Characterization of early autophagy signaling by quantitative phosphoproteomics

Rigbolt, Kristoffer; Zarei, Mostafa; Sprenger, Adrian; Becker, Andrea C.; Diedrich, Britta; Huang, Xun; Eiselein, Sven; Kristensen, Anders Riis; Gretzmeier, Christine; Andersen, Jens S.; Zi, Zhike; Dengjel, Jörn

doi:10.4161/auto.26864

Cited by 37 publications

(33 citation statements)

References 76 publications

(111 reference statements)

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“…NIPSNAP1 and -2 were previously identified as binding partners of LC3 and GABARAP proteins (Behrends et al, 2010;Rigbolt et al, 2014). In line with this, both endogenous NIPSNAP1 and in vitro translated NIPSNAP1 and -2 interacted with all human LC3 and GABARAP proteins when overexpressed as EGFP-tagged proteins in HeLa cells (Figure S1F) or expressed as recombinant GST-tagged proteins (Figure 1F), respectively.…”

Section: Nipsnap1 and Nipsnap2 Interact With Hatg8 Proteins Alfy Andsupporting

confidence: 76%

NIPSNAP1 and NIPSNAP2 Act as “Eat Me” Signals for Mitophagy

et al. 2019

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The clearance of damaged or dysfunctional mitochondria by selective autophagy (mitophagy) is important for cellular homeostasis and prevention of disease. Our understanding of the mitochondrial signals that trigger their recognition and targeting by mitophagy is limited. Here we show that the mitochondrial matrix proteins NIPSNAP1 (4-Nitrophenylphosphatase domain and non-neuronal SNAP25-like protein homolog 1) and NIPSNAP2 accumulate on the mitochondria surface upon mitochondrial depolarization. There they recruit proteins involved in selective autophagy, including autophagy receptors and ATG8 proteins, thereby functioning as an "eat-me signal" for mitophagy. NIPSNAP1 and NIPSNAP2 have a redundant function in mitophagy and are predominantly expressed in different tissues, with NIPSNAP1 being the most abundant in the brain. Zebrafish lacking a functional Nipsnap1 display reduced mitophagy in the brain and parkinsonian phenotypes, including loss of tyrosine hydroxylase (Th1) positive dopaminergic (DA) neurons, reduced motor activity and increased oxidative stress.

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Section: Nipsnap1 and Nipsnap2 Interact With Hatg8 Proteins Alfy Andsupporting

confidence: 76%

NIPSNAP1 and NIPSNAP2 Act as “Eat Me” Signals for Mitophagy

et al. 2019

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“…The question if specific sites are direct kinase targets or if the observed effects are of secondary nature conveyed by downstream effector kinases is not easy to address. The kinetic analysis of in vivo events may shed light onto primary and secondary events (Oliveira et al, 2015;Rigbolt et al, 2014). However, the gold standard for proving direct kinase-substrate interactions is still classical in vitro kinase assays.…”

Section: Discussionmentioning

confidence: 99%

Multilayered Control of Protein Turnover by TORC1 and Atg1

et al. 2019

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Graphical Abstract Highlights d S. cerevisiae proteins harbor a minimum of 45,000 distinct phosphorylation sites d TORC1 and Atg1 regulate at least 26 protein and lipid kinases d Atg1 phosphorylates upstream regulators of TORC1 d TORC1 phosphorylates Atg29 to inhibit autophagy SUMMARYThe target of rapamycin complex 1 (TORC1) is a master regulator of cell homeostasis, which promotes anabolic reactions and synchronously inhibits catabolic processes such as autophagy-mediated protein degradation. Its prime autophagy target is Atg13, a subunit of the Atg1 kinase complex that acts as the gatekeeper of canonical autophagy. To study whether the activities of TORC1 and Atg1 are coupled through additional, more intricate control mechanisms than simply this linear pathway, we analyzed the epistatic relationship between TORC1 and Atg1 by using quantitative phosphoproteomics. Our in vivo data, combined with targeted in vitro TORC1 and Atg1 kinase assays, not only uncover numerous TORC1 and Atg1 effectors, but also suggest distinct bi-directional regulatory feedback loops and characterize Atg29 as a commonly regulated downstream target of both TORC1 and Atg1. Thus, an exquisitely multilayered regulatory network appears to coordinate TORC1 and Atg1 activities to robustly tune autophagy in response to nutritional cues.

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“…New Phytologist kinases including cyclin-dependent protein kinases (CDKs) and mitogen-activated protein kinases (MAPKs) (Lu et al, 2002). Additionally, a phosphoproteomic study using mammalian cell line MCF7 identified the RXXS/TP motif identified in clusters 1 and 2 as a rapamycin-sensitive motif (Rigbolt et al, 2014). Other studies have also found RXRXXS/T and RXXS/T motifs (Demirkan et al, 2011;Harder et al, 2014) enriched among rapamycin-sensitive phosphosites that are recognized by mTORregulated kinases Akt, S6K1 and SGK1 (Hsu et al, 2011).…”

Section: Researchmentioning

confidence: 99%

Investigating the effect of target of rapamycin kinase inhibition on the Chlamydomonas reinhardtii phosphoproteome: from known homologs to new targets

et al. 2018

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Target of rapamycin (TOR) kinase is a conserved regulator of cell growth whose activity is modulated in response to nutrients, energy and stress. Key proteins involved in the pathway are conserved in the model photosynthetic microalga Chlamydomonas reinhardtii, but the substrates of TOR kinase and downstream signaling network have not been elucidated. Our study provides a new resource for investigating the phosphorylation networks governed by the TOR kinase pathway in Chlamydomonas. We used quantitative phosphoproteomics to investigate the effects of inhibiting Chlamydomonas TOR kinase on dynamic protein phosphorylation. Wild-type and AZD-insensitive Chlamydomonas strains were treated with TOR-specific chemical inhibitors (rapamycin, AZD8055 and Torin1), after which differentially affected phosphosites were identified. Our quantitative phosphoproteomic dataset comprised 2547 unique phosphosites from 1432 different proteins. Inhibition of TOR kinase caused significant quantitative changes in phosphorylation at 258 phosphosites, from 219 unique phosphopeptides. Our results include Chlamydomonas homologs of TOR signaling-related proteins, including a site on RPS6 with a decrease in phosphorylation. Additionally, phosphosites on proteins involved in translation and carotenoid biosynthesis were identified. Follow-up experiments guided by these phosphoproteomic findings in lycopene beta/epsilon cyclase showed that carotenoid levels are affected by TORC1 inhibition and carotenoid production is under TOR control in algae.

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Characterization of early autophagy signaling by quantitative phosphoproteomics

Cited by 37 publications

References 76 publications

NIPSNAP1 and NIPSNAP2 Act as “Eat Me” Signals for Mitophagy

NIPSNAP1 and NIPSNAP2 Act as “Eat Me” Signals for Mitophagy

Multilayered Control of Protein Turnover by TORC1 and Atg1

Investigating the effect of target of rapamycin kinase inhibition on the Chlamydomonas reinhardtii phosphoproteome: from known homologs to new targets

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