Regulation of urokinase plasminogen activator/plasmin-mediated invasion ofmelanoma cells by the integrin vitronectin receptor alpha(v)beta(3)

The integrin vitronectin receptor alpha (v)beta (3) is a mediator of cellul ar migration and invasion and has been identified as a marker of progressio n in malignant melanoma. Using a human melanoma model, we have previously s hown that this receptor was coordinately expressed with the receptor for th e urokinase plasminogen activator (uPAR). In our present study, the link be tween these receptors was further investigated by assessing the effect of a lpha (v)beta (3) ligation on uPAR transcription and function. Using the rev erse transcription-polymerase chain reaction, we found that receptor ligati on by immobilized monoclonal antibodies (MAbs) induced a rapid increase (up to 4.5 fold) in uPAR mRNA levels, which was maximal 4 hr after cell attach ment. An increase was also noted in plasminogen activator inhibitor type-1 (PAI-1) mRNA levels (2.7-fold), but none was noted in uPA levels. In additi on, ligation of alpha (v)beta (3) resulted in a significant increase in cel l surface-associated plasmin revels, which coincided with a 2- to 3-fold in crease in cell invasion as measured in the Matrigel invasion assay. This in crease in invasion could in turn be abolished by antibodies directed to UPA and uPAR and by the plasmin inhibitors epsilon -aminocaproic acid and apro tinin, Furthermore, ligation of the integrin alpha (v)beta (3) triggered a rapid increase of up to 12-fold in total cellular PKC activity, and this co incided with the redistribution of PKC beta, but not PKC alpha, from the cy tosol to the membrane. Treatment of the cells with the PKC beta -specific i nhibitor LY379196 blocked uPAR and PAI-1 mRNA induction and reduced the inc rease in cell invasion due to alpha (v)beta (3) ligation, confirming the in volvement of this isoform in the response. The results provide evidence tha t the vitronectin receptor can enhance invasion by regulating the uPAR/uPA/ plasmin system of proteolysis and implicate PKC beta as an intermediate in the activation pathway. (C) 2001 Wiley-Liss, Inc.