Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin

Bolotin, Alexander; Quinquis, Benoît; Sorokin, Alexeï; Ehrlich, S. Dusko

doi:10.1099/mic.0.28048-0

Cited by 1,487 publications

(922 citation statements)

References 33 publications

(28 reference statements)

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“…CRISPR arrays consist of tandem direct repeats (DRs) of 23 to 55 bp in length separated by similarly sized variable spacer sequences that are generally derived from bacteriophages or plasmids (7)(8)(9)(10).…”

Section: Organization Of Crispr Locimentioning

confidence: 99%

CRISPRs: Molecular Signatures Used for Pathogen Subtyping

Shariat

Dudley

2014

Appl Environ Microbiol

132

115

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Rapid and accurate strain identification is paramount in the battle against microbial outbreaks, and several subtyping approaches have been developed. One such method uses clustered regular interspaced short palindromic repeats (CRISPRs), DNA repeat elements that are present in approximately half of all bacteria. Though their signature function is as an adaptive immune system against invading DNA such as bacteriophages and plasmids, CRISPRs also provide an excellent framework for pathogen tracking and evolutionary studies. Analysis of the spacer DNA sequences that reside between the repeats has been tremendously useful for bacterial subtyping during molecular epidemiological investigations. Subtyping, or strain identification, using CRISPRs has been employed in diverse Gram-positive and Gram-negative bacteria, including Mycobacterium tuberculosis, Salmonella enterica, and the plant pathogen Erwinia amylovora. This review discusses the several ways in which CRISPR sequences are exploited for subtyping. This includes the well-established spoligotyping methodologies that have been used for 2 decades to type Mycobacterium species, as well as in-depth consideration of newer, higher-throughput CRISPR-based protocols.

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Section: Organization Of Crispr Locimentioning

confidence: 99%

CRISPRs: Molecular Signatures Used for Pathogen Subtyping

Shariat

Dudley

2014

Appl Environ Microbiol

132

115

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“…For a small fraction of the spacers, protospacers have been reported, often in viral and plasmid genomes, but the substantial majority of the spacers remain without a match, representing vast CRISPR "dark matter" (43)(44)(45)(46)(47)(48)(49).…”

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confidence: 99%

The CRISPR Spacer Space Is Dominated by Sequences from Species-Specific Mobilomes

Shmakov

Sitnik

Makarova

et al. 2017

mBio

202

195

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Clustered regularly interspaced short palindromic repeats and CRISPRassociated protein (CRISPR-Cas) systems store the memory of past encounters with foreign DNA in unique spacers that are inserted between direct repeats in CRISPR arrays. For only a small fraction of the spacers, homologous sequences, called protospacers, are detectable in viral, plasmid, and microbial genomes. The rest of the spacers remain the CRISPR "dark matter." We performed a comprehensive analysis of the spacers from all CRISPR-cas loci identified in bacterial and archaeal genomes, and we found that, depending on the CRISPR-Cas subtype and the prokaryotic phylum, protospacers were detectable for 1% to about 19% of the spacers (~7% global average). Among the detected protospacers, the majority, typically 80 to 90%, originated from viral genomes, including proviruses, and among the rest, the most common source was genes that are integrated into microbial chromosomes but are involved in plasmid conjugation or replication. Thus, almost all spacers with identifiable protospacers target mobile genetic elements (MGE). The GC content, as well as dinucleotide and tetranucleotide compositions, of microbial genomes, their spacer complements, and the cognate viral genomes showed a nearly perfect correlation and were almost identical. Given the near absence of self-targeting spacers, these findings are most compatible with the possibility that the spacers, including the dark matter, are derived almost completely from the species-specific microbial mobilomes. IMPORTANCEThe principal function of CRISPR-Cas systems is thought to be protection of bacteria and archaea against viruses and other parasitic genetic elements. The CRISPR defense function is mediated by sequences from parasitic elements, known as spacers, that are inserted into CRISPR arrays and then transcribed and employed as guides to identify and inactivate the cognate parasitic genomes. However, only a small fraction of the CRISPR spacers match any sequences in the current databases, and of these, only a minority correspond to known parasitic elements. We show that nearly all spacers with matches originate from viral or plasmid genomes that are either free or have been integrated into the host genome. We further demonstrate that spacers with no matches have the same properties as those of identifiable origins, strongly suggesting that all spacers originate from mobile elements.KEYWORDS CRISPR-Cas, bacteriophages, mobilome, oligonucleotide composition, spacer acquisition C RISPR-Cas (clustered regularly interspaced palindromic repeats and CRISPRassociated proteins) systems are adaptive (acquired) immune systems of archaea and bacteria that store memory of past encounters with foreign DNA in unique spacer Citation Shmakov SA, Sitnik V, Makarova KS, Wolf YI, Severinov KV, Koonin EV. 2017. The CRISPR spacer space is dominated by sequences from species-specific mobilomes.

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“…CRISPRs consist of short repeats interspersed with nonrepetitive nucleotides of 26 to 72 bp called spacers derived from exogenous genetic elements (9)(10)(11). CRISPRs are often associated with a set of cas genes that encode proteins that mediate the defense process.…”

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confidence: 99%

Role of the Streptococcus mutans CRISPR-Cas Systems in Immunity and Cell Physiology

et al. 2015

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b CRISPR-Cas systems provide adaptive microbial immunity against invading viruses and plasmids. The cariogenic bacterium Streptococcus mutans UA159 has two CRISPR-Cas systems: CRISPR1 (type II-A) and CRISPR2 (type I-C) with several spacers from both CRISPR cassettes matching sequences of phage M102 or genomic sequences of other S. mutans. The deletion of the cas genes of CRISPR1 (⌬C1S), CRISPR2 (⌬C2E), or both CRISPR1؉2 (⌬C1SC2E) or the removal of spacers 2 and 3 (⌬CR1SP13E) in S. mutans UA159 did not affect phage sensitivity when challenged with virulent phage M102. Using plasmid transformation experiments, we demonstrated that the CRISPR1-Cas system inhibits transformation of S. mutans by the plasmids matching the spacers 2 and 3. Functional analysis of the cas deletion mutants revealed that in addition to a role in plasmid targeting, both CRISPR systems also contribute to the regulation of bacterial physiology in S. mutans. Compared to wild-type cells, the ⌬C1S strain displayed diminished growth under cell membrane and oxidative stress, enhanced growth under low pH, and had reduced survival under heat shock and DNA-damaging conditions, whereas the ⌬C2E strain exhibited increased sensitivity to heat shock. Transcriptional analysis revealed that the two-component signal transduction system VicR/K differentially modulates expression of cas genes within CRISPR-Cas systems, suggesting that VicR/K might coordinate the expression of two CRISPR-Cas systems. Collectively, we provide in vivo evidence that the type II-A CRISPR-Cas system of S. mutans may be targeted to manipulate its stress response and to influence the host to control the uptake and dissemination of antibiotic resistance genes.

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Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin

Cited by 1,487 publications

References 33 publications

CRISPRs: Molecular Signatures Used for Pathogen Subtyping

CRISPRs: Molecular Signatures Used for Pathogen Subtyping

The CRISPR Spacer Space Is Dominated by Sequences from Species-Specific Mobilomes

Role of the Streptococcus mutans CRISPR-Cas Systems in Immunity and Cell Physiology

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