PCF11 ENCODES A 3RD PROTEIN-COMPONENT OF YEAST CLEAVAGE AND POLYADENYLATION FACTOR-I

Cleavage and polyadenylation factor I (CF I) is one of four factors re quired in vitro for yeast pre-mRNA 3'-end processing, Two protein comp onents of this factor, encoded by genes RNA14 and RNA15, have already been identified, We describe here another gene, PCF11 (for protein 1 o f CF I), that genetically interacts with RNA14 and RNA15 and which pre sumably codes for a third protein component of CF I. This gene was iso lated in a two-hybrid screening designed to identify proteins interact ing with Rna14 and Rna15. PCF11 is an essential gene encoding for a pr otein of 626 amino acids having an apparent molecular mass of 70 kDa. Thermosensitive mutations in PCF11 are synergistically lethal with the rmosensitive alleles of RNA14 and RNA15. The Pcf11-2 thermosensitive s train shows a shortening of the poly(A) tails and a strong decrease in the steady-state level of actin transcripts after a shift to the nonp ermissive temperature as do the thermosensitive alleles of RNA14 and R NA15. Extracts from the pcf11-1 and pcf11-2 thermosensitive strains an d the wild-type strain, when Pcf11 is neutralized by specific antibodi es, are deficient in cleavage and polyadenylation, Moreover, fractions obtained by anion exchange chromatography of extracts from the wild-t ype strain contain both Pcf11 and Rna15 in the same fractions, as show n by immunoblotting with a Pcf11-specific antibody.