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. 2018 Sep 7;42(6):3278–3290. doi: 10.3892/ijmm.2018.3866

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Copyright: © Huang et al.

This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.

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O-GlcNAc modification of SNAP29 inhibits SNAP29-STX17-VAMP8 complex formation and inhibits autophagy-mediated degradation. NRCMs were exposed to high glucose (25 mM) and were treated with TG (5 µM) or Don (40 µM) for 24 h. The combination of O-GlcNAc-modified SNAP29 into the SNAP29-STX17-VAMP8 complex was observed by co-IP. (A) SNAP29 antibody was used as pull down the VAMP8 and STX17, SNAP29 and Tubulin were used as the loading control; (B) VAMP8 antibody was applied to pull down the complex SNAP29 and STX17, VAMP8 and Tubulin were used as the loading control. (C) STX17 antibody was used to pull down the complex SANP29 and VAMP8, STX17 and Tubulin acted as the loading control. (^**P<0.01, ^***P<0.001 vs. Vehicle+Glu group; ^##P<0.01, ^###P<0.001 vs. TG+Glu group); NRCMs were transfected with sh-OGT or ad-OGA for 48 h and to high glucose for another 24 h, following which the formation of the SNAP29-STX17-VAMP8 complex was observed by co-IP. (D) SNAP29 antibody was used as pull down the VAMP8 and STX17, SNAP29 and Tubulin were used as the loading control; (E) VAMP8 antibody was applied to pull down the complex SNAP29 and STX17, VAMP8 and Tubulin were used as the loading control. (F) STX17 antibody was used to pull down the complex SANP29 and VAMP8, STX17 and Tubulin acted as the loading control. (^*P<0.05, ^**P<0.01, ^***P<0.001 vs. sh-Ctrl+Glu group; ^##P<0.01, ^###P<0.001 vs. sh-OGT+Glu group); NRCMs, neonatal rat cardiomyocytes; O-GlcNAc, O-linked β-N-acetylglucosamine; SNAP29, synaptosomal-associated protein 29; vesicle-associated membrane protein 8; STX17, syntaxin-17; Don, 6-diazo-5-oxo-L-norleucine; TG, thiamet G; OGT, O-GlcNAc transferase; sh-OGT, OGT-knockdown adenovirus; ad-OGA, OGA-overexpression adeno-virus; Ctrl, control; co-IP, coimmunoprecipitation.

O-GlcNAc modification of SNAP29 inhibits SNAP29-STX17-VAMP8 complex formation and inhibits autophagy-mediated degradation. NRCMs were exposed to high glucose (25 mM) and were treated with TG (5 µM) or Don (40 µM) for 24 h. The combination of O-GlcNAc-modified SNAP29 into the SNAP29-STX17-VAMP8 complex was observed by co-IP. (A) SNAP29 antibody was used as pull down the VAMP8 and STX17, SNAP29 and Tubulin were used as the loading control; (B) VAMP8 antibody was applied to pull down the complex SNAP29 and STX17, VAMP8 and Tubulin were used as the loading control. (C) STX17 antibody was used to pull down the complex SANP29 and VAMP8, STX17 and Tubulin acted as the loading control. (^**P<0.01, ^***P<0.001 vs. Vehicle+Glu group; ^##P<0.01, ^###P<0.001 vs. TG+Glu group); NRCMs were transfected with sh-OGT or ad-OGA for 48 h and to high glucose for another 24 h, following which the formation of the SNAP29-STX17-VAMP8 complex was observed by co-IP. (D) SNAP29 antibody was used as pull down the VAMP8 and STX17, SNAP29 and Tubulin were used as the loading control; (E) VAMP8 antibody was applied to pull down the complex SNAP29 and STX17, VAMP8 and Tubulin were used as the loading control. (F) STX17 antibody was used to pull down the complex SANP29 and VAMP8, STX17 and Tubulin acted as the loading control. (^*P<0.05, ^**P<0.01, ^***P<0.001 vs. sh-Ctrl+Glu group; ^##P<0.01, ^###P<0.001 vs. sh-OGT+Glu group); NRCMs, neonatal rat cardiomyocytes; O-GlcNAc, O-linked β-N-acetylglucosamine; SNAP29, synaptosomal-associated protein 29; vesicle-associated membrane protein 8; STX17, syntaxin-17; Don, 6-diazo-5-oxo-L-norleucine; TG, thiamet G; OGT, O-GlcNAc transferase; sh-OGT, OGT-knockdown adenovirus; ad-OGA, OGA-overexpression adeno-virus; Ctrl, control; co-IP, coimmunoprecipitation.