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      CRISPR-Cas in Streptococcus pyogenes

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          ABSTRACT

          The discovery and characterization of the prokaryotic CRISPR-Cas immune system has led to a revolution in genome editing and engineering technologies. Despite the fact that most applications emerged after the discovery of the type II-A CRISPR-Cas9 system of Streptococcus pyogenes, its biological importance in this organism has received little attention. Here, we provide a comprehensive overview of the current knowledge about CRISPR-Cas systems from S. pyogenes. We discuss how the interplay between CRISPR-mediated immunity and horizontal gene transfer might have modeled the evolution of this pathogen. We review the current literature about the CRISPR-Cas systems present in S. pyogenes (types I-C and II-A), and describe their distinctive biochemical and functional features. Finally, we summarize the main biotechnological applications that have arisen from the discovery of the CRISPR-Cas9 system in S. pyogenes.

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          Epigenome editing by a CRISPR/Cas9-based acetyltransferase activates genes from promoters and enhancers

          Isaac B. Hilton, Anthony M. D’Ippolito, Christopher M. Vockley … (2015)
          Technologies that facilitate the targeted manipulation of epigenetic marks could be used to precisely control cell phenotype or interrogate the relationship between the epigenome and transcriptional control. Here we have generated a programmable acetyltransferase based on the CRISPR/Cas9 gene regulation system, consisting of the nuclease-null dCas9 protein fused to the catalytic core of the human acetyltransferase p300. This fusion protein catalyzes acetylation of histone H3 lysine 27 at its target sites, corresponding with robust transcriptional activation of target genes from promoters, proximal enhancers, and distal enhancers. Gene activation by the targeted acetyltransferase is highly specific across the genome. In contrast to conventional dCas9-based activators, the acetyltransferase effectively activates genes from enhancer regions and with individual guide RNAs. The core p300 domain is also portable to other programmable DNA-binding proteins. These results support targeted acetylation as a causal mechanism of transactivation and provide a new robust tool for manipulating gene regulation.
          Article 0 comments Cited 670 times – based on 0 reviews     Review now
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            Intervening sequences of regularly spaced prokaryotic repeats derive from foreign genetic elements.

            Francisco Mojica, César Díez-Villaseñor, Jesús García-Martínez … (2005)
            Prokaryotes contain short DN repeats known as CRISPR, recognizable by the regular spacing existing between the recurring units. They represent the most widely distributed family of repeats among prokaryotic genomes suggesting a biological function. The origin of the intervening sequences, at present unknown, could provide clues about their biological activities. Here we show that CRISPR spacers derive from preexisting sequences, either chromosomal or within transmissible genetic elements such as bacteriophages and conjugative plasmids. Remarkably, these extrachromosomal elements fail to infect the specific spacer-carrier strain, implying a relationship between CRISPR and immunity against targeted DNA. Bacteriophages and conjugative plasmids are involved in prokaryotic population control, evolution, and pathogenicity. All these biological traits could be influenced by the presence of specific spacers. CRISPR loci can be visualized as mosaics of a repeated unit, separated by sequences at some time present elsewhere in the cell.
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              Phage response to CRISPR-encoded resistance in Streptococcus thermophilus.

              Hélène Deveau, Rodolphe Barrangou, Josiane E Garneau … (2008)
              Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated genes are linked to a mechanism of acquired resistance against bacteriophages. Bacteria can integrate short stretches of phage-derived sequences (spacers) within CRISPR loci to become phage resistant. In this study, we further characterized the efficiency of CRISPR1 as a phage resistance mechanism in Streptococcus thermophilus. First, we show that CRISPR1 is distinct from previously known phage defense systems and is effective against the two main groups of S. thermophilus phages. Analyses of 30 bacteriophage-insensitive mutants of S. thermophilus indicate that the addition of one new spacer in CRISPR1 is the most frequent outcome of a phage challenge and that the iterative addition of spacers increases the overall phage resistance of the host. The added new spacers have a size of between 29 to 31 nucleotides, with 30 being by far the most frequent. Comparative analysis of 39 newly acquired spacers with the complete genomic sequences of the wild-type phages 2972, 858, and DT1 demonstrated that the newly added spacer must be identical to a region (named proto-spacer) in the phage genome to confer a phage resistance phenotype. Moreover, we found a CRISPR1-specific sequence (NNAGAAW) located downstream of the proto-spacer region that is important for the phage resistance phenotype. Finally, we show through the analyses of 20 mutant phages that virulent phages are rapidly evolving through single nucleotide mutations as well as deletions, in response to CRISPR1.
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                Author and article information

                Journal
                Journal ID (nlm-ta): RNA Biol
                Journal ID (iso-abbrev): RNA Biol
                Journal ID (publisher-id): KRNB
                Journal ID (publisher-id): krnb20
                Title: RNA Biology
                Publisher: Taylor & Francis
                ISSN (Print): 1547-6286
                ISSN (Electronic): 1555-8584
                Publication date Collection: April 2019
                Publication date (Electronic): 11 March 2019
                Publication date PMC-release: 11 March 2019
                Volume: 16
                Issue: 4
                Pages: 380-389
                Affiliations
                [a ]Max Planck Unit for the Science of Pathogens , Berlin, Germany
                [b ]Institute for Biology, Humboldt University , Berlin, Germany
                Author notes
                CONTACT Emmanuelle Charpentier research-charpentier@ 123456mpiib-berlin.mpg.de Max Planck Unit for the Science of Pathogens , Berlin, Germany
                Author information
                http://orcid.org/0000-0003-2211-2153
                http://orcid.org/0000-0002-0254-0778
                Article
                Publisher ID: 1582974
                DOI: 10.1080/15476286.2019.1582974
                PMC ID: 6546361
                PubMed ID: 30856357
                SO-VID: 5679dce2-1f9a-48e2-8768-2dcfd210f02a
                Copyright © © 2019 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
                License:

                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 work is properly cited.

                History
                Date received : 31 March 2018
                Date revision received : 6 February 2019
                Date accepted : 11 February 2019
                Page count
                Figures: 4, References: 122, Pages: 10
                Funding
                Funded by: German Research Foundation (DFG)
                Award ID: Leibniz Prize
                This work was supported by the Max-Planck-Gesellschaft;German Research Foundation (DFG) [Leibniz Prize].
                Categories
                Subject: Review

                ScienceOpen disciplines: Molecular biology
                Keywords: streptococcus pyogenes,mobile genetic elements,phages,adaptive immune system,crispr,cas9,genome editing
                Data availability:
                ScienceOpen disciplines: Molecular biology
                Keywords: streptococcus pyogenes, mobile genetic elements, phages, adaptive immune system, crispr, cas9, genome editing

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