REGULATION OF TRANSFER-RNA SUPPRESSOR ACTIVITY BY AN INTRON-ENCODED POLYADENYLATION SIGNAL

A 26-nt sequence from the 3' UTR of the yeast GAL7 mRNA directs accura te and efficient cleavage and polyadenylation to form the 3' end of th e GAL7 mRNA in vivo and in vitro. Here we asked whether this polyadeny lation signal can function within the context of a tRNA. Insertion of the GAL7 signal into the intron of the dominant SUP4 nonsense suppress or allowed us to judge the effect of the insert on SUP4 function by ob servation of nonsense suppression efficiency in vivo. The GAL7 signal impairs the function of SUP4 in an orientation-dependent manner in viv o, consistent with its ability to specify cleavage and polyadenylation in this context in vitro. Mutation of a UA repeat within the GAL7 sig nal restores SUP4 function partially, consistent with the role of this repeat as an efficiency element in polyadenylation. Mutations that im pair the mRNA 3' end-processing factors Rna14p and Rna15p restore supp ressor function partially. Northern blot analysis, PCR amplification, and DNA sequence analysis show that the GAL7 signal directs polyadenyl ation within the body of pre-SUP4 and within the terminator, suggestin g that polyadenylation inhibits 5' and 3' end processing, as well as r emoval of the pre-tRNA intron. These findings indicate that the GAL7 p olyadenylation signal is capable of targeting a pre-tRNA to the mRNA p rocessing pathway.