The C-Terminal Domain of Rpb1 Functions on Other RNA Polymerase II Subunits

Abstract

SUMMARY The C-terminal domain (CTD) of Rpb1, the largest subunit of RNA polymerase II (RNApII), coordinates recruitment of RNA- and chromatin-modifying factors to transcription complexes. It is unclear whether the CTD communicates with the catalytic core region of Rpb1 and thus must be physically connected, or instead can function as an independent domain. To address this question, CTD was transferred to other RNApII subunits. Fusions to Rpb4 or Rpb6, two RNApII subunits located near the original position of CTD, support viability in a strain carrying a truncated Rpb1. In contrast, CTD fusion to Rpb9 on the other side of RNApII does not. Rpb4-CTD and Rpb6-CTD proteins are functional for phosphorylation and recruitment of various factors, albeit with some restrictions and minor defects. Normal CTD functions are not transferred to RNApI or RNApIII by Rbp6-CTD. These results show that, with some spatial constraints, CTD can function even when disconnected from Rpb1.

Structure of a Complete Mediator-RNA Polymerase II Pre-Initiation Complex

“…The CTD path leading to TFIIK explains why Mediator enhances the rate of phosphorylation by TFIIH in vitro (Kim et al, 1994). Further consistent with the structure, splicing the CTD onto either Rpb4 or Rpb6 near its point of exit from the surface of the wild type enzyme rescued a lethal Rpb1-ΔCTD phenotype in yeast, whereas splicing onto Rpb9, on the opposite side of the enzyme, failed to rescue the lethal phenotype (Suh et al, 2013). These CTD variants, which contained a modified linker region, showed diminished levels of phosphorylation by TFIIK, possibly due to an influence of linker length and topology on the kinetics of CTD phosphorylation.…”

Determinants of Histone H3K4 Methylation Patterns

“…Rpb4 was chosen because the Rpb4-Rpb7 “stalk” protrudes away from the Pol II core, close to the presumed position of the CTD. This should be an acceptable location for CTD binding proteins, as the CTD itself can be transferred from Rpb1 to Rpb4 without loss of function (Suh et al, 2013). Unfortunately, Rpb4 fused to full-length Set1 was unstable (Fig 5A, lane 7), consistent with observations that Set1 levels are kept low through protein degradation signals in the N-terminal domains (Soares et al, 2014).…”

The Evolutionarily Conserved Pol II Flap Loop Contributes to Proper Transcription Termination on Short Yeast Genes

“…Interestingly, the CTD can be relocated to other Pol II subunits and still support viability, provided the CTD remains on the same face of Pol II (Suh et al, 2013). This observation suggests that the CTD must be spatially coordinated with other Pol II domains located at defined positions on the surface of Pol II and supports the hypothesis that a dual CTD-Pol II body interface supports termination factor recruitment.…”