Adaptation mechanisms in CRISPR-Cas immunity

Hayun Lee and Dipali G. Sashital

Roy J. Carver Department of Biochemistry, Biophysics & Molecular Biology, Iowa State University, Ames, IA USA


CRISPR–Cas systems provide immunological memory of prior infections to bacteria and archaea. Host cells store genetic information from viruses and mobile genetic elements in CRISPR arrays, enabling rapid immune response upon subsequent infections by the invader. The basis of memory formation is the integration of short segments of invader DNA as spacers within the CRISPR array, a process called adaptation. The Cas1-Cas2 integration complex is found universally amongst all CRISPR–Cas systems, and recent biochemical and structural studies have elucidated the molecular mechanism of spacer integration. Other core cas genes, such as Cas4, have been implicated in the adaptation stage, but their exact functions have remained elusive. Here, we show that Cas4 directly interacts with Cas1, enabling formation of a Cas4-Cas1-Cas2 complex. While the Cas1-Cas2 complex integrates optimal substrate DNA into the CRISPR locus, the complex is unable to efficiently process and integrate longer precursor substrates. Addition of Cas4 to the Cas1-Cas2 complex enables substrate processing through the combined endonucleolytic activity of Cas4 and Cas1, and significantly enhances integration activity into longer CRISPR loci. Together, these data establish Cas4 as an integral member of the Type I-C adaptation complex and answer long-standing questions about the mechanism of substrate processing prior to spacer integration.


Dipali Sashital is an assistant professor in the Department of Biochemistry, Biophysics & Molecular Biology at Iowa State University. She graduated with a BS from the University of Michigan, and received her PhD from the University of Wisconsin, Madison, in 2006. Following postdoctoral studies at University of California, Berkeley and The Scripps Research Institute, La Jolla, Dr. Sashital joined the faculty at Iowa State in January, 2014. Her primary research interests are the structure and function of RNA-protein complexes, and specifically the mechanisms of CRISPR-Cas RNA-guided adaptive immunity.