Addition of purified plasmin or plasminogen (0.1 microM) to serum-free culture media elevated cellular D-myo-inositol 1,4,5-trisphosphate (InsP3) levels in human colorectal carcinoma cells within 1 h to double those of control cells. This was accompanied by decreases in cellular phosphatidylinositol bisphosphate by 40% in cells exposed to fibrinolytic ligands for up to 1 h. The effect was not due to opening of Ca2+ channels of the type blocked by 5 microM nifedipine, and 100 microM EGTA, a Ca2+ chelator, did not suppress plasmin's ability to elevate InsP3. Binding assays at 4 degrees C with 125I-labelled plasmin indicated maximum binding within 1 h suggesting that the effects of plasmin may be associated with its cell-binding function. These cells could convert exogenous plasminogen to plasmin with endogenous activation and this was accompanied by a decrease in radioactive phosphatidylinositol well below control levels (13% of control). Our results contribute to evidence for the association of plasmin-binding sites with a signalling system. A cell signalling system indirectly or directly associated with plasmin binding, would permit carcinoma cells to coordinate extracellular fibrinolysis with cell migration and motility through second messengers.