title: CRISPR adaptation biases explain preference for acquisition of foreign DNA.
abstract: CRISPR-Cas (clustered, regularly interspaced short palindromic repeats coupled with CRISPR-associated proteins) is a bacterial immunity system that protects against invading phages or plasmids. In the process of CRISPR adaptation, short pieces of DNA ('spacers') are acquired from foreign elements and integrated into the CRISPR array. So far, it has remained a mystery how spacers are preferentially acquired from the foreign DNA while the self chromosome is avoided. Here we show that spacer acquisition is replication-dependent, and that DNA breaks formed at stalled replication forks promote spacer acquisition. Chromosomal hotspots of spacer acquisition were confined by Chi sites, which are sequence octamers highly enriched on the bacterial chromosome, suggesting that these sites limit spacer acquisition from self DNA. We further show that the avoidance of self is mediated by the RecBCD double-stranded DNA break repair complex. Our results suggest that, in Escherichia coli, acquisition of new spacers largely depends on RecBCD-mediated processing of double-stranded DNA breaks occurring primarily at replication forks, and that the preference for foreign DNA is achieved through the higher density of Chi sites on the self chromosome, in combination with the higher number of forks on the foreign DNA. This model explains the strong preference to acquire spacers both from high copy plasmids and from phages.
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Asaf
authors: A
authors: Levy
authors: Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors: Moran G
authors: MG
authors: Goren
authors: Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors: Ido
authors: I
authors: Yosef
authors: Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors: Oren
authors: O
authors: Auster
authors: Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors: Miriam
authors: M
authors: Manor
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Gil
authors: G
authors: Amitai
authors: Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors: Rotem
authors: R
authors: Edgar
authors: Department of Clinical Microbiology and Immunology, Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors: Udi
authors: U
authors: Qimron
authors: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors: Rotem
authors: R
authors: Sorek
mesh_terms: D000222:Adaptation, Physiological; D001435:Bacteriophages / Q000235:genetics*; D064113:CRISPR-Cas Systems / Q000235:genetics; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D016384:Consensus Sequence / Q000235:genetics; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004261:DNA Replication / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics*; D004279:DNA, Viral / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D043211:Exodeoxyribonuclease V / Q000378:metabolism; D008954:Models, Biological; D010957:Plasmids / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial; D004279:DNA, Viral; D043211:Exodeoxyribonuclease V