title: A type III-B CRISPR-Cas effector complex mediating massive target DNA destruction.
abstract: The CRISPR (clustered regularly interspaced short palindromic repeats) system protects archaea and bacteria by eliminating nucleic acid invaders in a crRNA-guided manner. The Sulfolobus islandicus type III-B Cmr-α system targets invading nucleic acid at both RNA and DNA levels and DNA targeting relies on the directional transcription of the protospacer in vivo. To gain further insight into the involved mechanism, we purified a native effector complex of III-B Cmr-α from S. islandicus and characterized it in vitro. Cmr-α cleaved RNAs complementary to crRNA present in the complex and its ssDNA destruction activity was activated by target RNA. The ssDNA cleavage required mismatches between the 5΄-tag of crRNA and the 3΄-flanking region of target RNA. An invader plasmid assay showed that mutation either in the histidine-aspartate acid (HD) domain (a quadruple mutation) or in the GGDD motif of the Cmr-2α protein resulted in attenuation of the DNA interference in vivo. However, double mutation of the HD motif only abolished the DNase activity in vitro. Furthermore, the activated Cmr-α binary complex functioned as a highly active DNase to destroy a large excess DNA substrate, which could provide a powerful means to rapidly degrade replicating viral DNA.
authors.affiliation: Archaea Centre, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, Copenhagen Biocenter, DK-2200 Copenhagen N, Denmark.
authors.forename: Wenyuan
authors.initials: W
authors.lastname: Han
authors.affiliation: State Key Laboratory of Agricultural Microbiology and College of Life Science and Technology, Huazhong Agricultural University, 430070 Wuhan, China.
authors.forename: Yingjun
authors.initials: Y
authors.lastname: Li
authors.affiliation: Archaea Centre, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, Copenhagen Biocenter, DK-2200 Copenhagen N, Denmark.
authors.forename: Ling
authors.initials: L
authors.lastname: Deng
authors.affiliation: State Key Laboratory of Agricultural Microbiology and College of Life Science and Technology, Huazhong Agricultural University, 430070 Wuhan, China.
authors.forename: Mingxia
authors.initials: M
authors.lastname: Feng
authors.affiliation: Archaea Centre, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, Copenhagen Biocenter, DK-2200 Copenhagen N, Denmark.
authors.forename: Wenfang
authors.initials: W
authors.lastname: Peng
authors.affiliation: Archaea Centre, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, Copenhagen Biocenter, DK-2200 Copenhagen N, Denmark.
authors.forename: Søren
authors.initials: S
authors.lastname: Hallstrøm
authors.affiliation: Biomedical Sciences Research Complex, University of St Andrews, Fife KY16 9ST, UK.
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Agricultural Microbiology and College of Life Science and Technology, Huazhong Agricultural University, 430070 Wuhan, China.
authors.forename: Nan
authors.initials: N
authors.lastname: Peng
authors.affiliation: State Key Laboratory of Agricultural Microbiology and College of Life Science and Technology, Huazhong Agricultural University, 430070 Wuhan, China.
authors.forename: Yun Xiang
authors.initials: YX
authors.lastname: Liang
authors.affiliation: Biomedical Sciences Research Complex, University of St Andrews, Fife KY16 9ST, UK.
authors.forename: Malcolm F
authors.initials: MF
authors.lastname: White
authors.affiliation: Archaea Centre, Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, Copenhagen Biocenter, DK-2200 Copenhagen N, Denmark.|State Key Laboratory of Agricultural Microbiology and College of Life Science and Technology, Huazhong Agricultural University, 430070 Wuhan, China.
authors.forename: Qunxin
authors.initials: Q
authors.lastname: She
mesh_terms: D064130:CRISPR-Associated Proteins / Q000378:metabolism*; D064113:CRISPR-Cas Systems; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D053837:DNA Cleavage; D019641:DNA, Archaeal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D046912:Multiprotein Complexes / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D019642:RNA, Archaeal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism; D016944:Sulfolobus / Q000235:genetics / Q000378:metabolism
chemical_list: D064130:CRISPR-Associated Proteins; D019641:DNA, Archaeal; D046912:Multiprotein Complexes; D019642:RNA, Archaeal; D012261:Ribonucleoproteins