CDC73P AND PAF1P ARE FOUND IN A NOVEL RNA-POLYMERASE II-CONTAINING COMPLEX DISTINCT FROM THE SRBP-CONTAINING HOLOENZYME

The products of the yeast CDC73 and PAF1 genes, were originally identi fied as RNA polymerase II-associated proteins, Paf1p is a nuclear prot ein important for cell growth and transcriptional regulation of a subs et of yeast genes. In this study we demonstrate that the product of CD C73 is a nuclear protein that interacts directly with purified RNA pol ymerase II in vitro. Deletion of CDC73 confers a temperature-sensitive phenotype, Combination of the cdc73 mutation with the more severe paf 1 mutation does not result in an enhanced phenotype, indicating that t he two proteins may function in the same cellular processes, To determ ine the relationship between Cdc73p and Paf1p and the recently describ ed holoenzyme form of RNA polymerase II, we created yeast strains cont aining glutathione S-transferase (GST)-tagged forms of CDC73, PAF1, an d TFGZ functionally replacing the chromosomal copies of the genes, Iso lation of GST-tagged Cdc73p and Paf1p complexes has revealed a unique form of RNA polymerase II that contains both Cdc73p and Paf1p but lack s the Srbps found in the holoenzyme, The Cdc73p-Paf1p-RNA polymerase I I-containing complex also includes Gal11p, and the general initiation factors TFIIB and TFIIF, but lacks TBP, TFIIH, and transcription elong ation factor TFIIS as well as the Srbps, The Srbp-containing holoenzym e does not include either Paf1p or Cdc73p, demonstrating that these tw o forms of RNA polymerase II are distinct, In confirmation of the hypo thesis that the two forms coexist in yeast cells, we found that a TFII F-containing complex isolated via the GST-tagged Tfg2p construct conta ins both (i) the Srbps and (ii) Cdc73p and Paf1p. The Srbps and Cdc73p -Paf1p therefore appear to define two complexes with partially redunda nt, essential functions in the yeast cell, Using the technique of diff erential display, we have identified several genes whose transcripts r equire Cdc73p and/or Paf1p for normal levels of expression, Our analys is suggests that there are multiple RNA polymerase II-containing compl exes involved in the expression of different classes of protein-coding genes.