MULTIPLE ROLES FOR U6 SNRNA IN THE SPLICING PATHWAY

作者：MADHANI HD^*; BORDONNE R; GUTHRIE C

来源：Genes & Development, 1990, 4(12B): 2264-2277.

DOI：10.1101/gad.4.12b.2264

• SACCHAROMYCES-CEREVISIAE
• SPLICING
• U6 SNRNA
• U4 SNRNA

U6 is the most highly conserved of the five spliceosomal RNAs. It is associated with U4 by an extensive base-pairing interaction, which is disrupted immediately prior to the first nucleolytic step of splicing. It has been proposed that this event activates catalysis by unmasking U6. Using a combination of doped synthesis and site-directed mutagenesis to generate point mutations in U6, we have now identified 12 positions, in three domains, at which single nucleotide substitutions or deletions result in lethal or temperature-sensitive phenotypes. Biochemical analysis demonstrates that most of these mutants retain the ability to assemble into U4/U6 and U4/U5/U6 snRNPs. Notably although mutations at three positions in U6 that base-pair with U4 are lethal, mutations in the complementary residues in U4 are fully viable. Furthermore, compensatory mutations in U4 that restore base-pairing fail to suppress the phenotypes of the U6 mutations. This demonstrates a function for U6 independent of its role in base-pairing. Remarkably, two of the three essential regions in U6 identified genetically correspond to intron insertion points in two yeast species. A temperature-sensitive mutation at one of these sites is defective in the second step of splicing in vitro.

• 出版日期1990-12