GENETIC INTERACTION BETWEEN U6 SNRNA AND THE FIRST INTRON NUCLEOTIDE IN SACCHAROMYCES-CEREVISIAE

Nuclear pre-mRNA splicing necessitates specific recognition of the pre -mRNA splice sites. It is known that 5' splice site selection requires base pairing of U6 snRNA with intron positions 4-6. However, no facto r recognizing the highly conserved 5' splice site GU has yet been iden tified. We have tested if the known U6 snRNA-pre-mRNA interaction coul d be extended to include the first intron nucleotides and the conserve d (50)GAG(52) sequence of U6 snRNA. We observe that some combinations of 5' splice site and Us snRNA mutations produce a specific synthetic block to the first splicing step. In addition, the U6-G52U allele can switch between two competing 5' splice sites harboring different nucle otides following the cleavage site. These results indicate that U6 snR NA position 52 interacts with the first nucleotide of the intron befor e 5' splice site cleavage. Some combinations of U6 snRNA and pre-mRNA mutations also blocked the second splicing step, suggesting a role for the corresponding nucleotides in a proofreading step before exon liga tion. From studies in diverse organisms, various functions have been a scribed to the conserved U6 snRNA (47)ACAGAG(52) sequence. Our results suggest that these discrepancies might reflect variations between dif ferent experimental systems and point to an important conserved role o f this sequence in the splicing reaction.