Receptor-mediated mitophagy is a crucial process involved in mitochondria quality control. AMBRA1 is a mitophagy receptor for the selective removal of damaged mitochondria in mammalian cells. A critical unresolved issue is how AMBRA1-mediated mitophagy is controlled in response to cellular stress. Here, we investigated the role of BCL2-family proteins on AMBRA1-dependent mitophagy and showed that MCL1 delays AMBRA1-dependent mitophagy. Indeed, MCL1 overexpression is sufficient to inhibit recruitment to mitochondria of the E3 Ubiquitin ligase HUWE1, a crucial dynamic partner of AMBRA1, upon AMBRA1-mediated mitophagy induction. In addition, we found that during mitophagy induced by AMBRA1, MCL1 levels decreased but were sustained by inhibition of the GSK-3β kinase, which delayed AMBRA1-mediated mitophagy. Also, we showed that MCL1 was phosphorylated by GSK-3β at a conserved GSK-3 phosphorylation site (S159) during AMBRA1-mediated mitophagy and that this event was accompanied by HUWE1-dependent MCL1 degradation. Altogether, our results demonstrate that MCL1 stability is regulated by the kinase GSK-3β and the E3 ubiquitin ligase HUWE1 in regulating AMBRA1-mediated mitophagy. Our work thus defines MCL1 as an upstream stress-sensitive protein, functional in AMBRA1-mediated mitophagy.
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We wish to thank Dr Muriel Priault for kindly providing us the constructs BCL2L1 and MCL1; Prof. M. Campanella for the gift of Short Hairpin PARKIN and Dr V. Stagni and Prof. Barilà for the gift of MCF7 cells. We are grateful to Dr Salvatore Rizza for his help with Fig. 7. We thank M. Acuña Villa for secretarial work and F. Maria Orecchio, S. Verna, T. Maiorino, K. Bruqi for technical assistance.
This work was also supported by grants: GR2011-02351433 (Italian Ministry of Health), ROCHE (Roche per la ricerca 2017) and 5XMILLE Italian Ministry of Health (2017) to FS; AIRC (MFAG#15523) to AP; MRC, LazioInnova (85-2017-14986) and AIRC (IG#20473) to GM; grants from the Italian Ministry of Education, University and Research (Research Project of National Relevance 2017 ID 2017WC8499) and AIRC (Associazione Italiana Ricerca sul Cancro, 5 × 1000 ID 9962, AIRC IG 2018 ID 21724) to FL. P.L was supported by FUV Grant 2019. Francesco Cecconi’s laboratory is supported by grants from the Bjarne Saxhof Foundation, the Danish Cancer Society (KBVU R72-A4408, R146-A9364), the Novo Nordisk Foundation (7559, 22544), the Lundbeckfonden (R233-2016-3360), the LEO Foundation (LF17024), “Associazione Italiana per la Ricerca sul Cancro” (AIRC IG-15180). FC lab in Copenhagen are part of the Center of Excellence for Autophagy, Recycling and Disease (CARD), funded by the Danmarks Grundforskningsfond (DNRF125).
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Strappazzon, F., Di Rita, A., Peschiaroli, A. et al. HUWE1 controls MCL1 stability to unleash AMBRA1-induced mitophagy. Cell Death Differ 27, 1155–1168 (2020). https://doi.org/10.1038/s41418-019-0404-8
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