mTOR inhibits autophagy by controlling ULK1 ubiquitylation, self-association and function through AMBRA1 and TRAF6

Nazio, Francesca; Strappazzon, Flavie; Antonioli, Manuela; Bielli, Pamela; Cianfanelli, Valentina; Bordi, Matteo; Gretzmeier, Christine; Dengjel, Jörn; Piacentini, Mauro; Fimia, Gian María; Cecconi, Francesco

doi:10.1038/ncb2708

Cited by 687 publications

(635 citation statements)

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“…ULK1 is a relatively stable protein and is subject to proteasome-mediated degradation. Post-translational modifications including K63-linked ubiquitylation 16,17 and phosphorylation [18][19][20] have been reported to modulate the rates of ULK1 turnover and kinase activity in different cellular contexts. Hsp90 and Cdc37 have been shown to regulate ULK1 stability and activity by forming complex with ULK1, which subsequently influences Atg13-mediated mitophagy.…”

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confidence: 99%

“…ULK1 has been suggested to undergo proteasome-mediated degradation. 17,21 We, therefore, tested whether ULK1 reduction in p32-depleted cells could be attributed to increased proteolysis of ULK1. Indeed, proteasomal inhibitor MG132 treatment completely blocked downregulation of ULK1 in p32-ablated cells (Figure 2e), demonstrating that depletion of p32 leads to increased proteasomal degradation of ULK1.…”

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confidence: 99%

“…Previous studies reported that ULK1 is a K63-linked but not K48-linked ubiquitylated protein. 16,17 We investigated whether p32 regulates ULK1 stability via modulating its ubiquitylation. Unlike previous studies, when we used ubiquitin mutants Ub-K48 and Ub-K63, which contain arginine substitutions on all their lysine residues except the one at position 48 and 63, respectively, we found ULK1 is subject to both K48-linked and K63-linked ubiquitylation in vivo (Figure 3a).…”

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confidence: 99%

“…p32 regulates ULK1 stability and autophagy H Jiao et al TRAF6 is the only known E3 ligase that mediates ULK1 K63 ubiquitylation and its subsequent stabilization. 17 We, therefore, tested whether there is an interplay between TRAF6 and p32 in regulating the steady state of ULK1. As both TRAF6 and p32 could bind to ULK1, we wondered whether these two molecules may form protein complex.…”

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Chaperone-like protein p32 regulates ULK1 stability and autophagy

Jiao¹,

Gq²,

Dong³

et al. 2015

Cell Death Differ

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Mitophagy mediates clearance of dysfunctional mitochondria, and represents one type of mitochondrial quality control, which is essential for optimal mitochondrial bioenergetics. p32, a chaperone-like protein, is crucial for maintaining mitochondrial membrane potential and oxidative phosphorylation. However, the relationship between p32 and mitochondrial homeostasis has not been addressed. Here, we identified p32 as a key regulator of ULK1 stability by forming complex with ULK1. p32 depletion potentiated K48-linked but impaired K63-linked polyubiquitination of ULK1, leading to proteasome-mediated degradation of ULK1. As a result, silencing p32 profoundly impaired starvation-induced autophagic flux and the clearance of damaged mitochondria caused by mitochondrial uncoupler. Importantly, restoring ULK1 expression in p32-depleted cells rescued autophagy and mitophagy defects. Our findings highlight a cytoprotective role of p32 under starvation conditions by regulating ULK1 stability, and uncover a crucial role of the p32-ULK1-autophagy axis in coordinating stress response, cell survival and mitochondrial homeostasis. Mitophagy is a selective form of autophagy by which mitochondria are degraded in autolysosomes. p32 is a critical regulator of mitochondrial bioenergetics. 1 It primarily localizes to the mitochondrial matrix, but has also been reported to be present in other subcellular locations. 2-5 Many human tumors exhibit higher p32 expression levels than their nonmalignant counterpart tissues. 6-9 Depleting p32 in human cancer cells strongly shifts their metabolism from oxidative phosphorylation to glycolysis. 1 Consistently, p32 knockout causes mid-gestation lethality of knockout embryos and defects in oxidative phosphorylation. Mouse embryonic fibroblasts (MEFs) generated from p32 knockout embryos exhibited impaired ATP production and reduced mitochondrial membrane potential, which is in agreement with the observation that p32 silencing leads to increased mitochondrial fragmentation. 10,11 Notably, p32 was found to form protein complex with a variety of molecules 7,12,13 and has been suggested that it may act as a multifunctional chaperone protein. [12][13][14] ULK1 has a crucial role in mitophagy induction. 15 Despite the pivotal role of ULK1 in mitochondrial clearance, little is known as how ULK1 itself is regulated. ULK1 is a relatively stable protein and is subject to proteasome-mediated degradation. Post-translational modifications including K63-linked ubiquitylation 16,17 and phosphorylation [18][19][20] have been reported to modulate the rates of ULK1 turnover and kinase activity in different cellular contexts. Hsp90 and Cdc37 have been shown to regulate ULK1 stability and activity by forming complex with ULK1, which subsequently influences Atg13-mediated mitophagy. 21 Here, we found p32 regulates ULK1 stability by forming protein complex with ULK1. The interaction between ULK1 and p32 is crucial for maintaining the steady-state levels and activity of ULK1. We further show that p32 ablation results in a ...

