Cyclin-dependent kinase inhibitors (cdkis), such as p21, are believed to control proliferation through an ability to function as stoichiometric antagonists of cyclin-dependent kinases (cdks). The p21 gene is a direct transcriptional target for the p53 protein, and its activation is likely to be important in effecting the p53 response. It is widely accepted that p21 can influence cell cycle progression by controlling the activity of cdks that act on the retinoblastoma tumour suppressor protein (pRb) which, in a hypophosphorylated state, associates with E2F transcription factors to prevent the activation of genes required for progression into S phase. Phosphorylation of pRb by G1 cdk complexes releases E2F and thereby enables progress through the cell cycle. Here, we describe results which suggest a p21-dependent mechanism that facilitates the regulation of E2F through a pathway that is independent of the cdk control of pRb activity. As p21 can associate with E2F subunits, it is possible that these effects are exerted through a complex with E2F. Furthermore, we find that p21 can regulate transcription in vitro. The results suggest that p21 may control E2F activity through a pathway that acts independently of pRb.