Although hyperthermophilic archaea arguably have a great need for efficient DNA repair, they lack members of several DNA repair protein families broadly conserved among bacteria and eukaryotes. Conversely, the putative DNA repair genes that do occur in these archaea often do not generate the expected phenotype when deleted. The prospect that hyperthermophilic archaea have some unique strategies for coping with DNA damage and replication errors has intellectual and technological appeal, but resolving this question will require alternative coping mechanisms to be proposed and tested experimentally. This review evaluates a combination of four enigmatic properties that distinguishes the hyperthermophilic archaea from all other organisms: DNA polymerase stalling at dU, apparent lack of conventional NER, lack of MutSL homologs, and apparent essentiality of homologous recombination proteins. Hypothetical damage-coping strategies that could explain this set of properties may provide new starting points for efforts to define how archaea differ from conventional models of DNA repair and replication fidelity.

Dennis W. Grogan

Understanding DNA Repair in Hyperthermophilic Archaea: Persistent Gaps and Other Reasons to Focus on the Fork

Our understanding of the genus Natrinema is presently limited due to the lack of available genetic tools. Auxotrophic markers have been widely used to construct genetic systems in bacteria and eukaryotes and in some archaeal species. Here, we isolated four auxotrophic mutants of Natrinema sp. J7-2, via 1-methyl-3-nitro-1-nitroso-guanidin mutagenesis, and designated them as J7-2-1, J7-2-22, J7-2-26, and J7-2-52, respectively. The mutant phenotypes were determined to be auxotrophic for leucine (J7-2-1), arginine (J7-2-22 and J7-2-52), and lysine (J7-2-26). The complete genome and the biosynthetic pathways of amino acids in J7-2 identified that the auxotrophic phenotype of three mutants was due to gene mutations in leuB (J7-2-1), dapD (J7-2-26), and argC (J7-2-52). These auxotrophic phenotypes were employed as selectable makers to establish a transformation method. The transformation efficiencies were determined to be approximately 103 transformants per <svg xmlns:xlink="http://www.w3.org/1999/xlink" xmlns="http://www.w3.org/2000/svg" style="vertical-align:-3.40335pt" id="M1" height="9.4034pt" version="1.1" viewBox="-0.0498162 -6.00005 7.4591 9.4034" width="7.4591pt"><g transform="matrix(.013,0,0,-0.013,0,0)"><path id="g50-234" d="M546 97L529 123C499 91 459 65 450 65C443 65 439 73 445 106C468 226 496 341 523 451H513L436 424C423 357 412 311 403 276C356 195 245 59 190 59C165 59 158 99 166 135L236 445L227 451L152 428L60 18C43 -58 24 -145 24 -183C24 -237 43 -257 66 -257C96 -257 123 -236 130 -224L129 -216C115 -205 96 -156 96 -102C96 -57 105 -7 113 26H115C127 -1 137 -12 158 -12S198 -5 230 19C284 60 332 117 385 186H387C381 154 378 142 370 107C347 1 367 -12 388 -12C422 -12 492 32 546 97Z"/></g></svg>g DNA. And strains J7-2-1 and J7-2-26 were transformed into prototrophic strains with the wild type genomic DNA, amplified fragments of the corresponding genes, or the integrative plasmids carrying the corresponding genes. Additionally, exogenous genes, bgaH or amyH gene, were expressed successfully in J7-2-1. Thus, we have developed a genetic manipulation system for the Natrinema genus based on the isolated auxotrophic mutants of Natrinema sp. J7-2.

Jie Lv

Shuai Wang

Yuchen Wang

Yuping Huang

Xiangdong Chen

Isolation and Molecular Identification of Auxotrophic Mutants to Develop a Genetic Manipulation System for the Haloarchaeon Natrinema sp. J7-2

In this study the archaeal communities in extreme saline-alkaline soils of the former lake Texcoco, Mexico, with electrolytic conductivities (EC) ranging from 0.7 to 157.2&#x2009;dS/m and pH from 8.5 to 10.5 were explored. Archaeal communities in the 0.7&#x2009;dS/m pH 8.5 soil had the lowest alpha diversity values and were dominated by a limited number of phylotypes belonging to the mesophilic Candidatus Nitrososphaera. Diversity and species richness were higher in the soils with EC between 9.0 and 157.2&#x2009;dS/m. The majority of OTUs detected in the hypersaline soil were members of the Halobacteriaceae family. Novel phylogenetic branches in the Halobacteriales class were detected in the soil, and more abundantly in soil with the higher pH (10.5), indicating that unknown and uncharacterized Archaea can be found in this soil. Thirteen different genera of the Halobacteriaceae family were identified and were distributed differently between the soils. Halobiforma, Halostagnicola, Haloterrigena, and Natronomonas were found in all soil samples. Methanogenic archaea were found only in soil with pH between 10.0 and 10.3. Retrieved methanogenic archaea belonged to the Methanosarcinales and Methanomicrobiales orders. The comparison of the archaeal community structures considering phylogenetic information (UniFrac distances) clearly clustered the communities by pH.

