INDUCTION OF DROSOPHILA RNA POLYMERASE-III GENE-EXPRESSION BY THE PHORBOL ESTER 12-O-TETRADECANOYLPHORBOL-13-ACETATE (TPA) IS MEDIATED BY TRANSCRIPTION FACTOR-IIIB

We have previously found that the phorbol ester 12-O-tetradecanoylphor bol-13-acetate (TPA) induces specific transcription of tRNA and 5S RNA genes in Drosophila Schneider S-2 cells (M. Garber, S. Panchanathan, R. F. Fan, and D. L. Johnson, J. Biol. Chem. 266:20598-20601, 1991). H aving derived cellular extracts from TPA-treated cells, that are capab le of reproducing this stimulation in vitro, we have examined the mech anism for this regulatory event. Using conditions that limit reinitiat ion and produce single rounds of transcription from active gene comple xes, we find that the number of functional transcription complexes is increased in extracts prepared from TPA-induced cells. We have analyze d the activities of the transcription factors TFIIIB and TFIIIC derive d from extracts prepared from TPA-induced and noninduced cells. Examin ation of the relative activities of TFIIIC showed that both its abilit y to reconstitute transcription with TFIIIB and RNA polymerase Ill and its ability to stably bind to the DNA template are unchanged. However , the activity of TFIIIB derived from the TPA-induced cells is substan tially increased compared with that derived from the noninduced cells. The differences in TFIIIB activity account for the differences in the overall transcriptional activities observed in the unfractionated ext racts. Western blot analysis of the TATA-binding protein subunit of TF IIIB revealed that there is an increase in the amount of this polypept ide present in the induced cell extracts and TFIIIB fraction. Together , these results indicate that the TPA response in Drosophila cells sti mulates specific transcription of RNA polymerase III genes by increasi ng the activity of the limiting transcription component, TFIIIB, and t hereby increasing the number of functional transcription complexes.