The E2F transcription factor plays a major role in cell cycle regulation, differentiation and apoptosis, but it is not clear how it is regulated by non‐mitogenic signaling cascades. Here we report that two kinases involved in signal transduction have opposite effects on E2F function: the stress‐induced kinase JNK1 inhibits E2F1 activity whereas the related p38 kinase reverses Rb‐mediated repression of E2F1. JNK1 phosphorylates E2F1 in vitro, and co‐transfection of JNK1 reduces the DNA binding activity of E2F1; treatment of cells with TNFα had a similar effect. Fas stimulation of Jurkat cells is known to induce p38 kinase and we find a pronounced increase in Rb phosphorylation within 30 min of Fas stimulation. Phosphorylation of Rb correlated with a dissociation of E2F and increased transcriptional activity. The inactivation of Rb by Fas was blocked by SB203580, a p38‐specific inhibitor, as well as a dominant‐negative p38 construct; cyclin‐dependent kinase (cdk) inhibitors as well as dominant‐negative cdks had no effect. These results suggest that Fas‐mediated inactivation of Rb is mediated via the p38 kinase, independent of cdks. The Rb/E2F‐mediated cell cycle regulatory pathway appears to be a normal target for non‐mitogenic signaling cascades and could be involved in mediating the cellular effects of such signals.