Abstract
Autophagy is a highly conserved catabolic process through which defective or otherwise harmful cellular components are targeted for degradation via the lysosomal route. Regulatory pathways, involving post‐translational modifications such as phosphorylation, play a critical role in controlling this tightly orchestrated process. Here, we demonstrate that TBK1 regulates autophagy by phosphorylating autophagy modifiers LC3C and GABARAP‐L2 on surface‐exposed serine residues (LC3C S93 and S96; GABARAP‐L2 S87 and S88). This phosphorylation event impedes their binding to the processing enzyme ATG4 by destabilizing the complex. Phosphorylated LC3C/GABARAP‐L2 cannot be removed from liposomes by ATG4 and are thus protected from ATG4‐mediated premature removal from nascent autophagosomes. This ensures a steady coat of lipidated LC3C/GABARAP‐L2 throughout the early steps in autophagosome formation and aids in maintaining a unidirectional flow of the autophagosome to the lysosome. Taken together, we present a new regulatory mechanism of autophagy, which influences the conjugation and de‐conjugation of LC3C and GABARAP‐L2 to autophagosomes by TBK1‐mediated phosphorylation.
Highlights
Autophagy is a highly conserved catabolic process through which defective or otherwise harmful cellular components are targeted for degradation via the lysosomal route
LC3A, LC3C, GABARAP-L1, and GABARAP-L2 are directly phosphorylated by TBK1 in vitro, and the phosphorylation sites of LC3 family proteins were identified by mass spectrometry
TBK1 is recruited to the site of autophagosome formation by autophagy receptor proteins, where TBK1 phosphorylates OPTN and p62 to promote autophagy flux [19,20,22,23,24,25,26,27,31,32]
Summary
Autophagy is a highly conserved catabolic process through which defective or otherwise harmful cellular components are targeted for degradation via the lysosomal route. We demonstrate that TBK1 regulates autophagy by phosphorylating autophagy modifiers LC3C and GABARAP-L2 on surfaceexposed serine residues (LC3C S93 and S96; GABARAP-L2 S87 and S88). This phosphorylation event impedes their binding to the processing enzyme ATG4 by destabilizing the complex. Phosphorylated LC3C/GABARAP-L2 cannot be removed from liposomes by ATG4 and are protected from ATG4-mediated premature removal from nascent autophagosomes. This ensures a steady coat of lipidated LC3C/GABARAP-L2 throughout the early steps in autophagosome formation and aids in maintaining a unidirectional flow of the autophagosome to the lysosome. We present a new regulatory mechanism of autophagy, which influences the conjugation and de-conjugation of LC3C and GABARAPL2 to autophagosomes by TBK1-mediated phosphorylation
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