Yendi E. Navarro-Noya

C&#xe9;sar Valenzuela-Encinas

Alonso Sandoval-Yuriar

Norma G. Jim&#xe9;nez-Bueno

Rodolfo Marsch

Luc Dendooven

Archaeal Communities in a Heterogeneous Hypersaline-Alkaline Soil

Guillaume Borrel

Nadia Gaci

Pierre Peyret

Paul W. O&#x2019;Toole

Simonetta Gribaldo

Jean-Fran&#xe7;ois Brug&#xe8;re

Corrigendum to &#x201c;Unique Characteristics of the Pyrrolysine System in the 7th Order of Methanogens: Implications for the Evolution of a Genetic Code Expansion Cassette&#x201d;

Hypersaline systems near salt saturation levels represent an extreme environment, in which organisms grow and survive near the limits of life. One of the abundant members of the microbial communities in hypersaline systems is the square archaeon, Haloquadratum walsbyi. Utilizing a short-read metagenome from Lake Tyrrell, a hypersaline ecosystem in Victoria, Australia, we performed a comparative genomic analysis of H. walsbyi to better understand the extent of variation between strains/subspecies. Results revealed that previously isolated strains/subspecies do not fully describe the complete repertoire of the genomic landscape present in H. walsbyi. Rearrangements, insertions, and deletions were observed for the Lake Tyrrell derived Haloquadratum genomes and were supported by environmental de novo sequences, including shifts in the dominant genomic landscape of the two most abundant strains. Analysis pertaining to halomucins indicated that homologs for this large protein are not a feature common for all species of Haloquadratum. Further, we analyzed ATP-binding cassette transporters (ABC-type transporters) for evidence of niche partitioning between different strains/subspecies. We were able to identify unique and variable transporter subunits from all five genomes analyzed and the de novo environmental sequences, suggesting that differences in nutrient and carbon source acquisition may play a role in maintaining distinct strains/subspecies.

Benjamin J. Tully

Joanne B. Emerson

Karen Andrade

Jochen J. Brocks

Eric E. Allen

Jillian F. Banfield

Karla B. Heidelberg

De Novo Sequences of Haloquadratum walsbyi from Lake Tyrrell, Australia, Reveal a Variable Genomic Landscape

Halococcus hamelinensis was the first archaeon isolated from stromatolites. These geomicrobial ecosystems are thought to be some of the earliest known on Earth, yet, despite their evolutionary significance, the role of Archaea in these systems is still not well understood. Detailed here is the genome sequencing and analysis of an archaeon isolated from stromatolites. The genome of H. hamelinensis consisted of 3,133,046 base pairs with an average G+C content of 60.08&#x25; and contained 3,150 predicted coding sequences or ORFs, 2,196 (68.67&#x25;) of which were protein-coding genes with functional assignments and 954 (29.83&#x25;) of which were of unknown function. Codon usage of the H. hamelinensis genome was consistent with a highly acidic proteome, a major adaptive mechanism towards high salinity. Amino acid transport and metabolism, inorganic ion transport and metabolism, energy production and conversion, ribosomal structure, and unknown function COG genes were overrepresented. The genome of H. hamelinensis also revealed characteristics reflecting its survival in its extreme environment, including putative genes/pathways involved in osmoprotection, oxidative stress response, and UV damage repair. Finally, genome analyses indicated the presence of putative transposases as well as positive matches of genes of H. hamelinensis against various genomes of Bacteria, Archaea, and viruses, suggesting the potential for horizontal gene transfer.

Reema K. Gudhka

Brett A. Neilan

Brendan P. Burns

Adaptation, Ecology, and Evolution of the Halophilic Stromatolite Archaeon Halococcus hamelinensis Inferred through Genome Analyses

