We report in vitro studies showing that tRNA gene transcription in yeast is down-regulated during the transition from logarithmic to stationary phase growth. Transcription in a postdiauxic (early stationary) phase extract of a wild-type strain decreased 3-fold relative to a log phase extract. This growth stage-related difference in transcription was amplified to 20-fold in extracts of a strain containing a mutation (pcf1-4) in the 131-kDa subunit of TFIIIC. The reduction in transcription activity in both wild-type and mutant postdiauxic phase extracts was correlated with a decrease in the amount of TFIIIB70, the limiting factor in these extracts. However, the 3.7 +/- 0.5-fold decrease in amount of TFIIIB70 in mutant extracts does not, by itself, account for the 20-fold decrease in transcription. Accordingly, transcription in the mutant postdiauxic phase extract could be reconstituted to a level equal to the mutant log phase extract by the addition of two components, TFIIIB70 and TFIIIC. Addition of TFIIIB70 increased transcription 10-fold, while a 2-fold effect of TFIIIC was seen at saturating levels of TFIIIB70. The data suggest that both TFIIIB70 and TFIIIC play a role in coordinating the level of polymerase III transcription with cell growth rate.