AMPK S-sulfuration contributes to H2S donors-induced AMPK phosphorylation and autophagy activation in dopaminergic cells

Abstract

Hydrogen sulfide (H₂S) serves as a neuromodulator and regulator of neuroinflammation. It is reported to be therapeutic for Parkinson's disease (PD) animal and cellular models. However, whether it affects α-synuclein accumulation in dopaminergic cells, the key pathological feature in PD, is poorly understood. In this study we reported that exogenous H₂S donors NaHS and GYY4137 (GYY) enhanced the autophagy activity, as indicated by the increases of autophagy marker LC3-II expression and LC3 dots formation even during lysosome inhibition in dopaminergic cell lines and HEK293 cells. The enhancement of H₂S donors on autophagic flux was mediated by adenosine 5'-monophosphate-activated protein kinase (AMPK)-dependent mammalian target of rapamycin (mTOR) inhibition, as H₂S donors activated AMPK but reduced the mTOR activity and H₂S donors-induced LC3-II increase was diminished by mTOR activator. Moreover, point mutation of Cys302 into alanine (C302A) in AMPKα2 subunit abolished the AMPK activation and mTOR inhibition, as well as autophagic flux increase elicited by NaHS. Interestingly, NaHS triggered AMPK S-sulfuration, which was not observed in AMPK C302A-transfected cells. Further, NaHS was able to attenuate α-synuclein accumulation in a cellular model induced by dopamine oxidized metabolite 3, 4-dihydroxyphenylacetaldehyde (DOPAL), and this effect was interfered by autophagy inhibitor wortmannin and also eliminated in AMPK Cys302A-transfected cells. In sum, the findings identified a role of Cys302 S-sulfuration in AMPK activation induced by exogenous H₂S and demonstrated that H₂S donors could enhance the autophagic flux via AMPK-mTOR signaling and thus reduce α-synuclein accumulation in vitro.

Keywords: AMPK; Autophagy; Hydrogen sulfide; S-sulfuration; α-synuclein.