Genetic interactions within TFIIIC, the promoter-binding factor of yeast RNA polymerase III

Abstract.

TFIIIC is a heteromultimeric protein, made of six distinct subunits in Saccharomyces cerevisiae, that binds to RNA polymerase III promoters and triggers the assembly of the transcription complex. The largest yeast subunit tau138, encoded by TFC3, binds to the B-box promoter element. This binding is defective in the temperature-sensitive mutant tfc3-G349E; the mutation responsible is located in one of two conserved motifs shared with the B-binding component of human TFIIIC. Rare dominant gain-of-function mutations that restore growth at high temperature were obtained following ultraviolet mutagenesis of tfc3-G349E. All of them resulted from single amino acid substitutions that alter the structure of TFIIIC. Three were due to reversion or intragenic suppression (TFC3-K754E and TFC3-L804H) events. Three were identical isolates of TFC6-E330K, a previously described mutation of the tau91 subunit. The remaining suppressors mapped in TFC4, and resulted in amino acid replacements in the second largest subunit of TFIIIC (tau131). With the exception TFC4-E711K, these affect positions that are invariant between the S. cerevisiae and Homo sapiens proteins, and are localised in conserved tetratricopeptide motifs. These findings demonstrate a close functional interaction between the two largest subunits of TFIIIC and underscore the importance of the tetratricopeptide motif of tau131.