BACKGROUND The human prostate carcinoma cell line, LNCaP, proliferates under stimulation by a limited number of mitogenic signals, which include members of the growth factor and steroid hormone families. Androgens and epidermal growth factor (EGF) are among the LNCaP cell mitogens. We tested the hypothesis that these mitogens stimulate LNCaP cell proliferation at least in part through the induction of cyclin D1, a protein requisite for cell cycle progression, which is expressed in the G1 phase of the cell cycle. METHODS LNCaP cells were grown in serum-free medium with 10 ng/ml or 100 ng/ml EGF, 0.1 nM or 1.0 nM mibolerone (a potent androgen agonist), or vehicle (distilled water or 0.01% ethanol). Expression of cyclin D, mRNA, and protein were assessed by Northern and Western blot analyses. Transcription regulation was assessed by nuclear runoff assay. RESULTS Western analyses demonstrated that EGF stimulated cyclin D1 protein expression 4-fold over 12 hr. Northern analyses showed a 4-fold increase in mRNA expression, peaking within 4 hr of EGF stimulation. There were no effects on cyclin D1 protein or mRNA expression with mibolerone treatments. We further explored the mechanism of cyclin D1 induction. LNCaP cells stimulated for 1 hr with EGF demonstrated a 2-fold increase in cyclin D1 message, as assayed by nuclear runoff transcription assay. In addition, we demonstrated the involvement of the protein kinase C pathway in mediating the EGF induction of cyclin D1. CONCLUSIONS We conclude that one of the mechanisms by which growth factors such as EGF may stimulate prostate cell proliferation is through the direct induction of cyclin proteins, which are necessary for entry of cells into mitosis.