The effect of hydrocortisone on the transcriptional activity of exponentially and parasynchronously growing WI38 cells was measured using E. coli RNA polymerase or endogenous chromatinbound RNA polymerase to catalyse RNA synthesis in vitro. Results indicated that the transcriptional activity of cells was increased 8 h after hydrocortisone addition to exponentially growing cultures and 3–5 h after hormone addition to parasynchronously growing cultures. The hydrocortisone-induced alteration of transcriptional activity was not an artifact induced by different rates of RNA or DNA, degradation. These findings indicate that the hormone-dependent alteration of transcriptional activity is an early event in the hydrocortisone-mediated stimulation of WI38 cell proliferation.