Gintautas Tamulaitis, Migle Kazlauskiene, Elena Manakova, Česlovas Venclovas, Alison O Nwokeoji, Mark J Dickman, Philippe Horvath, Virginijus Siksnys
{"title":"嗜热链球菌III-A型CRISPR-Cas系统的可编程RNA切碎。","authors":"Gintautas Tamulaitis, Migle Kazlauskiene, Elena Manakova, Česlovas Venclovas, Alison O Nwokeoji, Mark J Dickman, Philippe Horvath, Virginijus Siksnys","doi":"10.1016/j.molcel.2014.09.027","DOIUrl":null,"url":null,"abstract":"<p><p>Immunity against viruses and plasmids provided by CRISPR-Cas systems relies on a ribonucleoprotein effector complex that triggers the degradation of invasive nucleic acids (NA). Effector complexes of type I (Cascade) and II (Cas9-dual RNA) target foreign DNA. Intriguingly, the genetic evidence suggests that the type III-A Csm complex targets DNA, whereas biochemical data show that the type III-B Cmr complex cleaves RNA. Here we aimed to investigate NA specificity and mechanism of CRISPR interference for the Streptococcus thermophilus Csm (III-A) complex (StCsm). When expressed in Escherichia coli, two complexes of different stoichiometry copurified with 40 and 72 nt crRNA species, respectively. Both complexes targeted RNA and generated multiple cuts at 6 nt intervals. The Csm3 protein, present in multiple copies in both Csm complexes, acts as endoribonuclease. In the heterologous E. coli host, StCsm restricts MS2 RNA phage in a Csm3 nuclease-dependent manner. Thus, our results demonstrate that the type III-A StCsm complex guided by crRNA targets RNA and not DNA. </p>","PeriodicalId":18950,"journal":{"name":"Molecular Cell","volume":null,"pages":null},"PeriodicalIF":14.5000,"publicationDate":"2014-11-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.molcel.2014.09.027","citationCount":"227","resultStr":"{\"title\":\"Programmable RNA shredding by the type III-A CRISPR-Cas system of Streptococcus thermophilus.\",\"authors\":\"Gintautas Tamulaitis, Migle Kazlauskiene, Elena Manakova, Česlovas Venclovas, Alison O Nwokeoji, Mark J Dickman, Philippe Horvath, Virginijus Siksnys\",\"doi\":\"10.1016/j.molcel.2014.09.027\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Immunity against viruses and plasmids provided by CRISPR-Cas systems relies on a ribonucleoprotein effector complex that triggers the degradation of invasive nucleic acids (NA). Effector complexes of type I (Cascade) and II (Cas9-dual RNA) target foreign DNA. Intriguingly, the genetic evidence suggests that the type III-A Csm complex targets DNA, whereas biochemical data show that the type III-B Cmr complex cleaves RNA. Here we aimed to investigate NA specificity and mechanism of CRISPR interference for the Streptococcus thermophilus Csm (III-A) complex (StCsm). When expressed in Escherichia coli, two complexes of different stoichiometry copurified with 40 and 72 nt crRNA species, respectively. Both complexes targeted RNA and generated multiple cuts at 6 nt intervals. The Csm3 protein, present in multiple copies in both Csm complexes, acts as endoribonuclease. In the heterologous E. coli host, StCsm restricts MS2 RNA phage in a Csm3 nuclease-dependent manner. Thus, our results demonstrate that the type III-A StCsm complex guided by crRNA targets RNA and not DNA. </p>\",\"PeriodicalId\":18950,\"journal\":{\"name\":\"Molecular Cell\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":14.5000,\"publicationDate\":\"2014-11-20\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1016/j.molcel.2014.09.027\",\"citationCount\":\"227\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Molecular Cell\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1016/j.molcel.2014.09.027\",\"RegionNum\":1,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2014/11/6 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q1\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular Cell","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/j.molcel.2014.09.027","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2014/11/6 0:00:00","PubModel":"Epub","JCR":"Q1","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
Programmable RNA shredding by the type III-A CRISPR-Cas system of Streptococcus thermophilus.
Immunity against viruses and plasmids provided by CRISPR-Cas systems relies on a ribonucleoprotein effector complex that triggers the degradation of invasive nucleic acids (NA). Effector complexes of type I (Cascade) and II (Cas9-dual RNA) target foreign DNA. Intriguingly, the genetic evidence suggests that the type III-A Csm complex targets DNA, whereas biochemical data show that the type III-B Cmr complex cleaves RNA. Here we aimed to investigate NA specificity and mechanism of CRISPR interference for the Streptococcus thermophilus Csm (III-A) complex (StCsm). When expressed in Escherichia coli, two complexes of different stoichiometry copurified with 40 and 72 nt crRNA species, respectively. Both complexes targeted RNA and generated multiple cuts at 6 nt intervals. The Csm3 protein, present in multiple copies in both Csm complexes, acts as endoribonuclease. In the heterologous E. coli host, StCsm restricts MS2 RNA phage in a Csm3 nuclease-dependent manner. Thus, our results demonstrate that the type III-A StCsm complex guided by crRNA targets RNA and not DNA.
期刊介绍:
Molecular Cell is a companion to Cell, the leading journal of biology and the highest-impact journal in the world. Launched in December 1997 and published monthly. Molecular Cell is dedicated to publishing cutting-edge research in molecular biology, focusing on fundamental cellular processes. The journal encompasses a wide range of topics, including DNA replication, recombination, and repair; Chromatin biology and genome organization; Transcription; RNA processing and decay; Non-coding RNA function; Translation; Protein folding, modification, and quality control; Signal transduction pathways; Cell cycle and checkpoints; Cell death; Autophagy; Metabolism.