TRANSCRIPTIONAL ACTIVATION DOMAINS STIMULATE INITIATION AND ELONGATION AT DIFFERENT TIMES AND VIA DIFFERENT RESIDUES

Transcriptional activators can stimulate multiple steps in the transcr iption process. We have used GAL4 fusion proteins to characterize the ability of different transcriptional activation domains to stimulate t ranscriptional elongation on the hsp70 gene irt vitro. Stimulation of elongation apparently occurs via a mechanistic pathway different from that of stimulation of initiation: the herpes simplex virus VP16, heat shock factor 1 (HSF1) and amphipathic helix (AH) activation domains a ll stimulate initiation, but only VP16 and HSF1 stimulate elongation; and mutations in hydrophobic residues of the HSF1 activation domains i mpair stimulation of elongation but not of initiation, while mutations in adjacent acidic residues impair stimulation of initiation more tha n of elongation. Experiments in which activators were exchanged betwee n initiation and elongation demonstrate that the elongation function o f HSF1 will stimulate RNA polymerase that has initiated and is transcr iptionally engaged. Transcriptional activators thus appear to have at least two distinct functions that reside in the same domain, and that act at different times to stimulate initiation and elongation.