A human postcatalytic spliceosome structure reveals essential roles of metazoan factors for exon ligation.

During exon ligation, the S. cerevisiae spliceosome recognizes the 3'-splice site (3'SS) of precursor mRNA through non-Watson-Crick pairing with the 5'SS and the branch adenosine, in a conformation stabilized by Prp18 and Prp8. Here we present the 3.3 Å cryoEM structure of a human post-catalytic spliceosome just after exon ligation. The 3'SS docks at the active site through conserved RNA interactions in the absence of Prp18. Unexpectedly, the metazoan-specific FAM32A directly bridges the 5'-exon and intron 3'SS of pre-mRNA and promotes exon-ligation, as shown by functional assays. CACTIN, SDE2, and NKAP - factors implicated in alternative splicing - further stabilize the catalytic conformation of the spliceosome during exon ligation. Together these four proteins act as exon ligation factors. Our study reveals how the human spliceosome has co-opted additional proteins to modulate a conserved RNA-based mechanism for 3'-splice site selection and to potentially fine-tune alternative splicing at the exon ligation stage.