In response to androgen stimuli, SC-3 cells cloned from mouse mammary carcinoma (Shionogi carcinoma 115) secrete heparin-binding growth factor, which is able to bind to fibroblast growth factor (FGF) receptor, ultimately resulting in SC-3 cell proliferation. A role for glycosaminoglycan in the process of this androgen-induced autocrine loop was examined. When quiescent SC-3 cells were stimulated with testosterone, enhanced DNA synthesis was demonstrated even at the early phase (12-24 h) of stimulation. During this early period, autocrine growth-promoting activity was found to be associated with cell surface and extracellular matrix, but not to be present in the conditioned medium. This bound form of growth-promoting activity was able to be extracted with 2 M NaCl (pH 7.5) and absorbed onto a heparin-Sepharose affinity column, from which it was eluted at a concentration of 1.1-1.3 M NaCl. Extracted growth factor, whose activity was partially blocked by anti-basic FGF antibody, up-regulated the expression of FGF receptor-1 mRNA. These characteristics were similar to those of a soluble form of SC-3-derived growth factor previously reported from our laboratory. Androgen-induced enhancement of DNA synthesis was inhibited by simultaneous treatment of SC-3 cells with 10 mIU/ml heparitinase or 25 mM sodium chlorate (an inhibitor of phosphoadenosine sulfate synthesis). However, chlorate treatment did not affect the synthesis and distribution of androgen-induced growth factor, suggesting that the response of chlorate-treated cells to this growth factor was impaired. These results indicate that heparan sulfate has important roles in concentrating androgen-induced heparin-binding growth factor on or very close to the cell surface and in potentiating its bioactivity.