Urokinase-dependent adhesion loss and shape change after cyclic adenosine monophosphate elevation in cultured rat mesangial cells

Mesangial cells in culture change shape and become less adhesive in response to cAMP elevation (e.g., treatment with isoproterenol plus isobutylmethylxanthine (IM). Inhibitors of serine proteases inhibit cellular shape change in response to IM. To further examine the role of cell surface proteases in shape change, adhesion plaque proteins (i.e., preparations of ventral membranes and extracellular matrix) were separated in SDS-polyacrylamide gels containing gelatin with and without plasminogen. Four discrete zones of lysis were evident in plasminogen gels (indicative of activation of plasminogen) from control adhesion plaques: one inconspicuous zone with a M(r) ~ 150 kD, another at ~ 115 kD, and a doublet at ~ 35-32 kD. Another diffuse zone of lysis centered around M(r) ~ 70 kD and contained a defined band of ~ 56 kD. Adhesion plaques contained most of the plasminogen activators (PA). 5 min after IM treatment, the M(r) ~ 150- and ~ 115-kD PA were increased in activity. Vasopressin (VP), which prevented shape change and adhesion loss when added along with IM, inhibited the increase in these PA. Preincubation with monoclonal or polyclonal antibodies to urokinase-type plaminogen activator (uPA) totally inhibited the IM-inducible shape change and ahesion loss. Activation of plasminogen throughout the gels revealed mutliple protease resistant bands that markedly increased with IM treatment (maximal at 45 min). These may represent focal control mechanisms. uPA thus may mediate focal proteolysis, which results in shape change and decreased adhesion.