Mechanisms that regulate the transition of metastases from clinically undetectable and dormant to progressively growing are the least understood aspects of cancer biology. Here, we show that a large ( approximately 70%) reduction in the urokinase plasminogen activator receptor (uPAR) level in human carcinoma HEp3 cells, while not affecting their in vitro growth, induced a protracted state of tumor dormancy in vivo, with G(0)/G(1) arrest. We have now identified the mechanism responsible for the induction of dormancy. We found that uPA/uPAR proteins were physically associated with alpha5beta1, and that in cells with low uPAR the frequency of this association was significantly reduced, leading to a reduced avidity of alpha5beta1 and a lower adhesion of cells to the fibronectin (FN). Adhesion to FN resulted in a robust and persistent ERK1/2 activation and serum-independent growth stimulation of only uPAR-rich cells. Compared with uPAR-rich tumorigenic cells, the basal level of active extracellular regulated kinase (ERK) was four to sixfold reduced in uPAR-poor dormant cells and its stimulation by single chain uPA (scuPA) was weak and showed slow kinetics. The high basal level of active ERK in uPAR-rich cells could be strongly and rapidly stimulated by scuPA. Disruption of uPAR-alpha5beta1 complexes in uPAR-rich cells with antibodies or a peptide that disrupts uPAR-beta1 interactions, reduced the FN-dependent ERK1/2 activation. These results indicate that dormancy of low uPAR cells may be the consequence of insufficient uPA/uPAR/alpha5beta1 complexes, which cannot induce ERK1/2 activity above a threshold needed to sustain tumor growth in vivo. In support of this conclusion we found that treatment of uPAR-rich cells, which maintain high ERK activity in vivo, with reagents interfering with the uPAR/beta1 signal to ERK activation, mimic the in vivo dormancy induced by downregulation of uPAR.