Abstract
The CRISPR system for prokaryotic adaptive immunity provides RNA-mediated protection from viruses and mobile genetic elements. When viral RNA transcripts are detected, type III systems adopt an activated state that licenses DNA interference and synthesis of cyclic oligoadenylate (cOA). cOA activates nucleases and transcription factors that orchestrate the antiviral response. We demonstrate that cOA synthesis is subject to tight temporal control, commencing on target RNA binding, and is deactivated rapidly as target RNA is cleaved and dissociates. Mismatches in the target RNA are well tolerated and still activate the cyclase domain, except when located close to the 3' end of the target. Phosphorothioate modification reduces target RNA cleavage and stimulates cOA production. The 'RNA shredding' activity originally ascribed to type III systems may thus be a reflection of an exquisite mechanism for control of the Cas10 subunit, rather than a direct antiviral defence.
Highlights
CRISPR systems provide adaptive immunity in prokaryotes against mobile genetic elements (MGE)
S. solfataricus Csm generates cyclic oligoadenylate (cOA) in response to target RNA binding cOA synthesis has so far been observed directly in two bacterial type III systems (Kazlauskiene et al, 2017; Niewoehner et al, 2017)
We have demonstrated that the type III-D Csm complex from S. solfataricus generates cOA on binding target RNA
Summary
CRISPR systems provide adaptive immunity in prokaryotes against mobile genetic elements (MGE). DNA sequences derived from MGE are incorporated into the host genome separated by short direct repeats, forming the CRISPR locus. The HD-nuclease domain of the large Cas subunit is responsible for target DNA degradation (Elmore et al, 2016; Estrella et al, 2016; Kazlauskiene et al, 2016; Jung et al, 2015; Han et al, 2017a) This is activated when target RNA is bound by the complex (Samai et al, 2015), providing a mechanism for transcription-dependent DNA targeting (Deng et al, 2013) that allows lysogenic phage to persist in the host chromosome (Goldberg et al, 2014). Type III CRISPR systems are much more tolerant of mismatches with nucleic acid targets than other types, making viral escape difficult (Pyenson et al, 2017; Manica et al, 2011; Manica et al, 2013)
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