The mammalian convertase furin plays a significant role in tumorigenesis and its overexpression was observed in a number of different cancer types. To date, however, few mechanisms of action have been described. Most of the information available concerns the invasion step and designates MT1-MMP, through the activation of MMP-2, as the bona fide substrate mediating furin activity. However, recent reports indicate furin-independent pathways for MT1-MMP activation. To gain further insights into the role of furin in the invasion process, we studied the in vitro invasive capacity of LoVo cells, a furin-deficient adenocarcinoma cell line transfected with wild-type furin. Furin complementation resulted in an increased cell invasiveness that correlated with their capacity to produce MMP-2. Chemical blockage of MMPs activity with BB-3103 or MMP-2-specific antibodies revealed that the increased invasive capacity of furin-complemented cells was mediated by MMP-2. Unexpectedly, furin complementation did not change the status of MT1-MMP expression or activation, but instead resulted in the production of mature and bioactive TGFbeta1. Western blot-analysis of TGFbeta1 fragmentation species indicated that TGFbeta maturation step required furin activity, whereas results from TGFbeta-inducible reporter assays in the presence of MMP inhibitors or exogenous MMP-2 suggested that the activation step was under MMP influence. In addition, blockage with TGFbeta neutralizing antibodies revealed that furin-induced invasiveness was mediated by endogenous production of TGFbeta. Taken together, our findings established the existence of a novel alternative/complementary pathway by which furin increases tumor cell invasion through an amplification/activation loop between MMP-2 and TGFbeta.