Differential effects of echistatin and thrombin on collagen production and prolidase activity in human dermal fibroblasts and their possible implication in β1-integrin-mediated signaling

Prolidase [E.C. 3.4.13.9] is a cytosolic imidodipeptidase that plays an important role in collagen biosynthesis. The enzyme contributes to the recovery of proline from protein degradation products (mainly collagen) for collagen resynthesis. Prolidase activity and collagen biosynthesis are supposed to be regulated by β1-integrins, which initiate a signaling pathway in which several kinases and intracellular proteins are involved, including focal adhesion kinase pp125FAK (FAK), Src, Shc, growth factor receptor bound protein 2 (Grb-2), son of sevenless protein (SOS), Ras, Raf and mitogen-activated protein kinases (MAPK), extracellular-signal regulated kinase 1 (ERK1) and kinase 2 (ERK2). We studied the effects of echistatin, a well-known disintegrin and thrombin, a serine protease capable of activation of platelet integrin α2β1 receptor on collagen production, prolidase activity, expression of prolidase, β1-integrin receptor, FAK, SOS-protein and phosphorylated MAP-kinases (ERK1 and ERK2) in confluent human dermal fibroblasts. It has been found that treatment of the cells with 100 nM echistatin contributes to inhibition of collagen production, as well as prolidase activity and expression compared to control cells. These phenomena were accompanied by a decrease in the expression of FAK, SOS-protein and phosphorylated MAP-kinases, ERK1 and ERK2. An opposite phenomenon was observed in fibroblasts treated with 0.1 IU thrombin. In this case, a significant increase in collagen production and prolidase activity, accompanied by a distinct raise in the expression of prolidase, FAK and phosphorylated MAP-kinases and a slight increase in expression of SOS compared to controls were found. The results suggest that regulation of prolidase activity and collagen biosynthesis in human dermal fibroblasts may involve β1-integrin-dependent signaling.