THE RNA polymerase II holoenzyme consists of RNA polymerase II, a subset of general transcription factors, and regulatory proteins known as SRB proteins(1,2). The genes encoding SRB proteins were isolated as suppressors of mutations in the RNA polymerase II carboxy-terminal domain (CTD3,4). The CTD and SRB proteins have been implicated in the response to transcriptional regulators(1-11). We report here the isolation of two new SRB genes, SRB10 and SRB11, which encode kinase- and cyclin-like proteins, respectively, Genetic and biochemical evidence indicates that the SRB10 and SRB11 proteins form a kinase-cyclin pair in the holoenzyme, The SRB10/11 kinase is essential for a normal transcriptional response to galactose induction in vivo. Holoenzymes lacking SRB10/11 kinase function are strikingly deficient in CTD phosphorylation, Although defects in the kinase substantially affect transcription in vivo, purified holoenzymes lacking SRB10/11 kinase function do not show defects in defined in vitro transcription systems, suggesting that the factors necessary to elicit the regulatory role of the SRB10/11 kinase are missing in these systems, These results indicate that the SRB10/11 kinase is involved in CTD phosphorylation and suggest that this modification has a role in the response to transcriptional regulators in vivo.