Mechanisms of spliceosome remodeling and splice site selection during the catalytic stage of mammalian pre-mRNA splicing (360G-Wellcome-220212_Z_20_Z)

£1,199,999

The spliceosome produces mRNAs in two sequential transesterifications – branching and exon ligation. The post-branching C complex is remodeled by the ATPase Prp16 into the C* complex. Prp16 allows binding of the exon ligation factors Cactin and FAM32A, which stabilise docking of the 3’-splice site (3’SS) at the active site. While in yeast Prp18 is constitutively bound to C* spliceosomes, in humans Prp18 and other exon ligation factors of unknown function appear to engage the C* spliceosome in a transcript-specific manner. Here, I will reconstitute the C to C* transition in vitro in mammals and use biochemistry and cryo-electron microscopy to isolate and visualize novel intermediates and dissect the structural mechanisms by which Cactin, FAM32A, Prp18, and additional exon ligation factors cooperate to promote Prp16-mediated remodeling and to ensure correct 3’SS choice. In parallel, iCLIP studies will reveal where these exon ligation factors bind across all human pre-mRNAs and how RNA sequences around the splice sites determine the association of subsets of factors on specific pre-mRNAs. This work will elucidate how Prp16 drives remodeling of the human catalytic spliceosome and will investigate whether exon ligation factors regulate alternative splicing in a sequence-specific manner while proofreading 3'SS recognition during exon ligation.

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Grant Details

Amount Awarded 1199999
Applicant Surname Fica
Approval Committee Sir Henry Dale Fellowship Interview Committee
Award Date 2020-03-04T00:00:00+00:00
Financial Year 2019/20
Grant Programme: Title Sir Henry Dale Fellowship
Internal ID 220212/Z/20/Z
Lead Applicant Dr Sebastian Fica
Partnership Name Royal Society/Wellcome Trust Sir Henry Dale Fellowship
Partnership Value 1199999
Planned Dates: End Date 2026-04-30T00:00:00+00:00
Planned Dates: Start Date 2021-05-01T00:00:00+00:00
Recipient Org: Country United Kingdom
Region South East
Sponsor(s) Prof Francis Barr