Yeast Pab1 Interacts with Rna15 and Participates in the Control of the Poly(A) Tail Length In Vitro

Abstract

In Saccharomyces cerevisiae, the single poly(A) binding protein, Pab1, is the major ribonucleoprotein associated with the poly(A) tails of mRNAs in both the nucleus and the cytoplasm. We found that Pab1 interacts with Rna15 in two-hybrid assays and in coimmunoprecipitation experiments. Overexpression of I partially but specifically suppressed the rna15-2 mutation in vivo. RNA15 codes for a component of the cleavage and polyadenylation factor CF I, one of the four factors needed for pre-mRNA 3′-end processing. We show that Pab1 and CF I copurify in anion-exchange chromatography. These data suggest that Pab1 is physically associated with CF I. Extracts from a thermosensitive pab1 mutant and from a wild-type strain immunoneutralized for Pab1 showed normal cleavage activity but a large increase in poly(A) tail length. A normal tail length was restored by adding recombinant Pab1 to the mutant extract. The longer poly(A) tails were not due to an inhibition of exonuclease activities. Pab1 has previously been implicated in the regulation of translation initiation and in cytoplasmic mRNA stability. Our data indicate that Pab1 is also a part of the 3′-end RNA-processing complex and thus participates in the control of the poly(A) tail lengths during the polyadenylation reaction.