Event Title
NuA4 interaction with RNA polymerase II is stimulated by phosphorylation of CTD serines 2 and 5
Faculty Mentor
Dr. Daniel Ginsburg
Major/Area of Research
CLS
Description
Transcription of protein-coding genes is carried out by RNA polymerase II (Pol II). The largest of Pol II’s 12 subunits (Rpb1) has a long C-terminal domain (CTD). The CTD in yeast is composed of 26 repeats of the heptapeptide Y1S2P3T4S5P6S7. Post-translational modification of the CTD is important for recruiting protein complexes to gene bodies. We investigated the importance of serine 2 and serine 5 phosphorylation (Ser2p/Ser5p) for interaction between the CTD and NuA4. NuA4 is the only essential lysine acetyltransferase complex in yeast and has been shown to be important for both transcription initiation and elongation. We have previously proposed that NuA4 occupancy in gene bodies is stimulated by interaction with the CTD phosphorylated on serine 5, but were not able to observe binding to Ser5p peptides in vitro. In this study, we examined NuA4 binding to Pol II in mutants affecting the CTD kinases Kin28, Bur1, and Ctk1. Kin28 phosphorylates serines 5 and 7, while Bur1 and Ctk1 phosphorylate serine 2. We found that NuA4 interaction with Pol II as measured by coimmunoprecipitation was reduced in kin28 mutant cells. Consistent with this, NuA4 occupancy at the ADH1 3’ ORF was also reduced in kin28 cells. Interestingly, loss of Ser2p in either bur2Δ (the cyclin for Bur1) or ctk1Δ led to reduced NuA4 binding to Pol II as well as decreased occupancy across GAL1. These results provide the first evidence that NuA4 may be able to bind both the Ser2p and Ser5p forms of the Pol II CTD
NuA4 interaction with RNA polymerase II is stimulated by phosphorylation of CTD serines 2 and 5
Transcription of protein-coding genes is carried out by RNA polymerase II (Pol II). The largest of Pol II’s 12 subunits (Rpb1) has a long C-terminal domain (CTD). The CTD in yeast is composed of 26 repeats of the heptapeptide Y1S2P3T4S5P6S7. Post-translational modification of the CTD is important for recruiting protein complexes to gene bodies. We investigated the importance of serine 2 and serine 5 phosphorylation (Ser2p/Ser5p) for interaction between the CTD and NuA4. NuA4 is the only essential lysine acetyltransferase complex in yeast and has been shown to be important for both transcription initiation and elongation. We have previously proposed that NuA4 occupancy in gene bodies is stimulated by interaction with the CTD phosphorylated on serine 5, but were not able to observe binding to Ser5p peptides in vitro. In this study, we examined NuA4 binding to Pol II in mutants affecting the CTD kinases Kin28, Bur1, and Ctk1. Kin28 phosphorylates serines 5 and 7, while Bur1 and Ctk1 phosphorylate serine 2. We found that NuA4 interaction with Pol II as measured by coimmunoprecipitation was reduced in kin28 mutant cells. Consistent with this, NuA4 occupancy at the ADH1 3’ ORF was also reduced in kin28 cells. Interestingly, loss of Ser2p in either bur2Δ (the cyclin for Bur1) or ctk1Δ led to reduced NuA4 binding to Pol II as well as decreased occupancy across GAL1. These results provide the first evidence that NuA4 may be able to bind both the Ser2p and Ser5p forms of the Pol II CTD