Sequence- and Structure-specific RNA Processing by a CRISPR Endonuclease

Overview

Journal Science

Specialty Science

Date 2010 Sep 11

PMID 20829488

Citations 366

Authors

Rachel E Haurwitz

Martin Jinek

Blake Wiedenheft

Kaihong Zhou

Jennifer A Doudna

Affiliations

Soon will be listed here.

Abstract

Many bacteria and archaea contain clustered regularly interspaced short palindromic repeats (CRISPRs) that confer resistance to invasive genetic elements. Central to this immune system is the production of CRISPR-derived RNAs (crRNAs) after transcription of the CRISPR locus. Here, we identify the endoribonuclease (Csy4) responsible for CRISPR transcript (pre-crRNA) processing in Pseudomonas aeruginosa. A 1.8 angstrom crystal structure of Csy4 bound to its cognate RNA reveals that Csy4 makes sequence-specific interactions in the major groove of the crRNA repeat stem-loop. Together with electrostatic contacts to the phosphate backbone, these enable Csy4 to bind selectively and cleave pre-crRNAs using phylogenetically conserved serine and histidine residues in the active site. The RNA recognition mechanism identified here explains sequence- and structure-specific processing by a large family of CRISPR-specific endoribonucleases.

Citing Articles

A trigger-inducible split-Csy4 architecture for programmable RNA modulation.

Zhang L, Qiu X, Zhou Y, Luo Z, Zhu L, Shao J Nucleic Acids Res. 2025; 53(2).

PMID: 39817512 PMC: 11734699. DOI: 10.1093/nar/gkae1319.

Delivery of Prime editing in human stem cells using pseudoviral NanoScribes particles.

Halegua T, Risson V, Carras J, Rouyer M, Coudert L, Jacquier A Nat Commun. 2025; 16(1):397.

PMID: 39755699 PMC: 11700204. DOI: 10.1038/s41467-024-55604-0.

Research Progress on the Mechanism and Application of the Type I CRISPR-Cas System.

Yang P, Zhang S, Hu D, Li X, Guo Y, Guo H Int J Mol Sci. 2024; 25(23).

PMID: 39684256 PMC: 11640852. DOI: 10.3390/ijms252312544.

Nonlinear mechanical response of finite-length soft composites with random dislocations.

Jalilvand S, Mirzaei M, Mousavi H Sci Rep. 2024; 14(1):27121.

PMID: 39511290 PMC: 11543800. DOI: 10.1038/s41598-024-78623-9.

Insight into the natural regulatory mechanisms and clinical applications of the CRISPR-Cas system.

Cheng H, Deng H, Ma D, Gao M, Zhou Z, Li H Heliyon. 2024; 10(20):e39538.

PMID: 39502233 PMC: 11535992. DOI: 10.1016/j.heliyon.2024.e39538.

References

Jansen R, van Embden J, Gaastra W, Schouls L . Identification of genes that are associated with DNA repeats in prokaryotes. Mol Microbiol. 2002; 43(6):1565-75. DOI: 10.1046/j.1365-2958.2002.02839.x. View

Tang T, Bachellerie J, Rozhdestvensky T, Bortolin M, Huber H, Drungowski M . Identification of 86 candidates for small non-messenger RNAs from the archaeon Archaeoglobus fulgidus. Proc Natl Acad Sci U S A. 2002; 99(11):7536-41. PMC: 124276. DOI: 10.1073/pnas.112047299. View

Grissa I, Vergnaud G, Pourcel C . The CRISPRdb database and tools to display CRISPRs and to generate dictionaries of spacers and repeats. BMC Bioinformatics. 2007; 8:172. PMC: 1892036. DOI: 10.1186/1471-2105-8-172. View

Cai Z, Gorin A, Frederick R, Ye X, Hu W, Majumdar A . Solution structure of P22 transcriptional antitermination N peptide-boxB RNA complex. Nat Struct Biol. 1998; 5(3):203-12. DOI: 10.1038/nsb0398-203. View

Marraffini L, Sontheimer E . CRISPR interference: RNA-directed adaptive immunity in bacteria and archaea. Nat Rev Genet. 2010; 11(3):181-90. PMC: 2928866. DOI: 10.1038/nrg2749. View

