AMBRA1 Interplay with Cullin E3 Ubiquitin Ligases Regulates Autophagy Dynamics

• CUL4 controls autophagy by regulating AMBRA1 protein levels
• Autophagy stimuli induce CUL4-AMBRA1 dissociation and AMBRA1 stabilization via ULK1
• AMBRA1 inhibits CUL5 to stabilize DEPTOR and accelerate mTOR inactivation
• CUL4-AMBRA1 reassociation triggers AMBRA1 degradation and autophagy termination

SummaryAutophagy maintains cellular homeostasis by degrading harmful or unnecessary intracellular components. How the autophagy response is induced rapidly and transiently remains largely unknown. We report that the E3 ubiquitin ligases Cullin-5 and Cullin-4 regulate the onset and termination of autophagy, respectively, by dynamically interacting with AMBRA1, a regulator of autophagy. Under normal conditions, Cullin-4 binding to AMBRA1 limits its protein abundance. Autophagy stimuli promote AMBRA1 stabilization by causing ULK1-dependent Cullin-4 release. Notably, Cullin-4/AMBRA1 dissociation is transient, and the re-established interaction triggers AMBRA1 degradation, terminating the autophagy response. Moreover, Cullin-4 inhibits the interaction between AMBRA1 and another Cullin E3 ligase. Indeed, upon Cullin-4 dissociation, AMBRA1 binds and inhibits Cullin-5, thus promoting the accumulation of the mTOR inhibitor DEPTOR. Through DEPTOR stabilization, AMBRA1 establishes a feedback loop that ensures the rapid onset of autophagy by enhancing mTOR inactivation. Our findings show that Cullin-mediated degradation of autophagy regulators temporally controls the autophagy response.

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