RNA-Guided RNA Cleavage by a CRISPR RNA-Cas Protein Complex

Abstract

SUMMARY Compelling evidence indicates that the CRISPR-Cas system protects prokaryotes from viruses and other potential genome invaders. This adaptive prokaryotic immune system arises from the clustered regularly interspaced short palindromic repeats (CRISPRs) found in prokaryotic genomes, which harbor short invader-derived sequences, and the CRISPR-associated (Cas) protein-coding genes. Here we have identified a CRISPR-Cas effector complex that is comprised of small invader-targeting RNAs from the CRISPR loci (termed prokaryotic silencing (psi)RNAs) and the RAMP module (or Cmr) Cas proteins. The psiRNA-Cmr protein complexes cleave complementary target RNAs at a fixed distance from the 3' end of the integral psiRNAs. In Pyrococcus furiosus, psiRNAs occur in two size forms that share a common 5' sequence tag but have distinct 3' ends that direct cleavage of a given target RNA at two distinct sites. Our results indicate that prokaryotes possess a unique RNA silencing system that functions by homology-dependent cleavage of invader RNAs.

The impact of CRISPR repeat sequence on structures of a Cas6 protein–RNA complex

“…The molecular process of new DNA incorporation remains completely unknown while the obstruction phase of the pathway has been shown to depend on crRNA. [13][14][15][16] CRISPR RNAs are predicted to interact and function with an impressive number of repeatassociated mysterious protein (RAMP) proteins found across different CRISPR subtypes. 9,10,17 Despite their prevalence in CRISPR functional pathways, how RAMP proteins interact with RNA remains partially understood.…”

“…The crRNAassociated ribonucleoprotein particle, called Cmr complex, recently isolated from P. furiosus was demonstrated to have crRNA-guided RNA cleavage activity. 15 The Cmr complex is comprised of six proteins (Cmr1-Cmr6), four of which (Cmr1, Cmr3, Cmr4, and Cmr6) are RAMP proteins. These RAMP proteins have been shown to be essential to RNA silencing activity.…”

Diversity, evolution, and therapeutic applications of small RNAs in prokaryotic and eukaryotic immune systems

“…The Csa, Cst, Csh, and Csm subtype systems are common in Archaea, and all of the Cas systems are found in bacteria except for Csa. The broad range of variation in which Cas genes are present in a given CRISPR-Cas system suggests that a variety of pathways are used to carry out CRISPR-Cas immune functions, although the differences in the utility of these systems as a result of their unique pathways is unknown [64].…”

“…Cas3 can associate with COG2254 family proteins. One such COG2254 protein, the Cas gene SSO200, is an endonuclease that shows no preference for the cleavage of dsRNA over dsDNA, and may partner with Cas3 protein SSO1999 in order to silence foreign invaders [64,69].…”

“…Comparisons of Cas gene sequences, adaptation mechanisms, and crRNA processing mechanisms have demonstrated that even though Bacteria and Archaea represent two entirely different domains of life, and Archaea is more closely related to eukaryotes than bacteria, the CRISPR-Cas systems found in these two domains are indeed very similar [63,64,98].…”