Cas5d Protein Processes Pre-crRNA and Assembles into a Cascade-like Interference Complex in Subtype I-C/Dvulg CRISPR-Cas System

Nam, Ki Hyun; Haitjema, Charles H.; Liu, Xueqi; Ding, Fran; Wang, Hongwei; DeLisa, Matthew P.; Ke, Ailong

doi:10.1016/j.str.2012.06.016

Cited by 200 publications

(277 citation statements)

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“…1B) and that csm2, csm3, and csm5 are required for crRNA maturation (19). Here, we show that Csm2, Csm3, and Csm5, along with Csm4 and the type III signature protein Cas10, are part of a ribonucleoprotein complex analogous to the Cascade (CRISPR-associated complex for antiviral defense) complex described for Escherichia coli and other organisms (2,5,16,20,21). This complex, here named Cas10⅐Csm, is enriched with mature crRNAs that range from 31 to 67 nucleotides, measured precisely in 6-nucleotide increments.…”

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confidence: 63%

“…In type I and III systems, Cas6 is considered the primary processing endonuclease (2,(13)(14)(15). One exception appears to be the type I-C system in Bacillus halodurans, where Cas5d was recently shown to catalyze the cleavage of repeat sequences (16). In contrast, primary processing in type II systems relies upon an antisense trans-encoded crRNA and the host RNase III to cleave within repeats (17).…”

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confidence: 99%

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A Ruler Protein in a Complex for Antiviral Defense Determines the Length of Small Interfering CRISPR RNAs

Hatoum-Aslan

Samai

Maniv

et al. 2013

Journal of Biological Chemistry

119

192

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Background: CRISPR immune systems protect prokaryotes from their viruses using small interfering RNAs (crRNAs), which require maturation events during their biogenesis. Results: In Staphylococcus epidermidis, crRNAs undergo maturation in a Cas10⅐Csm ribonucleoprotein complex; Csm3 modulates the extent of maturation. Conclusion: Csm3 acts as a ruler for crRNAs. Significance: Investigating CRISPR immunity is important to understand prokaryotic ecology and to develop biotechnological applications.

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confidence: 63%

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confidence: 99%

A Ruler Protein in a Complex for Antiviral Defense Determines the Length of Small Interfering CRISPR RNAs

Hatoum-Aslan

Samai

Maniv

et al. 2013

Journal of Biological Chemistry

119

192

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“…Common CRISPR-mediated immunity processes are usually defined into three stages: (i) acquisition of a short DNA segment (protospacer) from an invading virus or plasmid, and its insertion at the leader-proximal end of a CRISPR locus (7,8), (ii) generation of small mature CRISPR RNAs (crRNAs) from a longer transcript of a CRISPR locus (9)(10)(11), and (iii) interference of foreign nucleic acid invaders by a crRNA-containing ribonucleoprotein effector complex (12)(13)(14)(15)(16)(17)(18).…”

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confidence: 99%

Molecular insights into DNA interference by CRISPR-associated nuclease-helicase Cas3

Gong

Shin

Sun

et al. 2014

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

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Mobile genetic elements in bacteria are neutralized by a system based on clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins. Type I CRISPR-Cas systems use a "Cascade" ribonucleoprotein complex to guide RNA specifically to complementary sequence in invader double-stranded DNA (dsDNA), a process called "interference." After target recognition by Cascade, formation of an R-loop triggers recruitment of a Cas3 nuclease-helicase, completing the interference process by destroying the invader dsDNA. To elucidate the molecular mechanism of CRISPR interference, we analyzed crystal structures of Cas3 from the bacterium Thermobaculum terrenum, with and without a bound ATP analog. The structures reveal a histidine-aspartate (HD)-type nuclease domain fused to superfamily-2 (SF2) helicase domains and a distinct C-terminal domain. Binding of ATP analog at the interface of the SF2 helicase RecA-like domains rearranges a motif V with implications for the enzyme mechanism. The HDnucleolytic site contains two metal ions that are positioned at the end of a proposed nucleic acid-binding tunnel running through the SF2 helicase structure. This structural alignment suggests a mechanism for 3′ to 5′ nucleolytic processing of the displaced strand of invader DNA that is coordinated with ATP-dependent 3′ to 5′ translocation of Cas3 along DNA. In agreement with biochemical studies, the presented Cas3 structures reveal important mechanistic details on the neutralization of genetic invaders by type I CRISPR-Cas systems.M any bacteria and archaea can eliminate invading phages and plasmids by activating a defense system that is based on processing of clustered regularly interspaced short palindromic repeats (CRISPRs) by CRISPR-associated (Cas) proteins and various additional proteins. These CRISPR-Cas systems are diverse in structure and function, but common principles of action allow their classification into three major types (I, II, and III) with some further divisions into subtypes (e.g., type IA-F) (1-6).Common CRISPR-mediated immunity processes are usually defined into three stages: (i) acquisition of a short DNA segment (protospacer) from an invading virus or plasmid, and its insertion at the leader-proximal end of a CRISPR locus (7, 8), (ii) generation of small mature CRISPR RNAs (crRNAs) from a longer transcript of a CRISPR locus (9-11), and (iii) interference of foreign nucleic acid invaders by a crRNA-containing ribonucleoprotein effector complex (12-18).Different interference effector complexes characterize the three major CRISPR types. Cascade (Cas complex for antiviral defense) complexes are synonymous with interference in type I CRISPRs, comprising multiple proteins in a helical structure around crRNA that targets invader DNA (13,19, 20). Type II CRISPR systems are characterized by Cas9, a multidomain protein with a bilobed architecture harboring two nuclease sites that together catalyze cleavage of invading DNA (12, 21-24). The Csm system in CRISPR type IIIA and a related Cmr ...

