K48/K63-linked polyubiquitination of ATG9A at K581/K838 by TRAF6 E3 ligase

ABSTRACT: Excessive generation and accumulation of highly reactive oxidizing molecules causes oxidative stress and oxidative damage to cellular components. Accumulating evidence indicates that autophagy diminishes oxidative damage in cells and maintains redox homeostasis by degrading and recycling intracellular damaged components. However, the molecular mechanisms underlying the activation of oxidative stress-mediated autophagic response remain elusive. Here we show that ubiquitin E3 ligase TNF receptor-associated factor 6 (TRAF6) and the deubiquitinase A20 coordinate to regulate ATG9A ubiquitination and autophagy activation in cells responding to oxidative stress. The reactive oxygen species (ROS)-dependent TRAF6-mediated non-proteolytic, K48/63-linked ubiquitination of ATG9A enhances its association with Beclin 1 and the assembly of class III phosphatidylinositol-3-kinase (PI3KC3/VPS34) UVRAG complex, thereby stimulating autophagy. Notably, depletion of ATG9A or expression of the ATG9A ubiquitination mutants markedly decreases ROS-induced VPS34 activation and autophagy. We further find that lipopolysaccharide (LPS)-induced ROS production also stimulates TRAF6-mediated ATG9A ubiquitination and enhances the assembly of the UVRAG-containing VPS34 complex in an ATG9A-dependent manner. Ablation of ATG9A causes aberrant TLR4 endosomal trafficking and decreased phosphorylation of IRF-3 in LPS-stimulated macrophages. Our findings provide important insight into how K48/K63-linked ubiquitination of ATG9A contributes to the regulation of oxidative stress-induced autophagy.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Early Embryonic Cell

SUBMITTER: YI-TING WANG  

LAB HEAD: Guang-Chao Chen

PROVIDER: PXD030415 | Pride | 2022-02-24

REPOSITORIES: Pride