Seminars Archive

CRISPR-Cas machines: structural electron microscopy provides crucial functional insight

Abstract
The CRISPR (clustered regularly interspaced short palindromic repeats) and Cas (CRISPR-associated) genes are widely spread in bacteria and archaea, representing an intracellular defence system against invading viruses and plasmids. In the system, fragments from foreign nucleic acid are captured and integrated into the host genome at the CRISPR locus. The locus is transcribed and the resulting RNAs are processed by Cas6 into small crRNAs (CRISPR RNAs) that guide a variety of effector complexes to degrade the invading genetic elements. CRISPR machines are currently in the limelight for their application in gene editing. Recent work has shown that Type II CRISPR/Cas systems can be engineered to direct targeted double-stranded DNA breaks in vitro to specific sequences by using a single "guide RNA" with complementarity to the DNA target site and a Cas9 nuclease (Jinek et al., Science 2012). This targetable Cas9-based system also works efficiently in cultured human cells (Mali et al., Science 2013; Cong et al., Science 2013) and in vivo in zebrafish (Hwang and Fu et al., Nature Biotechnology 2013) and mice (Wang et al., Cell 2013) for inducing targeted alterations into endogenous genes. Structural biology is providing crucial information on the function and mechanism of these nanomachines. In my talk, I will present electron microscopy coupled to image processing work, which we performed to unravel the general mechanism of crRNA interference, targeting both non-self RNA (CMR complex) and DNA (CSM complex). In my talk, I will present electron microscopy coupled to image processing work, which we performed to unravel the general mechanism of crRNA interference, targeting both non-self RNA (CMR complex) and DNA (CSM complex).