A Functional Mini-Integrase in a Two-Protein-type V-C CRISPR System

Addison V. Wright, Joy Y. Wang, David Burstein, Lucas B. Harrington, David Paez-Espino, Nikos C. Kyrpides, Anthony T. Iavarone, Jillian F. Banfield, Jennifer A. Doudna*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

CRISPR-Cas immunity requires integration of short, foreign DNA fragments into the host genome at the CRISPR locus, a site consisting of alternating repeat sequences and foreign-derived spacers. In most CRISPR systems, the proteins Cas1 and Cas2 form the integration complex and are both essential for DNA acquisition. Most type V-C and V-D systems lack the cas2 gene and have unusually short CRISPR repeats and spacers. Here, we show that a mini-integrase comprising the type V-C Cas1 protein alone catalyzes DNA integration with a preference for short (17- to 19-base-pair) DNA fragments. The mini-integrase has weak specificity for the CRISPR array. We present evidence that the Cas1 proteins form a tetramer for integration. Our findings support a model of a minimal integrase with an internal ruler mechanism that favors shorter repeats and spacers. This minimal integrase may represent the function of the ancestral Cas1 prior to Cas2 adoption. Wright et al. demonstrate the function of an integrase comprised of a single protein, Cas1, that inserts unusually short fragments of viral DNA into the bacterial genome as part of the CRISPR-Cas adaptive immunity pathway. This minimal integrase may represent the function of the ancestral Cas1 prior to Cas2 adoption.

Original languageEnglish
Pages (from-to)727-737.e3
JournalMolecular Cell
Volume73
Issue number4
DOIs
StatePublished - 21 Feb 2019
Externally publishedYes

Funding

FundersFunder number
DOE Office of Science User FacilityDE-AC02-05CH11231
NIH S10 OD018174 instrumentationS10 OD018174
NIH T32 0666981S10OD020062-01, T32 066698
US Department of Energy Joint Genome Institute
US National Science Foundation
Vincent J. Coates Genomics Sequencing Laboratory at UC Berkeley
National Science Foundation1244557
NIH Office of the DirectorS10OD020062
Norges Idrettshøgskole

    Keywords

    • CRISPR
    • integrase
    • protein-DNA recognition
    • spacer acquisition

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