Cross-talk between the p42/p44 MAP kinase and Smad pathways in transforming growth factor beta 1-induced furin gene transactivation

Furin, a predominant convertase of the cellular constitutive secretory path way, is known to be involved in the maturation of a number of growth/differ entiation factors, but the mechanisms governing its expression remain elusi ve. We have previously demonstrated that transforming growth factor (TGF) b eta1, through the activation of Smad transducers, regulates its own convert ing enzyme, furin, creating a unique activation/regulation loop of potentia l importance in a variety of cell fate and functions. Here we studied the i nvolvement of the p42/p44 MAPK pathway in such regulation. Using HepG2 cell s transfected with fur P1 LUC (luciferase) promoter construct, we observed that forced expression of a dominant negative mutant form of the small G pr otein p21(ras) (RasN17) inhibited TGF beta1-induced far gene transcription, suggesting the involvement of the p42/p44 MAPK cascade. In addition, TGF b eta induced sustained activation/phosphorylation of endogenous p42/p44 MAPK . Further-more, the role of MAPK cascade in far gene transcription was high lighted by the use of the MEK1/2 inhibitors, PD98059 or U0126, or co-expres sion of a p44 antisense construct that repressed the induction of fur promo ter transactivation. Conversely, overexpression of a constitutively active form of MEK1 increased unstimulated, TGF beta1-stimulated, and Smad2-stimul ated promoter P1 transactivation, and the universal Smad inhibitor, Smad7, inhibited this effect. Activation of Smad2 by MEK1 or TGF beta1 resulted in an enhanced nuclear localization of Smad2, which was inhibited upon blocki ng MEK1 activity. Our findings clearly show that the activation of the p42/ p44 MAPK pathway is involved in far gene expression and led us to propose a co-operative model whereby TGF beta1-induced receptor activation stimulate s not only a Smad pathway but also a parallel p42/p44 MAPK pathway that tar gets Smad2 for an increased nuclear translocation and enhanced fur gene tra nsactivation. Such an uncovered mechanism may be a key determinant for the regulation of furin in embryogenesis and growth-related physiopathological conditions.