF locus which is among the most commonly dysregulated sequences in human cancer. By inhibiting cyclin-dependent kinases, p16 activates the G1-S checkpoint, and this response is often considered to be critical for establishing a senescence-like growth arrest. Not all studies support a universal role for p16 in senescence. Single-cell analysis of noncancerous human fibroblast cultures undergoing senescence as a function of culture age (replicative senescence) has revealed that p16 is not expressed in the majority (>90%) of cells that exhibit features of senescence (e.g., flattened and enlarged morphology coupled with senescence-associated β-galactosidase expression), ruling out a requirement for p16 in this process. In addition, ionizing radiation triggers premature senescence in human cancer cell lines that do not express p16. These observations are made with cells that express wild-type p53, a key mediator of the DNA damage response. In this paper, we examine the growing evidence suggesting a negative regulatory relationship between p16 and p53 and discuss recent reports that implicate a role for p16 in replicative senescence and ionizing radiation-induced premature senescence in human cells that lack wild-type p53 function.

Cardiomyopathy Classification: Ongoing Debate in the Genomics Era

Charles McCartan, Robert Mason, S. R. Jayasinghe, and Lyn R. Griffiths — Wed, 08 Aug 2012 10:30:20 +0000

Cardiomyopathies represent a group of diseases of the myocardium of the heart and include diseases both primarily of the cardiac muscle and systemic diseases leading to adverse effects on the heart muscle size, shape, and function. Traditionally cardiomyopathies were defined according to phenotypical appearance. Now, as our understanding of the pathophysiology of the different entities classified under each of the different phenotypes improves and our knowledge of the molecular and genetic basis for these entities progresses, the traditional classifications seem oversimplistic and do not reflect current understanding of this myriad of diseases and disease processes. Although our knowledge of the exact basis of many of the disease processes of cardiomyopathies is still in its infancy, it is important to have a classification system that has the ability to incorporate the coming tide of molecular and genetic information. This paper discusses how the traditional classification of cardiomyopathies based on morphology has evolved due to rapid advances in our understanding of the genetic and molecular basis for many of these clinical entities.

“Dead Cells Talking”: The Silent Form of Cell Death Is Not so Quiet

Richard Jäger and Howard O. Fearnhead — Tue, 07 Aug 2012 11:24:01 +0000

After more than twenty years of research, the molecular events of apoptotic cell death can be succinctly stated; different pathways, activated by diverse signals, increase the activity of proteases called caspases that rapidly and irreversibly dismantle condemned cell by cleaving specific substrates. In this time the ideas that apoptosis protects us from tumourigenesis and that cancer chemotherapy works by inducing apoptosis also emerged. Currently, apoptosis research is shifting away from the intracellular events within the dying cell to focus on the effect of apoptotic cells on surrounding tissues. This is producing counterintuitive data showing that our understanding of the role of apoptosis in tumourigenesis and cancer therapy is too simple, with some interesting and provocative implications. Here, we will consider evidence supporting the idea that dying cells signal their presence to the surrounding tissue and, in doing so, elicit repair and regeneration that compensates for any loss of function caused by cell death. We will discuss evidence suggesting that cancer cell proliferation may be driven by inappropriate or corrupted tissue-repair programmes that are initiated by signals from apoptotic cells and show how this may dramatically modify how we view the role of apoptosis in both tumourigenesis and cancer therapy.

Mitotic Kinases and p53 Signaling

Geun-Hyoung Ha and Eun-Kyoung Yim Breuer — Thu, 19 Jul 2012 15:12:58 +0000

Mitosis is tightly regulated and any errors in this process often lead to aneuploidy, genomic instability, and tumorigenesis. Deregulation of mitotic kinases is significantly associated with improper cell division and aneuploidy. Because of their importance during mitosis and the relevance to cancer, mitotic kinase signaling has been extensively studied over the past few decades and, as a result, several mitotic kinase inhibitors have been developed. Despite promising preclinical results, targeting mitotic kinases for cancer therapy faces numerous challenges, including safety and patient selection issues. Therefore, there is an urgent need to better understand the molecular mechanisms underlying mitotic kinase signaling and its interactive network. Increasing evidence suggests that tumor suppressor p53 functions at the center of the mitotic kinase signaling network. In response to mitotic spindle damage, multiple mitotic kinases phosphorylate p53 to either activate or deactivate p53-mediated signaling. p53 can also regulate the expression and function of mitotic kinases, suggesting the existence of a network of mutual regulation, which can be positive or negative, between mitotic kinases and p53 signaling. Therefore, deciphering this regulatory network will provide knowledge to overcome current limitations of targeting mitotic kinases and further improve the results of targeted therapy.

