Structural basis for conformational equilibrium of the catalytic spliceosome

Abstract

The ATPase Prp16 governs equilibrium between the branching (B^∗/C) and exon ligation (C^∗/P) conformations of the spliceosome. Here, we present the electron cryomicroscopy reconstruction of the Saccharomyces cerevisiae C-complex spliceosome at 2.8 Å resolution and identify a novel C-complex intermediate (C_i) that elucidates the molecular basis for this equilibrium. The exon-ligation factors Prp18 and Slu7 bind to C_i before ATP hydrolysis by Prp16 can destabilize the branching conformation. Biochemical assays suggest that these pre-bound factors prime the C complex for conversion to C^∗ by Prp16. A complete model of the Prp19 complex (NTC) reveals how the branching factors Yju2 and Isy1 are recruited by the NTC before branching. Prp16 remodels Yju2 binding after branching, allowing Yju2 to remain tethered to the NTC in the C^∗ complex to promote exon ligation. Our results explain how Prp16 action modulates the dynamic binding of step-specific factors to alternatively stabilize the C or C^∗ conformation and establish equilibrium of the catalytic spliceosome.

Keywords: ATPase; RNA; cryo-EM; spliceosome; splicing; thermodynamic equilibrium.