DnaQ exonuclease‐like domain of Cas2 promotes spacer integration in a type I‐E CRISPR‐Cas system. (11th June 2018)

• Record Type: Journal Article
• Authors: Drabavicius, Gediminas; Sinkunas, Tomas; Silanskas, Arunas; Gasiunas, Giedrius; Venclovas, Česlovas; Siksnys, Virginijus
• Is Part Of: EMBO reports. Volume 19:Number 7(2018)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2018-06-11
• Subjects: adaptation -- Cas1 -- Cas2 -- protospacer -- Streptococcus thermophilus; Molecular biology -- Periodicals; Molecular Biology -- Periodicals; Molecular biology; Periodicals; 572.8
• DOI: 10.15252/embr.201745543
• Languages: English
• ISSNs: 1469-221X

Abstract

Abstract: CRISPR‐Cas systems constitute an adaptive immune system that provides acquired resistance against phages and plasmids in prokaryotes. Upon invasion of foreign nucleic acids, some cells integrate short fragments of foreign DNA as spacers into the CRISPR locus to memorize the invaders and acquire resistance in the subsequent round of infection. This immunization step called adaptation is the least understood part of the CRISPR‐Cas immunity. We have focused here on the adaptation stage of Streptococcus thermophilus DGCC7710 type I‐E CRISPR4‐Cas (St4) system. Cas1 and Cas2 proteins conserved in nearly all CRISPR‐Cas systems are required for spacer acquisition. The St4 CRISPR‐Cas system is unique because the Cas2 protein is fused to an additional DnaQ exonuclease domain. Here, we demonstrate that St4 Cas1 and Cas2‐DnaQ form a multimeric complex, which is capable of integrating DNA duplexes with 3′‐overhangs (protospacers) in vitro . We further show that the DnaQ domain of Cas2 functions as a 3′–5′‐exonuclease that processes 3′‐overhangs of the protospacer to promote integration. Synopsis: The CRISPR4‐Cas system of the Streptococcus thermophilus spacer integration complex contains the Cas2‐DnaQ fusion protein. This study shows that the DnaQ domain is required for the processing of the protospacers prior to integration into the CRISPR array. The DnaQ domain fused to Cas2 in the S. thermophilus CRISPR4‐Cas system is a 3′–5′ exonuclease. The DnaQ domain in the Cas1:Cas2‐DnaQ complex trims 3′ ends of protospacers to generate overhangs optimal for integration. Cas1:Cas2‐DnaQ integrates trimmed protospacers into the CRISPR array. Abstract : The CRISPR4‐Cas system of the Streptococcus thermophilus spacer integration complex contains the Cas2‐DnaQ fusion protein. This study shows that the DnaQ domain is required for the processing of the protospacers prior to integration into the CRISPR array.