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Exogenous H2S Induces Hrd1 S-sulfhydration and Prevents CD36 Translocation via VAMP3 Ubiquitylation in Diabetic Hearts.

ABSTRACT: Hydrogen sulfide (H2S) plays physiological roles in vascular tone regulation, cytoprotection, and ATP synthesis. HMG-CoA reductase degradation protein (Hrd1), an E3 ubiquitin ligase, is involved in protein trafficking. H2S may play a role in controlling fatty acid uptake in diabetic cardiomyopathy (DCM) in a manner correlated with modulation of Hrd1 S-sulfhydration; however, this role remains to be elucidated. The aim of the present study was to examine whether H2S can attenuate lipid accumulation and to explain the possible mechanisms involved in the regulation of the H2S-Hrd1/VAMP3 pathway. Db/db mice and neonatal rat cardiomyocytes treated with high glucose, palmitate and oleate were used as animal and cellular models of type 2 diabetes, respectively. The expression of cystathionine-?-lyase (CSE), Hrd1, CD36 and VAMP3 was detected by Western blot analysis. In addition, Hrd1 was mutated at Cys115, and Hrd1 S-sulfhydration was examined using an S-sulfhydration assay. VAMP3 ubiquitylation was investigated by immunoprecipitation. Lipid droplet formation was tested by TEM, BODIPY 493/503 staining and oil red O staining. The expression of CSE and Hrd1 was decreased in db/db mice compared to control mice, whereas CD36 and VAMP3 expression was increased. NaHS administration reduced droplet formation, and exogenous H2S restored Hrd1 expression, modified S-sulfhydration, and decreased VAMP3 expression in the plasma membrane. Using LC-MS/MS analysis, we identified 85 proteins with decreased ubiquitylation, including 3 vesicle-associated membrane proteins, in the cardiac tissues of model db/db mice compared with NaHS-treated db/db mice. Overexpression of Hrd1 mutated at Cys115 diminished VAMP3 ubiquitylation, whereas it increased CD36 and VAMP3 expression and droplet formation. siRNA-mediated Hrd1 deletion increased the expression of CD36 in the cell membrane. These findings suggested that H2S regulates VAMP3 ubiquitylation via Hrd1 S-sulfhydration at Cys115 to prevent CD36 translocation in diabetes.

SUBMITTER: Yu M

PROVIDER: S-EPMC7069459 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

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Exogenous H2S Induces Hrd1 S-sulfhydration and Prevents CD36 Translocation via VAMP3 Ubiquitylation in Diabetic Hearts.

Yu Miao M Du Haining H Wang Bingzhu B Chen Jian J Lu Fangping F Peng Shuo S Sun Yu Y Liu Ning N Sun Xiaojiao X Shiyun Dong D Zhao Yajun Y Wang Yan Y Zhao Dechao D Lu Fanghao F Zhang Weihua W

Aging and disease 20200309 2

Hydrogen sulfide (H2S) plays physiological roles in vascular tone regulation, cytoprotection, and ATP synthesis. HMG-CoA reductase degradation protein (Hrd1), an E3 ubiquitin ligase, is involved in protein trafficking. H2S may play a role in controlling fatty acid uptake in diabetic cardiomyopathy (DCM) in a manner correlated with modulation of Hrd1 S-sulfhydration; however, this role remains to be elucidated. The aim of the present study was to examine whether H2</sub ...[more]

PMID: 32257542

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Project description:Pathologic calcification (PC) is a painful and disabling condition whereby calcium-containing crystals deposit in tissues that do not physiologically calcify: cartilage, tendons, muscle, vessels and skin. In cartilage, compression and inflammation triggered by PC leads to cartilage degradation typical of osteoarthritis (OA). The PC process is poorly understood and treatments able to target the underlying mechanisms of the disease are lacking. Here we show a crucial role of the gasotransmitter hydrogen sulfide (H2S) and, in particular, of the H2S-producing enzyme cystathionine γ-lyase (CSE), in regulating PC in cartilage. Cse deficiency (Cse KO mice) exacerbated calcification in both surgically-induced (menisectomy) and spontaneous (aging) murine models of cartilage PC, and augmented PC was closely associated with cartilage degradation (OA). On the contrary, Cse overexpression (Cse tg mice) protected from these features. In vitro, Cse KO chondrocytes showed increased calcification, potentially via enhanced alkaline phosphatase (Alpl) expression and activity and increased IL-6 production. The opposite results were obtained in Cse tg chondrocytes. In cartilage samples from patients with OA, CSE expression inversely correlated with the degree of tissue calcification and disease severity. Increased cartilage degradation in murine and human tissues lacking or expressing low CSE levels may be accounted for by dysregulated catabolism. We found higher levels of matrix-degrading metalloproteases Mmp-3 and -13 in Cse KO chondrocytes, whereas the opposite results were obtained in Cse tg cells. Finally, by high-throughput screening, we identified a novel small molecule CSE positive allosteric modulator (PAM), and demonstrated that it was able to increase cellular H2S production, and decrease murine and human chondrocyte calcification and IL-6 secretion. Together, these data implicate impaired CSE-dependent H2S production by chondrocytes in the etiology of cartilage PC and worsening of secondary outcomes (OA). In this context, enhancing CSE expression and/or activity in chondrocytes could represent a potential strategy to inhibit PC.

Dataset Information

Exogenous H2S Induces Hrd1 S-sulfhydration and Prevents CD36 Translocation via VAMP3 Ubiquitylation in Diabetic Hearts.

Publications

Exogenous H<sub>2</sub>S Induces Hrd1 S-sulfhydration and Prevents CD36 Translocation via VAMP3 Ubiquitylation in Diabetic Hearts.

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