Cross Talk between tRNA and rRNA synthesis in Saccharomyces cerevisiae

Temperature sensitive RNA polymerase In (rpc160-112 and rpc160-270) mutants were analyzed for the synthesis of tRNAs and rRNAs in vivo, using a double -isotopic-labeling technique in which cells are pulse-labeled with [P-33]or thophosphate and coextracted with [H-3]uracil-labeled wild-type cells. Indi vidual RNA species were monitored by Northern blot hybridization or amplifi ed by reverse transcription. These mutants impaired the synthesis of RNA po lymerase III transcripts with little or no influence on mRNA synthesis but also largely turned off the formation of the 25S, 18S, and 5.8S mature rRNA species derived from the common 35S transcript produced by RNA polymerase I. In the rpc160-270 mutant, this parallel inhibition of tRNA and rRNA synt hesis also occurred at the permissive temperature (25 degreesC) and correla ted with an accumulation of 20S pre-rRNA, In the rpc160-112 mutant, inhibit ion of rRNA synthesis and the accumulation of 20S pre-rRNA were found only at 37 degreesC. The steady-state rRNA/tRNA ratio of these mutants reflected their tRNA and rRNA synthesis pattern: the rpc160-112 mutant had the three fold shortage in tRNA expected from its preferential defect in tRNA synthes is at 25 degreesC, whereas rpc160-270 cells completely adjusted their rRNA/ tRNA ratio down to a wild-type level, consistent with the tight coupling of tRNA and rRNA synthesis in vivo. Finally, an RNA polymerase I (rpa190-2) m utant grown at the permissive temperature had an enhanced level of pre-tRNA , suggesting the existence of a physiological coupling between rRNA synthes is and pre-tRNA processing.