title: A dual function of the CRISPR-Cas system in bacterial antivirus immunity and DNA repair.
abstract: Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) and the associated proteins (Cas) comprise a system of adaptive immunity against viruses and plasmids in prokaryotes. Cas1 is a CRISPR-associated protein that is common to all CRISPR-containing prokaryotes but its function remains obscure. Here we show that the purified Cas1 protein of Escherichia coli (YgbT) exhibits nuclease activity against single-stranded and branched DNAs including Holliday junctions, replication forks and 5'-flaps. The crystal structure of YgbT and site-directed mutagenesis have revealed the potential active site. Genome-wide screens show that YgbT physically and genetically interacts with key components of DNA repair systems, including recB, recC and ruvB. Consistent with these findings, the ygbT deletion strain showed increased sensitivity to DNA damage and impaired chromosomal segregation. Similar phenotypes were observed in strains with deletion of CRISPR clusters, suggesting that the function of YgbT in repair involves interaction with the CRISPRs. These results show that YgbT belongs to a novel, structurally distinct family of nucleases acting on branched DNAs and suggest that, in addition to antiviral immunity, at least some components of the CRISPR-Cas system have a function in DNA repair.
authors.affiliation: Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario, M5G 1L6, Canada.
authors.forename: Mohan
authors.initials: M
authors.lastname: Babu
authors.forename: Natalia
authors.initials: N
authors.lastname: Beloglazova
authors.forename: Robert
authors.initials: R
authors.lastname: Flick
authors.forename: Chris
authors.initials: C
authors.lastname: Graham
authors.forename: Tatiana
authors.initials: T
authors.lastname: Skarina
authors.forename: Boguslaw
authors.initials: B
authors.lastname: Nocek
authors.forename: Alla
authors.initials: A
authors.lastname: Gagarinova
authors.forename: Oxana
authors.initials: O
authors.lastname: Pogoutse
authors.forename: Greg
authors.initials: G
authors.lastname: Brown
authors.forename: Andrew
authors.initials: A
authors.lastname: Binkowski
authors.forename: Sadhna
authors.initials: S
authors.lastname: Phanse
authors.forename: Andrzej
authors.initials: A
authors.lastname: Joachimiak
authors.forename: Eugene V
authors.initials: EV
authors.lastname: Koonin
authors.forename: Alexei
authors.initials: A
authors.lastname: Savchenko
authors.forename: Andrew
authors.initials: A
authors.lastname: Emili
authors.forename: Jack
authors.initials: J
authors.lastname: Greenblatt
authors.forename: Aled M
authors.initials: AM
authors.lastname: Edwards
authors.forename: Alexander F
authors.initials: AF
authors.lastname: Yakunin
mesh_terms: D064130:CRISPR-Associated Proteins; D003090:Coliphages / Q000254:growth & development*; D018360:Crystallography, X-Ray; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003851:Deoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000821:virology*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid
chemical_list: D064130:CRISPR-Associated Proteins; D029968:Escherichia coli Proteins; D050505:Mutant Proteins; D003851:Deoxyribonucleases; D004706:Endodeoxyribonucleases; C557650:YgbT protein, E coli; D045643:DNA Repair Enzymes