Pta1, a component of yeast CFII, is required for both cleavage and poly(A)addition of mRNA precursor

CF II, a factor required for cleavage of the 3' ends of mRNA precursor in S accharomyces cerevisiae, has been shown to contain four polypeptides. The t hree largest subunits, Cft1/Yhh1, Cft2/Ydh1, and Brr5/Ysh1, are homologs of the three largest subunits of mammalian cleavage-polyadenylation specifici ty factor (CPSF), an activity needed for both cleavage and poly(A) addition . In this report, we show by protein sequencing and immunoreactivity that t he fourth subunit of CF II is Pta1, an essential 90-kDa protein originally implicated in tRNA splicing. Yth1, the yeast homolog of the CPSF 30-kDa sub unit, is not detected in this complex. Extracts prepared from pta1 mutant s trains are impaired in the cleavage and the poly(A) addition of both GAL7 a nd CYC1 substrates and exhibit little processing activity even after prolon ged incubation. However, activity is efficiently rescued by the addition of purified CF II to the defective extracts. Extract from a strain with a mut ation in the CF IA subunit Rna14 also restored processing, but extract from a brr5-1 strain did not. The amounts of Pta1 and other CF II subunits are reduced in pta1 strains, suggesting that levels of the subunits may be coor dinately regulated. Coimmunoprecipitation experiments indicate that the CF Ii in extract can be found in a stable complex containing Pap1, CF II, and the Fip1 and Yth1 subunits of polyadenylation factor I. While purified CF I I does not appear to retain the association with these other factors, this larger complex may be the form recruited onto pre-mRNA in vivo. The involve ment of Pta1 in both steps of mRNA 3'-end formation supports the conclusion that CF II is the functional homolog of CPSF.