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Title: A newly discovered Bordetella species carries a transcriptionally active CRISPR-Cas with a small Cas9 endonuclease

Author
item IVANOV, YURY - Pennsylvania State University
item SHARIAT, NIKKI - Pennsylvania State University
item Register, Karen
item RIVERA, ISRAEL - Pennsylvania State University
item LINZ, BODO - Pennsylvania State University
item HU, K - Pennsylvania State University
item DUDLEY, EDWARD - Pennsylvania State University
item HARVILL, ERIC - Pennsylvania State University

Submitted to: BMC Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/21/2015
Publication Date: 10/26/2015
Publication URL: http://handle.nal.usda.gov/10113/61642
Citation: Ivanov, Y.V., Shariat, N., Register, K.B., Rivera, I., Linz, B., Hu, K., Dudley, E.G., Harvill, E.T. 2015. A newly discovered Bordetella species carries a transcriptionally active CRISPR-Cas with a small Cas9 endonuclease. Biomed Central (BMC) Genomics. 16(1):863. doi: 10.1186/s12864-015-2028-9.

Interpretive Summary: Approximately 40% of all bacteria have a DNA segment known as a Clustered Regularly Interspersed Short Palindromic Repeat, or CRISPR. These CRISPRs, along with proteins arising from nearby “cas” genes, provide immunity to the bacteria and may, in some cases, regulate the activity of genes in the host. Bacteria found among the Bordetella are well known respiratory disease agents. Bordetella pertussis and Bordetella parapertussis cause whooping cough in humans while other members of the Bordetella cause respiratory disease in a variety of mammals and some avian species. No CRISPR-Cas system has yet been identified in any Bordetella species. Here we report the identification and characterization of a novel, functional CRISPR-Cas system in 12 different isolates of a single, newly discovered species of Bordetella, tentatively designated Bordetella psudohinzii. This work provides new insights into the evolution and pathogenicity of Bordetella species. The B. psudohinzii CRISPR-Cas system has the potential to add significantly to the tools available for genome engineering.

Technical Abstract: The Cas9 endonuclease of the Type II-a clustered regularly interspersed short palindromic repeats (CRISPR), of Streptococcus pyogenes (SpCas9) has been adapted as a widely used tool for genome editing and genome engineering. Herein, we describe a gene encoding a novel Cas9 ortholog (BpsuCas9) and the surrounding Type II-c CRISPR-cas locus found exclusively in a single, newly discovered Bordetella species, tentatively designated B. psudohinzii. The cas genes and the CRISPR array are transcribed during both logarithmic and stationary growth in vitro. Several of the spacer sequences in this locus are homologous to mobile genetic elements, including prophages, tRNA-Gly, and a transposase of the IS3/IS911 family, suggesting this is a functional CRISPR-Cas system that provides adaptive immunity against invading mobile elements. Nucleotide content analysis suggests that this system was laterally acquired from an unknown source with a GC-content lower than the genome-wide average for B. psudohinzii. Its insertion point in B. psudohinzii is an apparent recombination hotspot. BpsuCas9 shares ~75% pair-wise amino acid identity with the Cas9 of Alicycliphilus denitrificans, but only ~23% identity with SpCas9, the Cas9 endonuclease most frequently used for genome editing. Together these observations indicate that this single Bordetella species has a novel CRISPR-cas locus that is functional and may be of additional utility in genome editing.