Cascade-mediated binding and bending of negatively supercoiled DNA

Bacteria and archaea have evolved sophisticated adaptive immune systems, known as CRISPR–Cas (clustered regularly interspaced short palindromic repeats–CRISPR-associated proteins) systems, which target and inactivate invading viruses and plasmids. Immunity is acquired by integrating short fragments of foreign DNA into CRISPR loci, and following transcription and processing of these loci, the CRISPR RNAs (crRNAs) guide the Cas proteins to complementary invading nucleic acid, which results in target interference. In this Review, we summarize the recent structural and biochemical insights that have been gained for the three major types of CRISPR–Cas systems, which together provide a detailed molecular understanding of the unique and conserved mechanisms of RNA-guided adaptive immunity in bacteria and archaea.

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“…The targeting of invading MGEs by the different crRNP complexes seems to proceed in a stepwise manner 33,82,84,98,99 (FIG. 6).…”

Section: Target Surveillance and Interferencementioning

confidence: 99%

Unravelling the structural and mechanistic basis of CRISPR–Cas systems

et al. 2014

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“…While the Csa and Cst complexes have not been extensively studied, Cas5, Cas7, Cas8 (large subunit), and Cas11 (small subunit) superfamily proteins comprise other characterized Type I crRNPs (the latter sometimes fused to the large subunit as a domain) (Brouns et al 2008;Lintner et al 2011b;Wiedenheft et al 2011a,b;Nam et al 2012;Plagens et al 2012;van Duijn et al 2012;Brendel et al 2014). The signature Cas3 (HD nuclease-DExH helicase) proteins of Type I systems interact with the crRNPs and cleave invader DNA (Cady and O'Toole 2011;Westra et al 2012a;Mulepati and Bailey 2013;Sinkunas et al 2013;Hochstrasser et al 2014;Plagens et al 2014). The cas gene clusters in Pfu also encode predicted adaptation proteins (Cas1, Cas2, Cas4) and a crRNA biogenesis protein (Cas6) (Fig.…”

Section: Introductionmentioning

confidence: 99%

“…CRISPR RNAs (crRNAs) are produced by processing of CRISPR locus transcripts that arise from promoter elements in the leader region (Jansen et al 2002;Tang et al 2002). Each crRNA contains a guide region comprised of invader-derived sequences that allow crRNA-Cas protein effector complexes to recognize and destroy invader nucleic acids (Barrangou et al 2007;Marraffini and Sontheimer 2008;Hale et al 2009Hale et al , 2012Garneau et al 2010;Jore et al 2011;Lintner et al 2011b;Wiedenheft et al 2011a;Fischer et al 2012;Gasiunas et al 2012;Jinek et al 2012;Westra et al 2012a;Elmore et al 2013;Maier et al 2013;Mulepati and Bailey 2013;Peng et al 2013Peng et al , 2015Sinkunas et al 2013;Plagens et al 2014).…”

Section: Introductionmentioning

confidence: 99%

Three CRISPR-Cas immune effector complexes coexist in Pyrococcus furiosus

Majumdar

Zhao

Pfister

et al. 2015

RNA

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CRISPR-Cas immune systems function to defend prokaryotes against potentially harmful mobile genetic elements including viruses and plasmids. The multiple CRISPR-Cas systems (Types I, II, and III) each target destruction of foreign nucleic acids via structurally and functionally diverse effector complexes (crRNPs). CRISPR-Cas effector complexes are comprised of CRISPR RNAs (crRNAs) that contain sequences homologous to the invading nucleic acids and Cas proteins specific to each immune system type. We have previously characterized a crRNP in Pyrococcus furiosus (Pfu) that contains Cmr (Type III-B) Cas proteins associated with one of two size classes of crRNAs and cleaves complementary target RNAs. Here, we have isolated and characterized two additional native Pfu crRNPs containing either Csa (Type I-A) or Cst (Type I-G) Cas proteins and distinct profiles of associated crRNAs. For each complex, the Cas proteins were identified by mass spectrometry and immunoblotting and the crRNAs by RNA sequencing and Northern blot analysis. The crRNAs associated with both the Csa and Cst complexes originate from all seven Pfu CRISPR loci and contain identical 5 ′ ends (8-nt repeat-derived 5 ′ tag sequences) but heterogeneous 3 ′ ends (containing variable amounts of downstream repeat sequences). These crRNA forms are distinct from Cmr-associated crRNAs, indicating different 3 ′ end processing pathways following primary cleavage of common pre-crRNAs. Like other previously characterized Type I CRISPR-Cas effector complexes, we predict that the newly identified Pfu Csa and Cst crRNPs each function to target invading DNA, adding an additional layer of protection beyond that afforded by the previously characterized RNA targeting Cmr complex.

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“…According to the most recent classification, CRISPR systems are divided into two classes, six types (types I-VI), and 19 subtypes (7,8). Type I is the most common, consisting of seven subtypes (I-A to I-F, and I-U) that all rely on multisubunit crRNA-guided surveillance complexes for target detection, and a trans-acting effector protein called Cas3, which is required for target destruction (9)(10)(11)(12)(13). Cas3 proteins are typically composed of an amino-terminal histidine-aspartate (HD)-nuclease domain, fused to a carboxylterminal superfamily II (SF2) helicase (10,(13)(14)(15)(16)(17).…”

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confidence: 99%

Cas1 and the Csy complex are opposing regulators of Cas2/3 nuclease activity

Rollins

Chowdhury

Carter

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

102

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Significance Prokaryotes have adaptive immune systems that rely on CRISPRs (clustered regularly interspaced short palindromic repeats) and diverse CRISPR-associated ( cas ) genes. Cas1 and Cas2 are conserved components of CRISPR systems that are essential for integrating fragments of foreign DNA into CRISPR loci. In type I-F immune systems, the Cas2 adaptation protein is fused to the Cas3 interference protein. Here we show that the Cas2/3 fusion protein from Pseudomonas aeruginosa stably associates with the Cas1 adaptation protein, forming a 375-kDa propeller-shaped Cas1–2/3 complex. We show that Cas1, in addition to being an essential adaptation protein, also functions as a repressor of Cas2/3 nuclease activity and that foreign DNA binding by the CRISPR RNA-guided surveillance complex activates the Cas2/3 nuclease.

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Cascade-mediated binding and bending of negatively supercoiled DNA

Cited by 37 publications

References 31 publications

Unravelling the structural and mechanistic basis of CRISPR–Cas systems

Unravelling the structural and mechanistic basis of CRISPR–Cas systems

Three CRISPR-Cas immune effector complexes coexist in Pyrococcus furiosus

Cas1 and the Csy complex are opposing regulators of Cas2/3 nuclease activity

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