Atg38 is required for autophagy-specific phosphatidylinositol 3-kinase complex integrity

Araki, Yasuhiro; Ku, Wei-Chi; Akioka, Manami; May, Alexander I.; Hayashi, Yu; Arisaka, Fumio; Ishihama, Yasushi; Ohsumi, Yoshinori

doi:10.1083/jcb.201304123

Cited by 104 publications

(142 citation statements)

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“…We previously reported the V-shaped architecture of the four-subunit version of PI3KC3-C1 consisting of VPS (vacuolar protein sorting) 34, VPS15, BECN1 (Beclin 1), and ATG (autophagy-related) 14. Here we show that a putative fifth subunit, nuclear receptor binding factor 2 (NRBF2), is a tightly bound component of the complex that profoundly affects its activity and architecture.…”

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confidence: 92%

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Dynamics and architecture of the NRBF2-containing phosphatidylinositol 3-kinase complex I of autophagy

Young

Cho

Lawrence

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The class III phosphatidylinositol 3-kinase complex I (PI3KC3-C1) is central to autophagy initiation. We previously reported the V-shaped architecture of the four-subunit version of PI3KC3-C1 consisting of VPS (vacuolar protein sorting) 34, VPS15, BECN1 (Beclin 1), and ATG (autophagy-related) 14. Here we show that a putative fifth subunit, nuclear receptor binding factor 2 (NRBF2), is a tightly bound component of the complex that profoundly affects its activity and architecture. NRBF2 enhances the lipid kinase activity of the catalytic subunit, VPS34, by roughly 10-fold. We used hydrogen-deuterium exchange coupled to mass spectrometry and negative-stain electron microscopy to map NRBF2 to the base of the V-shaped complex. NRBF2 interacts primarily with the N termini of ATG14 and BECN1. We show that NRBF2 is a homodimer and drives the dimerization of the larger PI3KC3-C1 complex, with implications for the higherorder organization of the preautophagosomal structure.hydrogen-deuterium exchange | electron microscopy | allostery A utophagy is a pathway of subcellular engulfment and lysosomal transport that is conserved throughout eukaryotes (1). Survival during starvation is probably the primordial function of autophagy (2). Most current research focuses on a growing array of selective autophagy pathways in human cells (3). These include autophagy of mitochondria (mitophagy), endoplasmic reticulum (ER-phagy), and intracellular microbes (xenophagy). Autophagy is generally considered protective against a range of diseases, including several neurodegenerative diseases (4), microbial infections, and cancer (5). Autophagy can also promote tumor growth in late stages of cancer, and therefore agonists and antagonists of autophagy are of interest as potential therapeutics (5). The centrality of autophagy to both basic cell processes and human health has leant urgency to understanding its underlying molecular mechanisms.Bulk and selective autophagy share common machinery for the initiation and growth of the double-membrane sheet known as the phagophore. The phagophore matures and closes to form the autophagosome. The proteins dedicated to autophagy number ∼41 in yeast and more in humans. They include the ULK1 (unc-51 like autophagy activating kinase 1) protein kinase complex [Atg1 (autophagy-related 1) complex in yeast], the phosphatidylinositol 3-kinase complexes I and II (PI3KC3-C1 and -C2), the integral membrane protein ATG9, the WIPIs as PI(3P) receptors, and the ubiquitin-like LC3 protein family and their conjugation pathway. PI3KC3-C1 functions in autophagy initiation (6, 7), whereas PI3KC3-C2 is important for autolysosome formation at a later step in autophagy (8). The two PI3KC3 complexes share three subunits in common: the lipid kinase subunit VPS34 (vacuolar protein sorting 34) and the regulatory subunits BECN1 (Beclin 1) and VPS15. PI3KC3-C2 contains the unique subunit UVRAG (UV radiation resistance associated), which is involved in regulating the fusion of autophagosomes to lysosomes (9). PI3KC3-C1 is targeted to ...

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confidence: 92%

“…Recently a putative fifth subunit of the PI3KC3-C1 complex was identified in yeast and named Atg38 (14). Atg38 consists of an N-terminal MIT (microtubule interaction and transport) domain and a C-terminal coiled coil, together with connecting linkers.…”

mentioning

confidence: 99%

Dynamics and architecture of the NRBF2-containing phosphatidylinositol 3-kinase complex I of autophagy

Young

Cho

Lawrence

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…In yeast, Vps15 (a putative regulatory protein kinase that is required for Vps34 membrane association), Vps30/Atg6, Vps34 (the PtdIns3K), Atg14 and Atg38, form the autophagy-specific class III PtdIns3K complex I (Figure 4), which localizes at the PAS [92][93][94]. One key role of the PtdIns3K complex, the generation of PtdIns-3P, is presumably to recruit PtdIns3P-binding proteins such as Atg18 to the PAS.…”