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“…Cleavage of the pre-crRNA generates a product comprising a full spacer sequence flanked by repeat-derived handles on both ends. In most cases, the 5' handles are 8 nt (nucleotides) long (11nt in type I-C systems (Nam et al, 2012b) and 13 nt in I-D systems (Scholz et al, 2013), whereas the 3' handles have variable lengths depending on the specific CRISPR-Cas type. In some types, the 3' handles form hairpin structures (Carte et al, 2010; Carte et al, 2008;Niewoehner et al, 2014;Wang et al, 2011).…”

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confidence: 99%

“…In some types, the 3' handles form hairpin structures (Carte et al, 2010; Carte et al, 2008;Niewoehner et al, 2014;Wang et al, 2011). In type I-C systems, initial processing is performed by Cas5 rather than Cas6 (Garside et al, 2012;Nam et al, 2012b), whereas in other CRISPR-Cas types, Cas5 is catalytically inactive and is thought to interact with the 5' handle of the crRNA . In most type I and type III systems, after initial processing, the crRNA is transferred to CRISPR ribonucleoprotein (crRNP) complexes for secondary processing (Hatoum-Aslan et al, 2011).…”

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confidence: 99%

CRISPR-Cas Systems in Prokaryotes

Burmistrz

Pyrć

2015

Pol J Microbiol

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A b s t r a c tProkaryotic organisms possess numerous strategies that enable survival in hostile conditions. Among others, these conditions include the invasion of foreign nucleic acids such as bacteriophages and plasmids. The clustered regularly interspaced palindromic repeats-CRISPRassociated proteins (CRISPR-Cas) system provides the majority of bacteria and archaea with adaptive and hereditary immunity against this threat. This mechanism of immunity is based on short fragments of foreign DNA incorporated within the hosts genome. After transcription, these fragments guide protein complexes that target foreign nucleic acids and promote their degradation. The aim of this review is to summarize the current status of CRISPR-Cas research, including the mechanisms of action, the classification of different types and subtypes of these systems, and the development of new CRISPR-Cas-based molecular biology tools. K e y w o r d s: CRISPR-Cas, prokaryotesBurmistrz M. and Pyrć K. 3 194 spacer sequences can be of either foreign or self-origin (Stern et al., 2010;Vercoe et al., 2013). The proteomic component is responsible for the incorporation of new template sequences, processing them into a form that enables base pairing with target nucleic acids, as well as for scanning and cleavage of target DNA or RNA. Of note, not all CRISPR-Cas systems discovered to date are active (Haft et al., 2005;van der Ploeg, 2009). Structure of the CRISPR-Cas systemThe genomic component of the CRISPR-Cas system is formed by a series of variable spacers, which in some cases share sequence similarity with viruses, plasmids, or bacteria. These regions are interspaced with repeat sequences that are identical or almost identical within a single CRISPR cassette. The length of the repeat sequences varies between 25 and 40 nt, whereas the length of the spacer sequences varies between 21 and 72 nt. As mentioned above, some spacers show high homology with foreign nucleic acids, but the origin of a significant percentage of spacers remains unknown. The sequence homology between the spacer and the target is the major determinant of nucleic acid degradation of the target. Some bacterial species contain more than one CRISPR locus within their genome (Louwen et al., 2014). Depending on the specific bacterial species or strain, a CRISPR locus may contain from a few to several hundred repeat spacer units; however, most commonly, a single CRISPR locus contains approximately 50 units. The CRISPR repeat sequences play an important role during both the acquisition of new spacers and the transcription and maturation of CRISPR RNA (crRNA). Based on the sequence similarity, the CRISPR repeats are assigned into groups (Kunin et al., 2007). These groups were taken into account in the current classification of CRISPR-Cas systems (Makarova et al., 2011). Although most CRISPR arrays are located on chromosomal DNA, there are examples of CRISPRs located on plasmids (Godde and Bickerton, 2006). In general, CRISPR loci are flanked by A/T rich leader sequences containing...

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Cas5d Protein Processes Pre-crRNA and Assembles into a Cascade-like Interference Complex in Subtype I-C/Dvulg CRISPR-Cas System

Cited by 200 publications

References 41 publications

A Ruler Protein in a Complex for Antiviral Defense Determines the Length of Small Interfering CRISPR RNAs

A Ruler Protein in a Complex for Antiviral Defense Determines the Length of Small Interfering CRISPR RNAs

Molecular insights into DNA interference by CRISPR-associated nuclease-helicase Cas3

CRISPR-Cas Systems in Prokaryotes

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