RNA polymerase II CTD phosphopeptides compete with RNA for the interaction with Pcf11.
In Saccharomyces cerevisiae, the cleavage/polyadenylation factor Pcf11 is an important regulatory factor required for recruiting the polyadenylation machinery to the elongating RNA polymerase II (RNAPII) and is necessary for correct transcriptional termination. The interaction with RNAPII is mediated by a CTD-interacting domain (CID) located in the N-terminal region of Pcf11 that binds in a phospho-dependent manner the heptad repeats in the RNAPII CTD. We have previously investigated this protein-protein interaction. We examine here the interaction of the CID with different RNA sequences and look at the effect of phosphopeptides derived from the CTD heptad repeats on the RNA-protein interaction. Our findings demonstrate that the CID displays weak RNA-binding activity, but with some degree of sequence preference, the RNA-protein and peptide-protein interfaces overlap and the CTD-derived phosphopeptides and RNA compete for the binding site. We propose that competition between the protein-peptide and the protein-RNA interaction is important mechanistically and required for the disengagement of polyadenylation factors from RNAPII.
Division of Molecular Structure, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, United Kingdom.
David
D
Hollingworth
Christian G
CG
Noble
Ian A
IA
Taylor
Andres
A
Ramos
D001483:Base Sequence; D001667:Binding, Competitive; D019906:Nuclear Magnetic Resonance, Biomolecular; D012319:RNA Polymerase II / Q000737:chemistry / Q000378:metabolism*; D012331:RNA, Fungal / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D020349:Surface Plasmon Resonance; D039221:mRNA Cleavage and Polyadenylation Factors / Q000378:metabolism*
C512637:PCF11 protein, S cerevisiae; D012331:RNA, Fungal; D029701:Saccharomyces cerevisiae Proteins; D039221:mRNA Cleavage and Polyadenylation Factors; D012319:RNA Polymerase II