Two Distinct DNA Binding Modes Guide Dual Roles of a CRISPR-Cas Protein Complex

被引：75

作者：

Blosser, Timothy R. ^{[1
,2
]}

Loeff, Luuk ^{[1
,2
]}

Westra, Edze R. ^{[3
]}

Vlot, Marnix ^{[3
]}

Kunne, Tim ^{[3
]}

Sobota, Malgorzata ^{[3
]}

Dekker, Cees ^{[1
,2
]}

Brouns, Stan J. J. ^{[3
]}

Joo, Chirlmin ^{[1
,2
]}

机构：

[1] Delft Univ Technol, Kavli Inst NanoSci, NL-2628 CJ Delft, Netherlands

[2] Delft Univ Technol, Dept BioNanoSci, NL-2628 CJ Delft, Netherlands

[3] Wageningen Univ, Dept Agrotechnol & Food Sci, Lab Microbiol, NL-6703 HB Wageningen, Netherlands

来源：

MOLECULAR CELL | 2015年 / 58卷 / 01期

基金：

欧洲研究理事会;

关键词：

IN-VITRO RECONSTITUTION; ADAPTIVE IMMUNE-SYSTEMS; MEMORY B-CELLS; SURVEILLANCE COMPLEX; CRYSTAL-STRUCTURE; RNA; INTERFERENCE; DEGRADATION; CASCADE; TRANSCRIPTION;

D O I：

10.1016/j.molcel.2015.01.028

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Small RNA-guided protein complexes play an essential role in CRISPR-mediated immunity in prokaryotes. While these complexes initiate interference by flagging cognate invader DNA for destruction, recent evidence has implicated their involvement in new CRISPR memory formation, called priming, against mutated invader sequences. The mechanism by which the target recognition complex mediates these disparate responses-interference and priming-remains poorly understood. Using single-molecule FRET, we visualize how bona fide and mutated targets are differentially probed by E. coli Cascade. We observe that the recognition of bona fide targets is an ordered process that is tightly controlled for high fidelity. Mutated targets are recognized with low fidelity, which is featured by short-lived and PAM- and seed-independent binding by any segment of the crRNA. These dual roles of Cascade in immunity with distinct fidelities underpin CRISPR-Cas robustness, allowing for efficient degradation of bona fide targets and priming of mutated DNA targets.

引用

页码：60 / 70

页数：11

共 56 条

[21] Crystal structure of a CRISPR RNA-guided surveillance complex bound to a ssDNA target [J].