Autophagy impairment mediated by S-nitrosation of ATG4B leads to neurotoxicity in response to hyperglycemia

The majority of diabetic patients develop neuropathy and there is an increasing prevalence of neurodegeneration in the central nervous system (CNS). However, the mechanism behind this is poorly understood. Here we first observed that macroautophagy/autophagy was suppressed in the hippocampus of diabetic GK rats with hyperglycemia, whereas it was unchanged in ob/ob mice without hyperglycemia. Autophagy could be directly inhibited by high glucose in mouse primary hippocampal neurons. Moreover, autophagy was protective in high-glucose-induced neurotoxicity. Further studies revealed that autophagic flux was suppressed by high glucose due to impaired autophagosome synthesis illustrated by mRFP-GFP-LC3 puncta analysis. We showed that decreased autophagy was dependent on NO produced under high glucose conditions. Therefore, (LC-MS/MS)-based quantitative proteomic analysis of protein S-nitrosation was performed and a core autophagy protein, ATG4B was found to be S-nitrosated in the hippocampus of GK rats. ATG4B was also verified to be S-nitrosated in neuronal cells cultured with high glucose. The activities of ATG4B in the processing of unmodified, precursor Atg8-family proteins and in the deconjugation of PE from lipidated Atg8-family proteins, which are essential for efficient autophagosome biogenesis were both compromised by S-nitrosation at Cys189 and Cys292 sites. In addition, ATG4B processing of the GABARAPL1 precursor was affected the least by S-nitrosation compared with other substrates. Finally, ATG4B S-nitrosation was verified to be responsible for decreased autophagy and neurotoxicity in response to high glucose. In conclusion, autophagy impairment mediated by S-nitrosation of ATG4B leads to neurotoxicity in response to hyperglycemia. Our research reveals a novel mechanism linking hyperglycemia with CNS neurotoxicity and shows that S-nitrosation is a novel post-transcriptional modification of the core autophagy machinery.