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Virulence regulates and boosts CRISPR-Cas9 immunity in Group B Streptococcus [RNA-seq]

ABSTRACT: Bacterial CRISPR-Cas9 immune systems protect against foreign DNA. However, immune efficiency is constrained by Cas9 off-target effects and toxicity. Here, we demonstrate that CRISPR-Cas9 immunity is regulated by CovR, the major regulator of virulence in Group B Streptococcus, a pathobiont responsible for neonatal invasive infections. We show that CovR binds to and represses a distal promoter of the cas operon, embedding immunity in the virulence regulatory network. Releasing CovR repression enhances immune efficiency against suboptimal spacers, originating from old immune memory or mutations, thereby transiently expanding the sequence space recognized and cleaved by Cas9. Furthermore, CovR inactivation promotes the acquisition of new spacers, enhancing immune memory. CovR-mediated immune regulation is conserved at the species level, with lineage-specific variability in the constitutive cas promoter and Cas9 variants, suggesting different evolutionary trajectories. Overall, we describe a coordinated regulatory mechanism between immunity and virulence that enhances the immune repertoire during acute infection phases.

ORGANISM(S): Streptococcus agalactiae

PROVIDER: GSE269472 | GEO | 2024/11/04

REPOSITORIES: GEO

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Virulence regulates and boosts CRISPR-Cas9 immunity in Group B Streptococcus [CrispRseq]

Project description:Bacterial CRISPR-Cas9 immune systems protect against foreign DNA. However, immune efficiency is constrained by Cas9 off-target effects and toxicity. Here, we demonstrate that CRISPR-Cas9 immunity is regulated by CovR, the major regulator of virulence in Group B Streptococcus, a pathobiont responsible for neonatal invasive infections. We show that CovR binds to and represses a distal promoter of the cas operon, embedding immunity in the virulence regulatory network. Releasing CovR repression enhances immune efficiency against suboptimal spacers, originating from old immune memory or mutations, thereby transiently expanding the sequence space recognized and cleaved by Cas9. Furthermore, CovR inactivation promotes the acquisition of new spacers, enhancing immune memory. CovR-mediated immune regulation is conserved at the species level, with lineage-specific variability in the constitutive cas promoter and Cas9 variants, suggesting different evolutionary trajectories. Overall, we describe a coordinated regulatory mechanism between immunity and virulence that enhances the immune repertoire during acute infection phases.

2024-11-04 | GSE269471 | GEO

Processing-Independent CRISPR RNAs Limit Natural Transformation in Neisseria meningitidis

Project description:In bacteria and archaea, CRISPR loci confer adaptive, sequence-based immunity against viruses and plasmids. CRISPR interference is specified by CRISPR RNAs (crRNAs) that are transcribed and processed from CRISPR spacers and repeats. Pre-crRNA processing is essential for CRISPR interference in all systems studied thus far. Here we examine crRNA biogenesis and CRISPR interference in naturally competent Neisseria spp., including the human pathogen N. meningitidis. Our studies reveal a unique crRNA maturation pathway in which crRNA transcription is driven by promoters that are embedded within each repeat, yielding crRNA 5’ ends are not formed by processing. Although crRNA 3’ end formation occurs through RNase III cleavage of a pre-crRNA/tracrRNA duplex, as in other Type II CRISPR systems, this processing event is dispensable for interference. The meningococcal pathway is the most streamlined CRISPR/cas system characterized to date. Endogenous CRISPR spacers frequently target genomic sequences of other Neisseria strains and so limit natural transformation, which is the primary source of genetic variation that contributes to immune evasion, antibiotic resistance, and virulence in N. meningitidis. dRNA-seq approach for RNA samples from cultures of N. lactamica 020-06, harvested at mid-log. Two cDNA libraries from total RNA were prepared to distinguish between transcripts with either primary orprocessed 5’ ends: one library is generated from untreated RNA, whereas the other is treated with terminator exonuclease (TEX),

2013-04-23 | E-GEOD-44582 | biostudies-arrayexpress

Processing-Independent CRISPR RNAs Limit Natural Transformation in Neisseria meningitidis

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Casposase structure and the mechanistic link between DNA transposition and spacer acquisition by CRISPR-Cas

Project description:Key to CRISPR-Cas adaptive immunity is maintaining an ongoing record of invading nucleic acids that are encountered, a process carried out by the Cas1-Cas2 complex that integrates short segments of foreign genetic material (spacers) into the CRISPR locus. It is hypothesized that Cas1 evolved from casposases, a novel class of transposases. We show here that casposase integration in vitro recapitulates several properties of CRISPR-Cas integrases. The X-ray structure of Methanosarcina mazei casposase bound to DNA representing the product of integration reveals a tetramer with target DNA bound snugly between two dimers in which single-stranded casposon end binding resembles that of spacer 3'-overhangs. The differences between transposase and CRISPR-Cas integrase are largely architectural, and it appears that evolutionary change involved changes in protein-protein interactions to favor Cas2 binding over tetramerization and the separation of Cas1 dimers. This, in turn, led to preferred integration of single spacers over two transposon ends.

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Hyperphosphorylation of the Group A Streptococcal Control of Virulence Regulator Increases Promoter Occupancy Specifically at Virulence Factor Encoding Genes

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CovS Inactivation Reduces CovR Promoter Binding at Diverse Virulence Factor Encoding Genes in Group A Streptococcus

Project description:We performed CovR ChIP-seq analysis in the emm1 strain MGAS2221 and its CovS kinase deficient derivative strain 2221-CovS-E281A. We identified that CovR bound in the promoter regions of nearly all virulence factor encoding genes in the CovR regulon. Additionally, direct CovR binding was observed for numerous genes encoding proteins involved in amino acid metabolism, but we found limited direct CovR binding to genes encoding other transcriptional regulators. The consensus sequence AATRANAAAARVABTAAA was present in the promoters of genes directly regulated by CovR, and mutations of highly conserved positions within this motif relieved CovR repression of the hasA and M2221_0187 promoters. Analysis of strain 2221-CovS-E281A revealed that binding of CovR at repressed, but not activated, promoters is highly dependent on CovR~P state. CovR repressed virulence factor encoding genes could be grouped dependent on how variation in CovR~P differentially impacted DNA binding and gene transcript levels.

2022-01-03 | GSE192795 | GEO

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2024-04-24 | MTBLS7972 | MetaboLights

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