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

During exon ligation, the Saccharomyces cerevisiae spliceosome recognizes the 3′-splice site (3′SS) of precursor messenger RNA (pre-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-angstrom cryo–electron microscopy structure of a human postcatalytic 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′SS selection and to potentially fine-tune alternative splicing at the exon ligation stage.