The RNA polymerase II C-terminal domain promotes splicing activation through recruitment of a U2AF65-Prp19 complex

Pre-mRNA splicing is frequently coupled to transcription by RNA polymerase II (RNAPII). This coupling requires the C-terminal domain of the RNAPII largest subunit (CTD), although the underlying mechanism is poorly understood. Using a biochemical complementation assay, we previously identified an activity that stimulates CTD-dependent splicing in vitro. We purified this activity and found that it consists of a complex of two well-known splicing factors: U2AF65 and the Prp19 complex (PRP19C). We provide evidence that both U2AF65 and PRP19C are required for CTD-dependent splicing activation, that U2AF65 and PRP19C interact both in vitro and in vivo, and that this interaction is required for activation of splicing. Providing the link to the CTD, we show that U2AF65 binds directly to the phosphorylated CTD, and that this interaction results in increased recruitment of U2AF65 and PRP19C to the pre-mRNA. Our results not only provide a mechanism by which the CTD enhances splicing, but also describe unexpected interactions important for splicing and its coupling to transcription.