Hrd1 participates in the regulation of collagen I synthesis in renal fibrosis.

The production and accumulation of collagen-rich extracellular matrix are common hallmarks during the process of renal fibrogenesis. However, the mechanisms of the regulation of collagen synthesis in renal fibrosis are still unclear. Hrd1, an E3 ubiquitin ligase, plays important roles for protein folding in ER and transport to Golgi. Here, we examined the hypothesis that Hrd1 posttranslationally regulates collagen synthesis in renal interstitial fibrogenesis. Unilateral ureteral obstruction induced Hrd1 expression, predominantly in the renal interstitium and tubular epithelium of fibrotic kidneys. Transforming growth factor β1, as a key mediator in kidney fibrosis, significantly increased the expressions of Hrd1, α-smooth muscle actin, fibronectin as well as procollagen I and mature collagen I in dose-dependent manner in tubular epithelial cells, suggesting that collagen I maturation might be modulated during renal fibrosis. In cultured renal fibroblasts, Hrd1 knockdown decreased secreted collagen I ~60% in the supernatant of NRK-49F cells. Conversely, Hrd1 overexpression increased secreted collagen I ~1.5-fold. Hrd1 overexpression significantly increased the expressions of both procollagen I and mature collagen I, ~2.2-fold and ~1.8-fold, respectively. However, Hrd1 knockdown markedly decreased the expression of mature collagen I ~80%, while procollagen I expression only was decreased ~21%. Moreover, short interfering RNA-induced knockdown of Sec23A blunted the increase in collagen I expression (both immature and mature form) by Hrd1 overexpression and returned collagen I expression toward control levels. These results indicate that Hrd1 plays an important role in the maturation of collagen I in renal fibrosis, and that Sec23A pathway is required for ER-to-Golgi procollagen trafficking to promote collagen synthesis.