Role of nucleotide identity in effective CRISPR target escape mutations.

Abstract:

:Prokaryotes use primed CRISPR adaptation to update their memory bank of spacers against invading genetic elements that have escaped CRISPR interference through mutations in their protospacer target site. We previously observed a trend that nucleotide-dependent mismatches between crRNA and the protospacer strongly influence the efficiency of primed CRISPR adaptation. Here we show that guanine-substitutions in the target strand of the protospacer are highly detrimental to CRISPR interference and interference-dependent priming, while cytosine-substitutions are more readily tolerated. Furthermore, we show that this effect is based on strongly decreased binding affinity of the effector complex Cascade for guanine-mismatched targets, while cytosine-mismatched targets only minimally affect target DNA binding. Structural modeling of Cascade-bound targets with mismatches shows that steric clashes of mismatched guanines lead to unfavorable conformations of the RNA-DNA duplex. This effect has strong implications for the natural selection of target site mutations that lead to effective escape from type I CRISPR-Cas systems.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Künne T,Zhu Y,da Silva F,Konstantinides N,McKenzie RE,Jackson RN,Brouns SJ

doi

10.1093/nar/gky687

subject

Has Abstract

pub_date

2018-11-02 00:00:00

pages

10395-10404

issue

19

eissn

0305-1048

issn

1362-4962

pii

5070486

journal_volume

46

pub_type

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