Type III-A CRISPR-Cas Csm complexes: Assembly, periodic RNA cleavage, DNase activity regulation, and autoimmunity Journal Article

Authors: Jia, N.; Mo, C. Y.; Wang, C.; Eng, E. T.; Marraffini, L. A.; Patel, D. J.
Article Title: Type III-A CRISPR-Cas Csm complexes: Assembly, periodic RNA cleavage, DNase activity regulation, and autoimmunity
Abstract: Type ΙΙΙ CRISPR-Cas systems provide robust immunity against foreign RNA and DNA by sequence-specific RNase and target RNA-activated sequence-nonspecific DNase and RNase activities. We report on cryo-EM structures of Thermococcus onnurineus CsmcrRNA binary, CsmcrRNA-target RNA and CsmcrRNA-target RNAanti-tag ternary complexes in the 3.1 Å range. The topological features of the crRNA 5′-repeat tag explains the 5′-ruler mechanism for defining target cleavage sites, with accessibility of positions -2 to -5 within the 5′-repeat serving as sensors for avoidance of autoimmunity. The Csm3 thumb elements introduce periodic kinks in the crRNA-target RNA duplex, facilitating cleavage of the target RNA with 6-nt periodicity. Key Glu residues within a Csm1 loop segment of CsmcrRNA adopt a proposed autoinhibitory conformation suggestive of DNase activity regulation. These structural findings, complemented by mutational studies of key intermolecular contacts, provide insights into CsmcrRNA complex assembly, mechanisms underlying RNA targeting and site-specific periodic cleavage, regulation of DNase cleavage activity, and autoimmunity suppression. © 2018 Cryo-EM structures of type III-A CsmcrRNA in the absence or presence of target RNA define the role of the 5′-repeat tag and kinks in positioning of 6-nt periodic RNA cleavage sites, the contribution of a Glu-rich loop to DNase activity regulation, and pairing capacity at −2 to −5 within 5′-repeat dictating autoimmunity. © 2018
Keywords: cryo-em; 5′ repeat tag pairing controls autoimmunity; dnase activity regulation by glu residues; rna cleavage with 6-nt periodicity; t. onnurineus csm; ternary complex assembly with crrna and target rna; type iii-a crispr-cas complexes
Journal Title: Molecular Cell
Volume: 73
Issue: 2
ISSN: 1097-2765
Publisher: Cell Press  
Date Published: 2019-01-17
Start Page: 264
End Page: 277.e5
Language: English
DOI: 10.1016/j.molcel.2018.11.007
PUBMED: 30503773
PROVIDER: scopus
PMCID: PMC6355164
Notes: Export Date: 1 February 2019 -- Source: Scopus
Citation Impact
MSK Authors
  1. Dinshaw J Patel
    426 Patel
  2. Chongyuan Wang
    2 Wang
  3. Ning Jia
    6 Jia