A gene encoding a cyclodextrinase from Thermococcus kodakarensis KOD1 (CDase-Tk) was identified and characterized. The gene encodes a protein of 656 amino acid residues with a molecular mass of 76.4&#x2009;kDa harboring four conserved regions found in all members of the &#x3b1;-amylase family. A recombinant form of the enzyme was purified by ion-exchange chromatography, and its catalytic properties were examined. The enzyme was active in a broad range of pH conditions (pHs 4.0&#x2013;10.0), with an optimal pH of 7.5 and a temperature optimum of 65&#xb0;C. The purified enzyme preferred to hydrolyze &#x3b2;-cyclodextrin (CD) but not &#x3b1;- or &#x3b3;-CD, soluble starch, or pullulan. The final product from &#x3b2;-CD was glucose. The <svg xmlns:xlink="http://www.w3.org/1999/xlink" xmlns="http://www.w3.org/2000/svg" style="vertical-align:-3.408751pt" id="M1" height="12.4067pt" version="1.1" viewBox="-0.0498162 -8.99795 24.7073 12.4067" width="24.7073pt"><g transform="matrix(.0135,0,0,-0.0135,0,0)"><path id="g113-87" d="M697 650H468L461 623L492 619C539 613 547 605 518 546C481 471 367 264 278 116H276C239 278 197 500 186 567C180 604 185 613 226 619L252 623L260 650H24L17 623C78 617 92 613 108 533L216 -11H247C365 200 515 462 560 529C616 612 624 615 689 623L697 650Z"/></g><g transform="matrix(.0095,0,0,-0.0095,7.641,3.264)"><path id="g190-110" d="M797 0V26C739 32 732 36 732 103V296C732 394 682 449 605 449C576 449 550 437 529 423C504 407 475 389 446 366C425 418 382 449 334 449C303 449 279 437 253 421C222 403 201 385 180 371V452C135 432 85 419 41 411V388C99 379 102 374 102 310V103C102 38 93 32 27 26V0H238V26C189 32 180 38 180 103V338C210 363 250 390 289 390C351 390 377 348 377 275V103C377 37 368 32 306 26V0H520V26C465 32 456 38 456 101V296C456 314 455 326 453 338C491 369 529 390 565 390C628 390 653 345 653 274V107C653 36 642 32 583 26V0H797Z"/></g><g transform="matrix(.0095,0,0,-0.0095,15.38,3.264)"><path id="g190-98" d="M433 39L423 65C413 59 399 54 387 54C370 54 352 69 352 114V299C352 352 342 392 307 422C285 440 255 449 225 449C168 437 102 399 75 379C56 365 44 353 44 339C44 315 69 296 87 296C101 296 111 303 116 319C124 349 133 371 145 385C156 397 171 404 190 404C241 404 275 364 275 291V274C253 256 180 229 120 209C65 190 39 159 39 110C39 47 88 -12 159 -12C189 -12 237 25 277 52C282 35 288 21 301 8C312 -3 333 -12 348 -12L433 39ZM275 84C256 65 221 48 195 48C164 48 124 73 124 124C124 161 146 180 185 198C206 208 254 229 275 240V84Z"/></g><g transform="matrix(.0095,0,0,-0.0095,19.543,3.264)"><path id="g190-121" d="M474 0V26C414 34 401 43 364 100L267 248C300 297 324 332 345 358C381 400 394 405 455 411V437H272V411C316 406 323 401 305 370C287 337 267 306 247 276L188 369C169 397 173 405 215 411V437H16V411C71 404 83 396 114 348L201 212C171 167 144 127 116 92C77 42 66 34 4 26V0H190V26C139 34 136 43 156 77C175 113 198 150 220 183L294 66C311 39 302 31 260 26V0H474Z"/></g></svg> and <svg xmlns:xlink="http://www.w3.org/1999/xlink" xmlns="http://www.w3.org/2000/svg" style="vertical-align:-3.427751pt" id="M2" height="12.4257pt" version="1.1" viewBox="-0.0498162 -8.99795 17.916 12.4257" width="17.916pt"><g transform="matrix(.0135,0,0,-0.0135,0,0)"><path id="g113-76" d="M743 650H503L496 622L527 618C563 613 564 603 532 573C449 495 371 431 323 392C301 374 272 355 246 346L280 522C297 609 300 614 379 622L385 650H135L129 622C209 614 215 609 198 522L124 133C106 39 99 35 23 28L17 0H271L277 28C193 35 192 39 208 133L239 316C264 328 280 325 303 288C368 183 435 90 502 0H652L659 28C602 34 584 43 543 94C495 154 403 283 347 369L574 554C634 603 659 612 735 624L743 650Z"/></g><g transform="matrix(.0095,0,0,-0.0095,9.619,3.264)"><path id="g50-110" d="M781 91L765 118C733 86 702 70 693 70C686 70 685 77 692 106L738 297C772 439 736 451 712 451S675 445 647 431C605 410 526 356 452 257H450L459 294C491 426 459 451 429 451C403 451 386 445 361 431C312 404 239 343 170 253H168L188 333C208 411 205 451 175 451C148 451 85 417 24 346L39 318C57 338 100 374 112 374C120 374 122 372 115 339C91 226 62 112 28 -6L37 -12C59 -4 87 3 114 6C125 68 142 127 154 168C184 227 316 383 372 383C397 383 396 366 381 289C362 192 338 92 312 -6L317 -12C339 -5 367 3 396 6L434 170C468 234 598 383 650 383C668 383 676 369 664 319L609 90C593 23 600 -12 629 -12C654 -12 722 23 781 91Z"/></g></svg> values were 3.13&#x2009;&#xb1;&#x2009;0.47&#x2009;U&#x2009;mg&#x2212;1 and 2.94&#x2009;&#xb1;&#x2009;0.16&#x2009;mg&#x2009;mL&#x2212;1 for &#x3b2;-CD. The unique characteristics of CDase-Tk with a low catalytic temperature and substrate specificity are discussed, and the starch utilization pathway in a broad range of temperatures is also proposed.

Ying Sun

Xiaomin Lv

Zhengqun Li

Jiaqiang Wang

Baolei Jia

Jinliang Liu

Recombinant Cyclodextrinase from Thermococcus kodakarensis KOD1: Expression, Purification, and Enzymatic Characterization

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Archaea publishes original research articles as well as review articles dealing with all aspects of archaea, providing a unique venue for exchanging information about these extraordinary prokaryotes.

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