title: DNA motifs determining the accuracy of repeat duplication during CRISPR adaptation in Haloarcula hispanica.
abstract: Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) acquire new spacers to generate adaptive immunity in prokaryotes. During spacer integration, the leader-preceded repeat is always accurately duplicated, leading to speculations of a repeat-length ruler. Here in Haloarcula hispanica, we demonstrate that the accurate duplication of its 30-bp repeat requires two conserved mid-repeat motifs, AACCC and GTGGG. The AACCC motif was essential and needed to be ∼10 bp downstream from the leader-repeat junction site, where duplication consistently started. Interestingly, repeat duplication terminated sequence-independently and usually with a specific distance from the GTGGG motif, which seemingly served as an anchor site for a molecular ruler. Accordingly, altering the spacing between the two motifs led to an aberrant duplication size (29, 31, 32 or 33 bp). We propose the adaptation complex may recognize these mid-repeat elements to enable measuring the repeat DNA for spacer integration.
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Ming
authors.initials: M
authors.lastname: Li
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Luyao
authors.initials: L
authors.lastname: Gong
authors.affiliation: Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Songnian
authors.initials: S
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China xiangh@im.ac.cn.
authors.forename: Hua
authors.initials: H
authors.lastname: Xiang
mesh_terms: D001483:Base Sequence; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D004261:DNA Replication; D019641:DNA, Archaeal / Q000235:genetics*; D020440:Gene Duplication; D019766:Genes, Archaeal; D019613:Haloarcula / Q000235:genetics*; D059372:Nucleotide Motifs; D017422:Sequence Analysis, DNA
chemical_list: D019641:DNA, Archaeal