Mutations in STS1 suppress the defect in 3' mRNA processing caused by the rna15-2 mutation in Saccharomyces cerevisiae

Abstract

In a search for proteins associated with Rna15p in processing the 3' ends of messenger RNAs, we have looked for suppressors that correct, even partially, the thermosensitive growth defect of the rna15-2 mutant. Mutations in a single locus that we named SSM5, were able to suppress both the thermosensitivity of cell growth and the mRNA 3' processing defect associated with the rna15-2 mutation, but only slightly alleviated the thermosensitive growth defect of an rna14-1 mutant. The ssm5-1 mutant is sensitive to hydroxyurea at 37 degrees C, a drug that inhibits DNA synthesis. By screening for complementation of the hydroxyurea-sensitive phenotype we cloned the corresponding wild-type gene and found that it corresponds to the essential gene STS1 (also named DBF8). Sts1p has an apparent molecular weight of 30 kDa and was confirmed to be a cytosolic protein by immunofluorescence analysis. Western blot analysis indicates that the thermosensitive mutant strains rna15-2, rna14-1 and pap1-1 present a very low level of the Rna15p at 37 degrees C. The ssm5-1 mutation restores the level of Rna15p in the rna15-2 ssm5-1 double mutant. Use of the two-hybrid system suggests that Sts1p does not interact directly with Rna15p, but may be active as a homodimer. The present data suggest that Sts1p may play a role in the transport of Rna15p from the cytoplasm to the nucleus.