@article{daa74a52c62e4d0688aa303874ded62c,
title = "Viral recombination systems limit CRISPR-Cas targeting through the generation of escape mutations",
abstract = "CRISPR-Cas systems provide immunity to bacteria by programing Cas nucleases with RNA guides that recognize and cleave infecting viral genomes. Bacteria and their viruses each encode recombination systems that could repair the cleaved viral DNA. However, it is unknown whether and how these systems can affect CRISPR immunity. Bacteriophage λ uses the Red system (gam-exo-bet) to promote recombination between related phages. Here, we show that λ Red also mediates evasion of CRISPR-Cas targeting. Gam inhibits the host E. coli RecBCD recombination system, allowing recombination and repair of the cleaved DNA by phage Exo-Beta, which promotes the generation of mutations within the CRISPR target sequence. Red recombination is strikingly more efficient than the host's RecBCD-RecA in the production of large numbers of phages that escape CRISPR targeting. These results reveal a role for Red-like systems in the protection of bacteriophages against sequence-specific nucleases, which may facilitate their spread across viral genomes.",
keywords = "CRISPR, Cas9, Lambda, Red, recombination",
author = "Hossain, {Amer A.} and Jon McGinn and Meeske, {Alexander J.} and Modell, {Joshua W.} and Marraffini, {Luciano A.}",
note = "Funding Information: We thank all members of the Marraffini lab for helpful discussion and encouragement. We thank B. Levin for providing the λvir phage; T.K. Wood for the E. coli K-12 MG1655 Δ9 strain; the Coli Genetic Stock Center at Yale University for E. coli Keio knockout strains; K.C. Murphy for the pKM208(red) plasmid; C.A. Voigt for the E. coli ACT-01 strain; A.Z. Fire for the pPD207.846 plasmid; K. Severinov and E. Semenova for the E. coli KD263 strain and KD263 λ-targeting strains (λ_E4-R, λ_L1-R, λ_L4-R, λ_L6-R); J.W. Roberts for the MG1655::λ strain; and the Rockefeller University Genomics Resource Center for assistance with next generation sequencing experiments. J.W.M. was supported by a Jane Coffin Childs Memorial Fund for Medical Research postdoctoral fellowship. A.J.M. was a Helen Hay Whitney postdoctoral fellow. L.A.M. is supported by a Burroughs Wellcome Fund PATH Award, and an NIH Director's Pioneer Award (DP1GM128184). L.A.M. is an investigator at the Howard Hughes Medical Institute. A.A.H. J.W.M. and L.A.M. designed and conceived the study. A.A.H. and J.W.M. performed all experiments with help from J.M. A.J.M. constructed plasmid pAM38. A.A.H. J.W.M. and L.A.M. wrote the manuscript. L.A.M. is a cofounder and Scientific Advisory Board member of Intellia Therapeutics and a cofounder of Eligo Biosciences. Funding Information: J.W.M. was supported by a Jane Coffin Childs Memorial Fund for Medical Research postdoctoral fellowship. A.J.M. was a Helen Hay Whitney postdoctoral fellow. L.A.M. is supported by a Burroughs Wellcome Fund PATH Award, and an NIH Director{\textquoteright}s Pioneer Award (DP1GM128184). L.A.M. is an investigator at the Howard Hughes Medical Institute . Publisher Copyright: {\textcopyright} 2021 Elsevier Inc.",
year = "2021",
month = oct,
day = "13",
doi = "10.1016/j.chom.2021.09.001",
language = "English (US)",
volume = "29",
pages = "1482--1495.e12",
journal = "Cell Host and Microbe",
issn = "1931-3128",
publisher = "Cell Press",
number = "10",
}