Lysosomal Traffic and Degradation of ßig-h3 Protein

Abstract

Purpose:: To investigate intracellular traffic and degradation of ßig-h3 protein, a protein associated with amyloid deposition in granular corneal dystrophy Type II (GCDII).

Methods:: Corneal fibroblasts were treated with proteasomal inhibitors, lactacystin and MG132, or lysosomal inhibitors, bafilomycin A1 for 12 h. We used immunoblot analysis to measure ßig-h3 protein in cell lysate.

Results:: The lysosomal inhibitors, bafilomycin A1, significantly blocked ßig-h3 protein degradation in a dose-dependent manner in cultured corneal fibroblasts. In contrast, proteasomal inhibitor, MG132, did not lead to significant accumulation of ßig-h3 protein. The internalization of ßig-h3 is inhibited by nystatin, which blocks lipid raft endocytosis, but not by chlorpromazine which is known to block clathrin-mediated endocytosis. Moreover, ßig-h3 endocytosis was decreased significantly in TGF-ß1-exposed cultured corneal fibroblasts.

Conclusions:: These results demonstrate that ßig-h3 protein degradation is mediated by an autophagy/lysosomes pathway. Alterations both in lysosomal degradation and in expression of ßig-h3 induced by TGF-ß1 may contribute to amyloid deposition of ßig-h3 in GCDII.

Keywords: cornea: basic science • degenerations/dystrophies • cornea: stroma and keratocytes