Marraffini L, Sontheimer E . CRISPR interference limits horizontal gene transfer in staphylococci by targeting DNA. Science. 2008; 322(5909):1843-5. PMC: 2695655. DOI: 10.1126/science.1165771. View

Horvath P, Barrangou R . CRISPR/Cas, the immune system of bacteria and archaea. Science. 2010; 327(5962):167-70. DOI: 10.1126/science.1179555. View

Ye X, Gorin A, Ellington A, Patel D . Deep penetration of an alpha-helix into a widened RNA major groove in the HIV-1 rev peptide-RNA aptamer complex. Nat Struct Biol. 1996; 3(12):1026-33. DOI: 10.1038/nsb1296-1026. View

Anand K, Schulte A, Vogel-Bachmayr K, Scheffzek K, Geyer M . Structural insights into the cyclin T1-Tat-TAR RNA transcription activation complex from EIAV. Nat Struct Mol Biol. 2008; 15(12):1287-92. DOI: 10.1038/nsmb.1513. View

10.

Haft D, Selengut J, Mongodin E, Nelson K . A guild of 45 CRISPR-associated (Cas) protein families and multiple CRISPR/Cas subtypes exist in prokaryotic genomes. PLoS Comput Biol. 2005; 1(6):e60. PMC: 1282333. DOI: 10.1371/journal.pcbi.0010060. View

11.

Legault P, Li J, Mogridge J, Kay L, GREENBLATT J . NMR structure of the bacteriophage lambda N peptide/boxB RNA complex: recognition of a GNRA fold by an arginine-rich motif. Cell. 1998; 93(2):289-99. DOI: 10.1016/s0092-8674(00)81579-2. View

12.

Holm L, Sander C . Protein structure comparison by alignment of distance matrices. J Mol Biol. 1993; 233(1):123-38. DOI: 10.1006/jmbi.1993.1489. View

13.

Lillestol R, Shah S, Brugger K, Redder P, Phan H, Christiansen J . CRISPR families of the crenarchaeal genus Sulfolobus: bidirectional transcription and dynamic properties. Mol Microbiol. 2009; 72(1):259-72. DOI: 10.1111/j.1365-2958.2009.06641.x. View

14.

Ebihara A, Yao M, Masui R, Tanaka I, Yokoyama S, Kuramitsu S . Crystal structure of hypothetical protein TTHB192 from Thermus thermophilus HB8 reveals a new protein family with an RNA recognition motif-like domain. Protein Sci. 2006; 15(6):1494-9. PMC: 2242536. DOI: 10.1110/ps.062131106. View

15.

Raines R . Ribonuclease A. Chem Rev. 2002; 98(3):1045-1066. DOI: 10.1021/cr960427h. View

16.

Huppler A, Nikstad L, Allmann A, Brow D, Butcher S . Metal binding and base ionization in the U6 RNA intramolecular stem-loop structure. Nat Struct Biol. 2002; 9(6):431-5. DOI: 10.1038/nsb800. View

17.

Sorek R, Kunin V, Hugenholtz P . CRISPR--a widespread system that provides acquired resistance against phages in bacteria and archaea. Nat Rev Microbiol. 2007; 6(3):181-6. DOI: 10.1038/nrmicro1793. View

18.

Barrangou R, Fremaux C, Deveau H, Richards M, Boyaval P, Moineau S . CRISPR provides acquired resistance against viruses in prokaryotes. Science. 2007; 315(5819):1709-12. DOI: 10.1126/science.1138140. View

19.

Makarova K, Grishin N, Shabalina S, Wolf Y, Koonin E . A putative RNA-interference-based immune system in prokaryotes: computational analysis of the predicted enzymatic machinery, functional analogies with eukaryotic RNAi, and hypothetical mechanisms of action. Biol Direct. 2006; 1:7. PMC: 1462988. DOI: 10.1186/1745-6150-1-7. View

20.

Kunin V, Sorek R, Hugenholtz P . Evolutionary conservation of sequence and secondary structures in CRISPR repeats. Genome Biol. 2007; 8(4):R61. PMC: 1896005. DOI: 10.1186/gb-2007-8-4-r61. View