Endocytosis of the Type III Transforming Growth Factor-β (TGF-β) Receptor through the Clathrin-independent/Lipid Raft Pathway Regulates TGF-β Signaling and Receptor Down-regulation*

Transforming growth factor-β (TGF-β) signals through three highly conserved cell surface receptors, the type III TGF-β receptor (TβRIII), the type II TGF-β receptor (TβRII), and the type I TGF-β receptor (TβRI) to regulate diverse cellular processes including cell proliferation, differentiation, migration, and apoptosis. Although TβRI and TβRII undergo ligand-independent endocytosis by both clathrin-mediated endocytosis, resulting in enhanced signaling, and clathrin-independent endocytosis, resulting in receptor degradation, the mechanism and function of TβRIII endocytosis is poorly understood. TβRIII is a heparan sulfate proteoglycan with a short cytoplasmic tail that functions as a TGF-β superfamily co-receptor, contributing to TGF-β signaling through mechanisms yet to be fully defined. We have reported previously that TβRIII endocytosis, mediated by a novel interaction with βarrestin-2, results in decreased TGF-β signaling. Here we demonstrate that TβRIII undergoes endocytosis in a ligand and glycosaminoglycan modification-independent and cytoplasmic domain-dependent manner, with the interaction of Thr-841 in the cytoplasmic domain of TβRIII with β-arrestin2 enhancing TβRIII endocytosis. TβRIII undergoes both clathrin-mediated and clathrin-independent endocytosis. Importantly, inhibition of the clathrin-independent, lipid raft pathway, but not of the clathrin-dependent pathway, results in decreased TGF-β1 induced Smad2 and p38 phosphorylation, supporting a specific role for clathrin-independent endocytosis of TβRIII in regulating both Smad-dependent and Smad-independent TGF-β signaling.