Tyrosine Phosphorylation of the Carboxyl-Terminal Domain in RNA Polymerase II Regulates Binding of RPRD1B

The carboxyl-terminal domain (CTD) of the mammalian RNA polymerase II consists of 52 heptapeptide repeats with the consensus sequence YSPTSPS. As the CTD undergoes phosphorylation, RNA polymerase II initiates transcription and acts as a scaffold for recruitment of pre-mRNA processing proteins to the nascent RNA to  perform processes such as 5’ capping, alternative splicing, 3’ polyadenylation, and cleavage. RPRD1B, which stands for regulation of nuclear pre-mRNA domain containing 1B, is a 36kD nuclear protein that contains a CTD-interacting domain (CID). We have found that RPRD1B interacts with serine-phosphorylated (pSer) CTD, consistent with previous findings that CID binds to pSer-CTD. Interestingly we found that RPRD1B interaction with the CTD is inhibited when the CTD becomes tyrosine phosphorylated (pTyr).  We have generated antibodies that react with pTyr/pSer-CTD or pTyr-CTD and shown that these antibodies do not react with pSer-CTD. Using these antibodies, we have detected pTyr-CTD on endogenous RNA polymerase II in mammalian cells.  Together, our results suggest that the in vivo interaction between RPRD1B and RNA polymerase II can be positively and negatively regulated by the differential phosphorylation of Tyr and/or Ser on the CTD.