Smad proteins differentially regulate transforming growth factor-β-mediated induction of chondroitin sulfate proteoglycans

Traumatic injury to the CNS results in increased expression and deposition of chondroitin sulfate proteoglycans (CSPGs) that are inhibitory to axonal regeneration. Transforming growth factor-beta (TGF-beta) has been implicated as a major mediator of these changes, but the mechanisms through which TGF-beta regulates CSPG expression are not known. Using lentiviral expressed Smad-specific ShRNA we show that TGF-beta induction of CSPG expression in astrocytes is Smad-dependent. However, we find a differential dependence of the synthetic machinery on Smad2 and/or Smad3. TGF-beta induction of neurocan and xylosyl transferase 1 required both Smad2 and Smad3, whereas induction of phosphacan and chondroitin synthase 1 required Smad2 but not Smad3. Smad3 knockdown selectively reduced induction of chondroitin-4-sulfotransferase 1 and the amount of 4-sulfated CSPGs secreted by astrocytes. Additionally, Smad3 knockdown in astrocytes was more efficacious in promoting neurite outgrowth of neurons cultured on the TGF-beta-treated astrocytes. Our data implicate TGF-beta Smad3-mediated induction of 4-sulfation as a critical determinant of the permissiveness of astrocyte secreted CSPGs for axonal growth.