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Programmable type III-A CRISPR-Cas DNA targeting modules

The CRISPR-Cas systems provide invader defense in a wide variety of prokaryotes, as well as technologies for many powerful applications. The Type III-A or Csm CRISPR-Cas system is one of the most widely distributed across prokaryotic phyla, and cleaves targeted DNA and RNA molecules. In this work, w...

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Autores principales: Ichikawa, H. Travis, Cooper, John C., Lo, Leja, Potter, Jason, Terns, Rebecca M., Terns, Michael P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5404769/
https://www.ncbi.nlm.nih.gov/pubmed/28441427
http://dx.doi.org/10.1371/journal.pone.0176221
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author Ichikawa, H. Travis
Cooper, John C.
Lo, Leja
Potter, Jason
Terns, Rebecca M.
Terns, Michael P.
author_facet Ichikawa, H. Travis
Cooper, John C.
Lo, Leja
Potter, Jason
Terns, Rebecca M.
Terns, Michael P.
author_sort Ichikawa, H. Travis
collection PubMed
description The CRISPR-Cas systems provide invader defense in a wide variety of prokaryotes, as well as technologies for many powerful applications. The Type III-A or Csm CRISPR-Cas system is one of the most widely distributed across prokaryotic phyla, and cleaves targeted DNA and RNA molecules. In this work, we have constructed modules of Csm systems from 3 bacterial species and heterologously expressed the functional modules in E. coli. The modules include a Cas6 protein and a CRISPR locus for crRNA production, and Csm effector complex proteins. The expressed modules from L. lactis, S. epidermidis and S. thermophilus specifically eliminate invading plasmids recognized by the crRNAs of the systems. Characteristically, activation of plasmid targeting activity depends on transcription of the plasmid sequence recognized by the crRNA. Activity was not observed when transcription of the crRNA target sequence was blocked, or when the opposite strand or a non-target sequence was transcribed. Moreover, the Csm module can be programmed to recognize plasmids with novel target sequences by addition of appropriate crRNA coding sequences to the module. These systems provide a platform for investigation of Type III-A CRISPR-Cas systems in E. coli, and for introduction of programmable transcription-activated DNA targeting into novel organisms.
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spelling pubmed-54047692017-05-12 Programmable type III-A CRISPR-Cas DNA targeting modules Ichikawa, H. Travis Cooper, John C. Lo, Leja Potter, Jason Terns, Rebecca M. Terns, Michael P. PLoS One Research Article The CRISPR-Cas systems provide invader defense in a wide variety of prokaryotes, as well as technologies for many powerful applications. The Type III-A or Csm CRISPR-Cas system is one of the most widely distributed across prokaryotic phyla, and cleaves targeted DNA and RNA molecules. In this work, we have constructed modules of Csm systems from 3 bacterial species and heterologously expressed the functional modules in E. coli. The modules include a Cas6 protein and a CRISPR locus for crRNA production, and Csm effector complex proteins. The expressed modules from L. lactis, S. epidermidis and S. thermophilus specifically eliminate invading plasmids recognized by the crRNAs of the systems. Characteristically, activation of plasmid targeting activity depends on transcription of the plasmid sequence recognized by the crRNA. Activity was not observed when transcription of the crRNA target sequence was blocked, or when the opposite strand or a non-target sequence was transcribed. Moreover, the Csm module can be programmed to recognize plasmids with novel target sequences by addition of appropriate crRNA coding sequences to the module. These systems provide a platform for investigation of Type III-A CRISPR-Cas systems in E. coli, and for introduction of programmable transcription-activated DNA targeting into novel organisms. Public Library of Science 2017-04-25 /pmc/articles/PMC5404769/ /pubmed/28441427 http://dx.doi.org/10.1371/journal.pone.0176221 Text en © 2017 Ichikawa et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Ichikawa, H. Travis
Cooper, John C.
Lo, Leja
Potter, Jason
Terns, Rebecca M.
Terns, Michael P.
Programmable type III-A CRISPR-Cas DNA targeting modules
title Programmable type III-A CRISPR-Cas DNA targeting modules
title_full Programmable type III-A CRISPR-Cas DNA targeting modules
title_fullStr Programmable type III-A CRISPR-Cas DNA targeting modules
title_full_unstemmed Programmable type III-A CRISPR-Cas DNA targeting modules
title_short Programmable type III-A CRISPR-Cas DNA targeting modules
title_sort programmable type iii-a crispr-cas dna targeting modules
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5404769/
https://www.ncbi.nlm.nih.gov/pubmed/28441427
http://dx.doi.org/10.1371/journal.pone.0176221
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