TRANSFER-RNA NUCLEAR EXPORT IN SACCHAROMYCES-CEREVISIAE - IN-SITU HYBRIDIZATION ANALYSIS

To understand the factors specifically affecting tRNA nuclear export, we adapted in situ hybridization procedures to locate endogenous level s of individual tRNA families in wild-type and mutant yeast cells. Our studies of tRNAs encoded by genes lacking introns show that nucleopor in Nup116p affects both poly(A) RNA and tRNA export, whereas Nup159p a ffects only poly(A) RNA export. Los1p is similar to exportin-t, which facilitates vertebrate tRNA export. A los1 deletion mutation affects t RNA but not poly(A) RNA export. The data support the notion that Los1p and exportin-t are functional homologues. Because LOS1 is nonessentia l, tRNA export in vertebrate and yeast cells likely involves factors i n addition to exportin-t. Mutation of RNA1, which encodes RanGAP, caus es nuclear accumulation of tRNAs and poly(A) RNA. Many yeast mutants, including those with the rna1-1 mutation, affect both pre-tRNA splicin g and RNA export. Our studies of the location of intron-containing pre -tRNAs in the rna1-1 mutant rule out the possibility that this results from tRNA export occurring before splicing. Our results also argue ag ainst inappropriate subnuclear compartmentalization causing defects in pre-tRNA splicing. Rather, the data support ''feedback'' of nucleus/c ytosol exchange to the pre-tRNA splicing machinery.