title: Autophagy is induced by raptor degradation via the ubiquitin/proteasome system in granular corneal dystrophy type 2.
abstract: Granular corneal dystrophy type 2 (GCD2) is an autosomal dominant disorder that is caused by a point mutation in transforming growth factor-β-induced gene-h3 (TGFBI), which encodes transforming growth factor-β-induced protein (TGFBIp). Recently, we found that the autophagic clearance of mutant-TGFBIp is delayed in GCD2 corneal fibroblasts; however, any potential correlation between mutant-TGFBIp turnover and autophagy-lysosome pathway remains unknown. Here, we report that mutant-TGFBIp is accumulated and that autophagy, a key clearance pathway for mutant-TGFBIp, is induced in primary cultured GCD2 homozygous (HO) and wild-type (WT) corneal fibroblasts that express exogenously introduced mutant-TGFBIp. Mutant-TGFBI colocalized with LC3-enriched cytosolic vesicles and cathepsin D in primary cultured GCD2 corneal fibroblasts. We also observed reduced levels of raptor (regulatory-associated protein of the mammalian target of rapamycin [mTOR]) in GCD2 corneal fibroblasts and WT corneal fibroblasts expressing mutant-TGFBIp. Strikingly, treatment with MG132, a ubiquitin/proteasome system inhibitor, significantly increased the levels of both total and ubiquitinated raptor in GCD2 corneal fibroblasts. The levels of the autophagy marker LC3-II were also increased in WT corneal fibroblasts that were treated with shRNA against raptor. However, mutant-TGFBIp accumulated in autophagosomes or/and lysosomes in spite of the significant activation of basal autophagy in GCD2 corneal fibroblasts. These results suggest that an insufficient autophagy-lysosome pathway might be responsible for the intracellular accumulation of mutant-TGFBIp during the pathogenesis of GCD2.
authors.affiliation: Cornea Dystrophy Research Institute and Department of Ophthalmology, Yonsei University College of Medicine, Seoul, Republic of Korea.
authors.forename: Seung-Il
authors.initials: SI
authors.lastname: Choi
authors.affiliation: Cornea Dystrophy Research Institute and Department of Ophthalmology, Yonsei University College of Medicine, Seoul, Republic of Korea.
authors.forename: Yong-Sun
authors.initials: YS
authors.lastname: Maeng
authors.affiliation: School of Medicine, Emory University, Atlanta, GA, United States.
authors.forename: Kyu Seo
authors.initials: KS
authors.lastname: Kim
authors.affiliation: Cornea Dystrophy Research Institute and Department of Ophthalmology, Yonsei University College of Medicine, Seoul, Republic of Korea.
authors.forename: Tae-Im
authors.initials: TI
authors.lastname: Kim
authors.affiliation: Cornea Dystrophy Research Institute and Department of Ophthalmology, Yonsei University College of Medicine, Seoul, Republic of Korea; BK21 Project for Medical Science and Severance Biomedical Science Institute, Yonsei University College of Medicine, Republic of Korea. Electronic address: eungkkim@yuhs.ac.
authors.forename: Eung Kweon
authors.initials: EK
authors.lastname: Kim
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D001343:Autophagy; D003317:Corneal Dystrophies, Hereditary / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D059748:Proteolysis; D000076223:Regulatory-Associated Protein of mTOR; D025801:Ubiquitin / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C463659:RPTOR protein, human; D000076223:Regulatory-Associated Protein of mTOR; D025801:Ubiquitin; D046988:Proteasome Endopeptidase Complex
keywords: Autophagy; Granular corneal dystrophy type 2; Raptor; TGFBIp; Ubiquitin/proteasome system; mTOR