Cryo-EM Structures Reveal Mechanism and Inhibition of DNA Targeting by a CRISPR-Cas Surveillance Complex

Author:

Guo, Tai Wei

Bartesaghi, Alberto

Yang, Hui

Falconieri, Veronica

Rao, Prashant

Merk, Alan

Eng, Edward T

Raczkowski, Ashleigh M

Fox, Tara

Earl, Lesley A

Patel, Dinshaw J

Subramaniam, Sriram
Author Address

Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA., Structural Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA., Simons Electron Microscopy Center, New York Structural Biology Center, New York, NY 10027, USA., Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA; Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Frederick, MD 21701, USA., Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA; Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20892, USA. Electronic address: subramas@mail.nih.gov.,

Abstract:

Prokaryotic cells possess CRISPR-mediated adaptive immune systems that protect them from foreign genetic elements, such as invading viruses. A central element of this immune system is an RNA-guided surveillance complex capable of targeting non-self DNA or RNA for degradation in a sequence- and site-specific manner analogous to RNA interference. Although the complexes display considerable diversity in their composition and architecture, many basic mechanisms underlying target recognition and cleavage are highly conserved. Using cryoelectron microscopy (cryo-EM), we show that the binding of target double-stranded DNA (dsDNA) to a type I-F CRISPR system yersinia (Csy) surveillance complex leads to large quaternary and tertiary structural changes in the complex that are likely necessary in the pathway leading to target dsDNA degradation by a trans-acting helicase-nuclease. Comparison of the structure of the surveillance complex before and after dsDNA binding, or in complex with three virally encoded anti-CRISPR suppressors that inhibit dsDNA binding, reveals mechanistic details underlying target recognition and inhibition. Published by Elsevier Inc.

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