Transcriptional Regulation of the p53 Tumor Suppressor Gene in S-Phase of the Cell-Cycle and the Cellular Response to DNA Damage

David Reisman, Paula Takahashi, Amanda Polson, and Kristy Boggs — Wed, 11 Jul 2012 14:12:02 +0000

The p53 tumor suppressor induces the transcription of genes that negatively regulate progression of the cell cycle in response to DNA damage or other cellular stressors and thus participates in maintaining genome stability. Numerous studies have demonstrated that p53 transcription is activated before or during early S-phase in cells progressing from G0/G1 into S-phase through the combined action of two DNA-binding factors RBP-Jκ and C/EBPβ-2. Here, we review evidence that this induction occurs to provide available p53 mRNA in order to prepare the cell for DNA damage in S-phase, this ensuring a rapid response to DNA damage before exiting this stage of the cell cycle.

The Mutations Associated with Dilated Cardiomyopathy

Ruti Parvari and Aviva Levitas — Mon, 09 Jul 2012 11:40:49 +0000

Cardiomyopathy is an important cause of heart failure and a major indication for heart transplantation in children and adults. This paper describes the state of the genetic knowledge of dilated cardiomyopathy (DCM). The identification of the causing mutation is important since presymptomatic interventions of DCM have proven value in preventing morbidity and mortality. Additionally, as in general in genetic studies, the identification of the mutated genes has a direct clinical impact for the families and population involved. Identifying causative mutations immediately amplifies the possibilities for disease prevention through carrier screening and prenatal testing. This often lifts a burden of social isolation from affected families, since healthy family members can be assured of having healthy children. Identification of the mutated genes holds the potential to lead to the understanding of disease etiology, pathophysiology, and therefore potential therapy. This paper presents the genetic variations, or disease-causing mutations, contributing to the pathogenesis of hereditary DCM, and tries to relate these to the functions of the mutated genes.

Length and PKA Dependence of Force Generation and Loaded Shortening in Porcine Cardiac Myocytes

Kerry S. McDonald, Laurin M. Hanft, Timothy L. Domeier, and Craig A. Emter — Thu, 05 Jul 2012 15:41:31 +0000

In healthy hearts, ventricular ejection is determined by three myofibrillar properties; force, force development rate, and rate of loaded shortening (i.e., power). The sarcomere length and PKA dependence of these mechanical properties were measured in porcine cardiac myocytes. Permeabilized myocytes were prepared from left ventricular free walls and myocyte preparations were calcium activated to yield ~50% maximal force after which isometric force was measured at varied sarcomere lengths. Porcine myocyte preparations exhibited two populations of length-tension relationships, one being shallower than the other. Moreover, myocytes with shallow length-tension relationships displayed steeper relationships following PKA. Sarcomere length-𝐾tr relationships also were measured and 𝐾tr remained nearly constant over ~2.30 μm to ~1.90 μm and then increased at lengths below 1.90 μm. Loaded-shortening and peak-normalized power output was similar at ~2.30 μm and ~1.90 μm even during activations with the same [Ca2+], implicating a myofibrillar mechanism that sustains myocyte power at lower preloads. PKA increased myocyte power and yielded greater shortening-induced cooperative deactivation in myocytes, which likely provides a myofibrillar mechanism to assist ventricular relaxation. Overall, the bimodal distribution of myocyte length-tension relationships and the PKA-mediated changes in myocyte length-tension and power are likely important modulators of Frank-Starling relationships in mammalian hearts.

Combination Effects of Docetaxel and Doxorubicin in Hormone-Refractory Prostate Cancer Cells

Eleftheria Tsakalozou, Allison M. Eckman, and Younsoo Bae — Sun, 01 Jul 2012 15:49:05 +0000

Combination effects of docetaxel (DOC) and doxorubicin (DOX) were investigated in prostate cancer cells (PC3 and DU145). Combination indices (CIs) were determined using the unified theory in various concentrations and mixing ratios (synergy: CI<0.9, additivity: 0.91.1). DOC showed a biphasic cytotoxicity pattern with the half maximal inhibitory concentration (IC50) at the picomolar range for PC3 (0.598 nM) and DU145 (0.469 nM), following 72 h drug exposure. The IC50s of DOX were 908 nM and 343 nM for PC3 and DU145, respectively. Strong synergy was seen when PC3 was treated with DOC at concentrations lower than its IC50 values (0.125~0.5 nM) plus DOX (2~8 times IC50). Equipotent drug combination treatments (7×7) revealed that the DOC/DOX combination leads to high synergy and effective cell death only in a narrow concentration range in DU145. This study provides a convenient method to predict multiple drug combination effects by the estimated CI values as well as cell viability data. The proposed DOC/DOX mixing ratios can be used to design combination drug cocktails or delivery systems to improve chemotherapy for cancer patients.

