Pre-mRNA splicing is an essential step of eukaryotic gene expression carried out by a series of dynamic macromolecular protein/RNA complexes, known collectively and individually as the spliceosome. This series of spliceosomal complexes define, assemble on, and catalyze the removal of introns. Molecular model snapshots of intermediates in the process have been created from cryo-EM data, however, many aspects of the dynamic changes that occur in the spliceosome are not fully understood. Caenorhabditis elegans follow the GU-AG rule of splicing, with almost all introns beginning with 5′ GU and ending with 3′ AG. These splice sites are identified early in the splicing cycle, but as the cycle progresses and ″custody″ of the pre-mRNA splice sites is passed from factor to factor as the catalytic site is built, the mechanism by which splice site identity is maintained or re-established through these dynamic changes is unclear. We performed a genetic screen in C. elegans for factors that are capable of changing 5′ splice site choice. We report that KIN17 and PRCC are involved in splice site choice, the first functional splicing role proposed for either of these proteins. Previously identified suppressors of cryptic 5′ splicing promote distal cryptic GU splice sites, however, mutations in KIN17 and PRCC instead promote usage of an unusual proximal 5′ splice site which defines an intron beginning with UU, separated by 1nt from a GU donor. We performed high-throughput mRNA sequencing analysis and found that mutations in PRCC but not KIN17 changed 5′ splice sites genome-wide, promoting usage of nearby non-consensus sites. We further found that mutations in KIN17 and PRCC changed dozens of 3′ splice sites, promoting non-consensus sites upstream of canonical splice sites. Our work has uncovered both fine and coarse mechanisms by which the spliceosome maintains splice site identity during the complex assembly process.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.