On the Origin of Reverse Transcriptase-Using CRISPR-Cas Systems and Their Hyperdiverse, Enigmatic Spacer Repertoires.

Autor:	Silas S; Departments of Pathology and Genetics, Stanford University, Stanford, California, USA.; Department of Chemical and Systems Biology, Stanford University, Stanford, California, USA., Makarova KS; National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA., Shmakov S; National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA.; Skolkovo Institute of Science and Technology, Skolkovo, Russia., Páez-Espino D; Department of Energy, Joint Genome Institute, Walnut Creek, California, USA., Mohr G; Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas, USA., Liu Y; Department of Biomedical Informatics, Stanford University, Stanford, California, USA., Davison M; Department of Plant Biology, Carnegie Institution for Science, Stanford, California, USA., Roux S; Department of Microbiology, The Ohio State University, Columbus, Ohio, USA., Krishnamurthy SR; Departments of Molecular Microbiology and Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA., Fu BXH; Departments of Pathology and Genetics, Stanford University, Stanford, California, USA., Hansen LL; Departments of Pathology and Genetics, Stanford University, Stanford, California, USA., Wang D; Departments of Molecular Microbiology and Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA., Sullivan MB; Department of Microbiology, The Ohio State University, Columbus, Ohio, USA.; Department of Civil, Environmental and Geodetic Engineering, The Ohio State University, Columbus, Ohio, USA., Millard A; Microbiology and Infection Unit, Warwick Medical School, University of Warwick, Warwick, United Kingdom., Clokie MR; Department Infection, Immunity and Inflammation, University of Leicester, Leicester, United Kingdom., Bhaya D; Department of Plant Biology, Carnegie Institution for Science, Stanford, California, USA., Lambowitz AM; Department of Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas, USA., Kyrpides NC; Department of Energy, Joint Genome Institute, Walnut Creek, California, USA., Koonin EV; National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA., Fire AZ; Departments of Pathology and Genetics, Stanford University, Stanford, California, USA afire@stanford.edu.
Jazyk:	angličtina
Zdroj:	MBio [mBio] 2017 Jul 11; Vol. 8 (4). Date of Electronic Publication: 2017 Jul 11.
DOI:	10.1128/mBio.00897-17
Abstrakt:	Cas1 integrase is the key enzyme of the clustered regularly interspaced short palindromic repeat (CRISPR)-Cas adaptation module that mediates acquisition of spacers derived from foreign DNA by CRISPR arrays. In diverse bacteria, the cas1 gene is fused (or adjacent) to a gene encoding a reverse transcriptase (RT) related to group II intron RTs. An RT-Cas1 fusion protein has been recently shown to enable acquisition of CRISPR spacers from RNA. Phylogenetic analysis of the CRISPR-associated RTs demonstrates monophyly of the RT-Cas1 fusion, and coevolution of the RT and Cas1 domains. Nearly all such RTs are present within type III CRISPR-Cas loci, but their phylogeny does not parallel the CRISPR-Cas type classification, indicating that RT-Cas1 is an autonomous functional module that is disseminated by horizontal gene transfer and can function with diverse type III systems. To compare the sequence pools sampled by RT-Cas1-associated and RT-lacking CRISPR-Cas systems, we obtained samples of a commercially grown cyanobacterium- Arthrospira platensis Sequencing of the CRISPR arrays uncovered a highly diverse population of spacers. Spacer diversity was particularly striking for the RT-Cas1-containing type III-B system, where no saturation was evident even with millions of sequences analyzed. In contrast, analysis of the RT-lacking type III-D system yielded a highly diverse pool but reached a point where fewer novel spacers were recovered as sequencing depth was increased. Matches could be identified for a small fraction of the non-RT-Cas1-associated spacers, and for only a single RT-Cas1-associated spacer. Thus, the principal source(s) of the spacers, particularly the hypervariable spacer repertoire of the RT-associated arrays, remains unknown. IMPORTANCE While the majority of CRISPR-Cas immune systems adapt to foreign genetic elements by capturing segments of invasive DNA, some systems carry reverse transcriptases (RTs) that enable adaptation to RNA molecules. From analysis of available bacterial sequence data, we find evidence that RT-based RNA adaptation machinery has been able to join with CRISPR-Cas immune systems in many, diverse bacterial species. To investigate whether the abilities to adapt to DNA and RNA molecules are utilized for defense against distinct classes of invaders in nature, we sequenced CRISPR arrays from samples of commercial-scale open-air cultures of Arthrospira platensis , a cyanobacterium that contains both RT-lacking and RT-containing CRISPR-Cas systems. We uncovered a diverse pool of naturally occurring immune memories, with the RT-lacking locus acquiring a number of segments matching known viral or bacterial genes, while the RT-containing locus has acquired spacers from a distinct sequence pool for which the source remains enigmatic. (Copyright © 2017 Silas et al.)
Databáze:	MEDLINE
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