title: RNA-guided complex from a bacterial immune system enhances target recognition through seed sequence interactions.
abstract: Prokaryotes have evolved multiple versions of an RNA-guided adaptive immune system that targets foreign nucleic acids. In each case, transcripts derived from clustered regularly interspaced short palindromic repeats (CRISPRs) are thought to selectively target invading phage and plasmids in a sequence-specific process involving a variable cassette of CRISPR-associated (cas) genes. The CRISPR locus in Pseudomonas aeruginosa (PA14) includes four cas genes that are unique to and conserved in microorganisms harboring the Csy-type (CRISPR system yersinia) immune system. Here we show that the Csy proteins (Csy1-4) assemble into a 350 kDa ribonucleoprotein complex that facilitates target recognition by enhancing sequence-specific hybridization between the CRISPR RNA and complementary target sequences. Target recognition is enthalpically driven and localized to a "seed sequence" at the 5' end of the CRISPR RNA spacer. Structural analysis of the complex by small-angle X-ray scattering and single particle electron microscopy reveals a crescent-shaped particle that bears striking resemblance to the architecture of a large CRISPR-associated complex from Escherichia coli, termed Cascade. Although similarity between these two complexes is not evident at the sequence level, their unequal subunit stoichiometry and quaternary architecture reveal conserved structural features that may be common among diverse CRISPR-mediated defense systems.
authors.affiliation: Howard Hughes Medical Institute, University of California, Berkeley, CA 94720, USA.
authors.forename: Blake
authors.initials: B
authors.lastname: Wiedenheft
authors.forename: Esther
authors.initials: E
authors.lastname: van Duijn
authors.forename: Jelle B
authors.initials: JB
authors.lastname: Bultema
authors.forename: Jelle
authors.initials: J
authors.lastname: Bultema
authors.forename: Sakharam P
authors.initials: SP
authors.lastname: Waghmare
authors.forename: Sakharam
authors.initials: S
authors.lastname: Waghmare
authors.forename: Kaihong
authors.initials: K
authors.lastname: Zhou
authors.forename: Arjan
authors.initials: A
authors.lastname: Barendregt
authors.forename: Wiebke
authors.initials: W
authors.lastname: Westphal
authors.forename: Albert J R
authors.initials: AJ
authors.lastname: Heck
authors.forename: Albert
authors.initials: A
authors.lastname: Heck
authors.forename: Egbert J
authors.initials: EJ
authors.lastname: Boekema
authors.forename: Egbert
authors.initials: E
authors.lastname: Boekema
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Dickman
authors.forename: Mark
authors.initials: M
authors.lastname: Dickman
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Doudna
mesh_terms: D056704:Adaptive Immunity / Q000235:genetics / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D004722:Endoribonucleases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology; D046911:Macromolecular Substances / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000276:immunology; D012313:RNA / Q000235:genetics* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D046911:Macromolecular Substances; D012313:RNA; D004722:Endoribonucleases