Title: Function of the Rpb4 subunit of RNA polymerase II during transcription
Description: The RNA polymerase II enzyme from the yeast Saccharomyces cerevisiae is a complex of 12 subunits, Rpb1-12, that can dissociate into a 10-subunit catalytic core and a heterodimer of the Rpb4 and Rpb7 subunits. While the functions of the 10-subunit core in transcription have been intensively studied, less is understood about the role of the Rpb4/7 heterodimer. Crystal structures of the complete 12-subunit RNA polymerase II show that the Rpb4/7 heterodimer is found near both the transcript-exit groove of the enzyme and the C-terminal domain of Rpb1, which serves as an important docking location for the various proteins recruited at different stages of the transcription cycle. This placement of the heterodimer within the context of the complete enzyme would allow for interactions with the nascent RNA transcript as well as protein factors involved in transcription regulation. To further characterize the role of the Rpb4/7 heterodimer during transcription in vivo , we have used chromatin immunoprecipitation to map the cross-linking patterns of RNA polymerase II and multiple associated factors along actively transcribed genes in both wild-type and rpb4Δ strains. We have found that loss of Rpb4 results in a reduction of core RNA polymerase II levels found towards the 3' ends of mRNA genes. This correlates with a loss of recruitment of the 3'-end processing factors Rna14 and Rna15 in our rpb4Δ strain, as well as a defect in cotranscriptional 3'-end processing at the RNA14 gene.

Additionally, we have found that loss of Rpb4 results in changes to chromatin structure along actively transcribed genes. Recruitment of the chromatin-associated factors Spt6 and Spt2 is lost in an rpb4Δ strain. Furthermore, loss of Rpb4 results in altered histone H3 occupancy and modification in transcribed regions. These results suggest that the Rpb4/7 heterodimer is required for proper cotranscriptional 3'-end processing as well as the maintenance of chromatin structure at transcribed genes. Overall, our findings demonstrate that the function of Rpb4 is not only important to the behavior of the polymerase itself, but also serves to regulate crucial interactions between the polymerase and its associated factors.