TY - JOUR
T1 - Crystal structure of the crispr-cas RNA silencing cmr complex bound to a target analog
AU - Osawa, Takuo
AU - Inanaga, Hideko
AU - Sato, Chikara
AU - Numata, Tomoyuki
N1 - Funding Information:
We thank Dr. Kozo Tomita (AIST) for valuable and critical comments and suggestions for this manuscript. We thank Professor Yoshizumi Ishino (Kyushu University) for providing the P. furiosus genomic DNA for the experiment. We also thank the beamline staff at BL-17A of the Photon Factory (Ibaraki, Japan) for technical assistance during data collection. This work was supported by a Grant-in-Aid for Scientific Research on Innovative Areas from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) to T.N. and C.S. and by a Grant-in-Aid for Young Scientists from the Japan Society for the Promotion of Science (JSPS) and grants from the Kato Memorial Bioscience Foundation, the Kurata Memorial Hitachi Science and Technology Foundation, and the Institute for Fermentation, Osaka (IFO) to T.N.
Publisher Copyright:
© 2015 Elsevier Inc.
PY - 2015/5/7
Y1 - 2015/5/7
N2 - In prokaryotes, Clustered regularly interspaced shortpalindromic repeat (CRISPR)-derived RNAs (crRNAs), together with CRISPR-associated (Cas) proteins, capture and degrade invading genetic materials. In the type III-B CRISPR-Cas system, six Cas proteins (Cmr1-Cmr6) and a crRNA form an RNA silencing Cmr complex. Here we report the 2.1Å crystal structure of the Cmr1-deficient, functional Cmr complex bound to single-stranded DNA, a substrate analog complementary to the crRNA guide. Cmr3 recognizes the crRNA 5' tag and defines the start position of the guide-target duplex, using its idiosyncratic loops. The β-hairpins of three Cmr4 subunits intercalate within the duplex, causing nucleotide displacements with 6 nt intervals, and thus periodically placing the scissile bonds near the crucial aspartate of Cmr4. The structure reveals the mechanism for specifying the periodic target cleavage sites from the crRNA 5' tag and provides insights into the assembly of the type III interference machineries and the evolution of the Cmr and Cascade complexes.
AB - In prokaryotes, Clustered regularly interspaced shortpalindromic repeat (CRISPR)-derived RNAs (crRNAs), together with CRISPR-associated (Cas) proteins, capture and degrade invading genetic materials. In the type III-B CRISPR-Cas system, six Cas proteins (Cmr1-Cmr6) and a crRNA form an RNA silencing Cmr complex. Here we report the 2.1Å crystal structure of the Cmr1-deficient, functional Cmr complex bound to single-stranded DNA, a substrate analog complementary to the crRNA guide. Cmr3 recognizes the crRNA 5' tag and defines the start position of the guide-target duplex, using its idiosyncratic loops. The β-hairpins of three Cmr4 subunits intercalate within the duplex, causing nucleotide displacements with 6 nt intervals, and thus periodically placing the scissile bonds near the crucial aspartate of Cmr4. The structure reveals the mechanism for specifying the periodic target cleavage sites from the crRNA 5' tag and provides insights into the assembly of the type III interference machineries and the evolution of the Cmr and Cascade complexes.
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U2 - 10.1016/j.molcel.2015.03.018
DO - 10.1016/j.molcel.2015.03.018
M3 - Article
C2 - 25921071
AN - SCOPUS:84928925211
SN - 1097-2765
VL - 58
SP - 418
EP - 430
JO - Molecular Cell
JF - Molecular Cell
IS - 3
ER -