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Internal translation of large subunit transcripts drives small subunit synthesis in type I CRISPR-Cas interference complexes

By Tess M. McBride, Evan A. Schwartz, Abhishek Kumar, David W Taylor, Peter C. Fineran, Robert D. Fagerlund

Posted 18 Apr 2020
bioRxiv DOI: 10.1101/2020.04.18.045682

CRISPR-Cas adaptive immune systems provide bacteria and archaea with defense against their viruses and other mobile genetic elements. CRISPR-Cas immunity involves the formation of ribonucleoprotein complexes that specifically bind and degrade foreign nucleic acids. Despite advances in the biotechnological exploitation of select systems, multiple CRISPR-Cas types remain uncharacterized. Here, we investigated a type I-D system from Synechocystis and revealed the Cascade complex, which is required for interference, forms a hybrid ribonucleoprotein complex that is structurally and genetically related to both type I and III systems. Surprisingly, the type I-D complex contained multiple functionally-important small subunit proteins encoded from an internal in-frame translation initiation site within the large subunit gene, cas10d . Structural analysis revealed that these small subunits bound the complex similarly to Cas11 small subunits in other type I and III systems, where they are encoded as a separate gene. We show that internal translation of small subunits from within large subunit genes is conserved across diverse type I-D, I-B and I-C systems, which account for ~23% of all sequenced CRISPR-Cas systems. Indeed, we demonstrate that small subunits are expressed from within the cas8c large subunit gene in the Desulfovibrio vulgaris type I-C system. Our work reveals an unexpected aspect of CRISPR-Cas evolution and expansion of the coding potential from within single cas genes. ### Competing Interest Statement The authors have declared no competing interest.

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