Ubiquitin C-Terminal Hydrolase L1 in Tumorigenesis

Jennifer Hurst-Kennedy, Lih-Shen Chin, and Lian Li — Sun, 01 Jul 2012 11:24:00 +0000

Ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1, aka PGP9.5) is an abundant, neuronal deubiquitinating enzyme that has also been suggested to possess E3 ubiquitin-protein ligase activity and/or stabilize ubiquitin monomers in vivo. Recent evidence implicates dysregulation of UCH-L1 in the pathogenesis and progression of human cancers. Although typically only expressed in neurons, high levels of UCH-L1 have been found in many nonneuronal tumors, including breast, colorectal, and pancreatic carcinomas. UCH-L1 has also been implicated in the regulation of metastasis and cell growth during the progression of nonsmall cell lung carcinoma, colorectal cancer, and lymphoma. Together these studies suggest UCH-L1 has a potent oncogenic role and drives tumor development. Conversely, others have observed promoter methylation-mediated silencing of UCH-L1 in certain tumor subtypes, suggesting a potential tumor suppressor role for UCH-L1. In this paper, we provide an overview of the evidence supporting the involvement of UCH-L1 in tumor development and discuss the potential mechanisms of action of UCH-L1 in oncogenesis.

Strategies to Identify Recognition Signals and Targets of SUMOylation

Sun, 01 Jul 2012 08:31:48 +0000

SUMOylation contributes to the regulation of many essential cellular factors. Diverse techniques have been used to explore the functional consequences of protein SUMOylation. Most approaches consider the identification of sequences on substrates, adaptors, or receptors regulating the SUMO conjugation, recognition, or deconjugation. The large majority of the studied SUMOylated proteins contain the sequence [IVL]KxE. SUMOylated proteins are recognized by at least 3 types of hydrophobic SUMO-interacting motifs (SIMs) that contribute to coordinate SUMO-dependent functions. Typically, SIMs are constituted by a hydrophobic core flanked by one or two clusters of negatively charged amino acid residues. Multiple SIMs can integrate SUMO binding domains (SBDs), optimizing binding, and control over SUMO-dependent processes. Here, we present a survey of the methodologies used to study SUMO-regulated functions and provide guidelines for the identification of cis and trans sequences controlling SUMOylation. Furthermore, an integrative analysis of known and putative SUMO substrates illustrates an updated landscape of several SUMO-regulated events. The strategies and analysis presented here should contribute to the understanding of SUMO-controlled functions and provide rational approach to identify biomarkers or choose possible targets for intervention in processes where SUMOylation plays a critical role.

Regulator of G-Protein Signaling 5 Reduces HeyA8 Ovarian Cancer Cell Proliferation and Extends Survival in a Murine Tumor Model

Mon, 25 Jun 2012 11:08:59 +0000

The regulator of G-protein signaling 5 (RGS5) belongs to a family of GTPase activators that terminate signaling cascades initiated by extracellular mediators and G-protein-coupled receptors. RGS5 has an interesting dual biological role. One functional RGS5 role is as a pericyte biomarker influencing the switch to angiogenesis during malignant progression. Its other functional role is to promote apoptosis in hypoxic environments. We set out to clarify the extent to which RGS5 expression regulates tumor progression—whether it plays a pathogenic or protective role in ovarian tumor biology. We thus constructed an inducible gene expression system to achieve RGS5 expression in HeyA8-MDR ovarian cancer cells. Through this we observed that inducible RGS5 expression significantly reduces in vitro BrdU-positive HeyA8-MDR cells, although this did not correlate with a reduction in tumor volume observed using an in vivo mouse model of ovarian cancer. Interestingly, mice bearing RGS5-expressing tumors demonstrated an increase in survival compared with controls, which might be attributed to the vast regions of necrosis observed by pathological examination. Additionally, mice bearing RGS5-expressing tumors were less likely to have ulcerated tumors. Taken together, this data supports the idea that temporal expression and stabilization of RGS5 could be a valuable tactic within the context of a multicomponent approach for modulating tumor progression.

Endoplasmic Reticulum Stress and Lipid Metabolism

Huiping Zhou, Kezhong Zhang, Sabina Janciauskiene, and Xiaokun Li — Tue, 19 Jun 2012 10:07:53 +0000