The formin DAAM1 regulates the deubiquitinase activity of USP10 and integrin homeostasis

The differentiation of fibroblasts into pathological myofibroblasts during wound healing is characterized by increased cell surface expression of & alpha;v-integrins. Our previous studies found that the deubiquitinase (DUB) USP10 removes ubiquitin from & alpha;v-integrins, leading to cell surface integrin accumulation, subsequent TGF & beta;1 activation, and pathological myofibroblast differentiation. In this study, a yeast two-hybrid screen revealed a novel binding partner for USP10, the formin, DAAM1. We found that DAAM1 binds to and inhibits USP10's DUB activity through the FH2 domain of DAAM1 independent of its actin functions. The USP10/DAAM1 interaction was also supported by proximity ligation assay (PLA) in primary human corneal fibroblasts. Treatment with TGF & beta;1 significantly increased USP10 and DAAM1 protein expression, PLA signal, and co-localization to actin stress fibers. DAAM1 siRNA knockdown significantly reduced co-precipitation of USP10 and DAAM1 on purified actin stress fibers, and & beta;1- and & beta;5-integrin ubiquitination. This resulted in increased & alpha;v-, & beta;1-, and & beta;5-integrin total protein levels, & alpha;v-integrin recycling, and extracellular fibronectin (FN) deposition. Together, our data demonstrate that DAAM1 inhibits USP10's DUB activity on integrins subsequently regulating cell surface & alpha;v-integrin localization and FN accumulation.