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confidence: 99%

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confidence: 99%

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confidence: 99%

mentioning

confidence: 99%

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Chaperone-like protein p32 regulates ULK1 stability and autophagy

Jiao¹,

Gq²,

Dong³

et al. 2015

Cell Death Differ

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“…Altogether, these findings suggest that the role of WASH in autophagy is dependent on its subcellular localization and its partners in intracellular membranes. some formation via its E3 ligase activity with DDB1-CUL4A to enhance Beclin 1 association with VPS34 [7] and with TRAF6 to stabilize ULK1, which acts in a complex upstream of the PI3K complex 1 in autophagy [9]. Finally, the study by Xia et al emphasizes that autophagy must be tightly regulated to avoid deleterious effects on cell homeostasis.…”

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Autophagy regulation: RNF2 targets AMBRA1

2014

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npgThe phosphatidylinositol 3-kinase complex I (PI3K complex I) is a crucial regulator of autophagy, which contains Beclin 1 (or ATG6), ATG14L, VPS34 (or the class III phosphatidylinositol 3-kinase and its adaptor VPS15) and AMBRA1, and controls autophagosome formation. In a paper recently published in Cell Research, Xia et al. report that during nutrient deprivation the ubiquitin E3 ligase RNF2 is recruited to the PI3K complex I, and ubiquitinates AMBRA1 to trigger its degradation and downregulate autophagy.Macroautophagy (hereafter called autophagy) is a lysosomal degradation pathway for cytoplasmic components [1]. The ubiquitin-proteasome pathway is also critical during the process of autophagy. The formation of autophagosomes (double-membrane bound vacuoles that sequester cargo in bulk or in a selective manner before their delivery to the lysosomal compartment) depends on ubiquitination-like activity [2]. The selective removal of cargo (e.g., protein aggregates, organelles) by autophagy is dependent in many cases on the ubiquitination of the cargo [3]. Recent studies also indicate that ubiquitination regulates the activity of autophagy proteins that comprise autophagosomes [4].Autophagosome formation is dependent on evolutionarily conserved ATG (autophagy-related) proteins initially identified in yeast [2]. These proteins function in complexes or functional modules on a membrane known as the phagophore that matures into the autophagosome via stages of initiation, elongation, and sealing. Phosphati-

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A historical perspective of macroautophagy regulation by biochemical and biomechanical stimuli

Dupont,

Claude‐Taupin,

Codogno

2023

FEBS Letters

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Macroautophagy is a lysosomal degradative pathway for intracellular macromolecules, protein aggregates and organelles. The formation of the autophagosome, a double membrane‐bound structure that sequesters cargoes before their delivery to the lysosome, is regulated by several stimuli in multicellular organisms. Pioneering studies in rat liver showed the importance of amino acids, insulin and glucagon in controlling macroautophagy. Thereafter, many studies have deciphered the signaling pathways downstream of these biochemical stimuli to control autophagosome formation. Two signaling hubs have emerged: the kinase mTOR, in a complex at the surface of lysosomes which is sensitive to nutrients and hormones; and AMPK, which is sensitive to the cellular energetic status. Besides nutritional, hormonal and energetic fluctuations, many organs have to respond to mechanical forces (compression, stretching and shear stress). Recent studies have shown the importance of mechanotransduction in controlling macroautophagy. This regulation engages cell surface sensors, such as the primary cilium, in order to translate mechanical stimuli into biological responses.

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mTOR inhibits autophagy by controlling ULK1 ubiquitylation, self-association and function through AMBRA1 and TRAF6

Cited by 687 publications

References 33 publications

Chaperone-like protein p32 regulates ULK1 stability and autophagy

Chaperone-like protein p32 regulates ULK1 stability and autophagy

Autophagy regulation: RNF2 targets AMBRA1

A historical perspective of macroautophagy regulation by biochemical and biomechanical stimuli

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