Section: Yeast Ptdins3k Complexes I and Iimentioning

confidence: 99%

The machinery of macroautophagy

et al. 2013

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Autophagy is a primarily degradative pathway that takes place in all eukaryotic cells. It is used for recycling cytoplasm to generate macromolecular building blocks and energy under stress conditions, to remove superfluous and damaged organelles to adapt to changing nutrient conditions and to maintain cellular homeostasis. In addition, autophagy plays a critical role in cytoprotection by preventing the accumulation of toxic proteins and through its action in various aspects of immunity including the elimination of invasive microbes and its participation in antigen presentation. The most prevalent form of autophagy is macroautophagy, and during this process, the cell forms a double-membrane sequestering compartment termed the phagophore, which matures into an autophagosome. Following delivery to the vacuole or lysosome, the cargo is degraded and the resulting macromolecules are released back into the cytosol for reuse. The past two decades have resulted in a tremendous increase with regard to the molecular studies of autophagy being carried out in yeast and other eukaryotes. Part of the surge in interest in this topic is due to the connection of autophagy with a wide range of human pathophysiologies including cancer, myopathies, diabetes and neurodegenerative disease. However, there are still many aspects of autophagy that remain unclear, including the process of phagophore formation, the regulatory mechanisms that control its induction and the function of most of the autophagy-related proteins. In this review, we focus on macroautophagy, briefly describing the discovery of this process in mammalian cells, discussing the current views concerning the donor membrane that forms the phagophore, and characterizing the autophagy machinery including the available structural information.

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“…[81][82][83]159 The yeast ortholog of NRBF2 is presumed to be Atg38, although there are sequence diversities. 82,160 Likewise, Atg38 is recognized as a component of the Atg14-containing Vps34 complex I in yeast, which is required for the autophagy pathway. 160 Intriguingly, in yeast Atg38 is responsible for the stability of Vps34 complex I, which connects Vps34-Vps15 with Atg14-Vps30, 160 reminiscent of the function of NRBF2 in mediating ATG14, PIK3C3-PIK3R4, and BECN1 interaction in mammalian cells.…”

Section: Regulation Of Autophagy By Pik3c3 Via Other Interacting Partmentioning

confidence: 99%

“…82,160 Likewise, Atg38 is recognized as a component of the Atg14-containing Vps34 complex I in yeast, which is required for the autophagy pathway. 160 Intriguingly, in yeast Atg38 is responsible for the stability of Vps34 complex I, which connects Vps34-Vps15 with Atg14-Vps30, 160 reminiscent of the function of NRBF2 in mediating ATG14, PIK3C3-PIK3R4, and BECN1 interaction in mammalian cells. 82 However, it is a question under debate whether NRBF2 positively 81,83 or negatively 82 regulates autophagy in higher eukaryotes.…”

Section: Regulation Of Autophagy By Pik3c3 Via Other Interacting Partmentioning

confidence: 99%

Differential regulatory functions of three classes of phosphatidylinositol and phosphoinositide 3-kinases in autophagy

2015

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Abbreviations: 3-MA, 3-methyladenine; AMBRA1, autophagy/Beclin 1 regulator 1; ATG, autophagy related; ATM, ATM serine/threonine kinase; ATR, ATR serine/threonine kinase; BH3, Bcl-2 homology 3; CCD, coiled-coil domain; CDK, cyclin-dependent kinase; CISD2, CDGSH iron sulfur domain 2; class I/II PI3K, class I/II phosphoinositide 3-kinase; class III PtdIns3K, class III phosphatidylinositol 3-kinase; DAPK1, death-associated protein kinase 1; DDR, DNA damage response; EIF4EBP1, eukaryotic translation initiation factor 4E binding protein 1; ER, endoplasmic reticulum; FOXO, forkhead box O; FYCO1, FYVE and coiledcoil domain containing 1; GAP, GTPase-activating protein; HMGB1, high mobility group box 1; HSPA1A, heat shock 70kDa protein 1A; IR, ionizing radiation; MAPK8, mitogen-activated protein kinase 8; MEFs, mouse embryonic fibroblasts; MTMR, myotubularin-related protein; MTOR, mechanistic target of rapamycin (serine/threonine kinase); MTORC1, mechanistic target of rapamycin complex 1; MVBs, spherical multivesicular bodies; NRBF2, nuclear receptor binding factor 2; PAS, phagophore assembly site; PDPK1, 3-phosphoinositide dependent protein kinase 1; PE, phosphatidylethanolamine; PIKFYVE, phosphoinositide kinase, FYVE finger containing; PINK1, PTEN-induced putative kinase 1; PtdIns3K-C1, class III phosphatidylinositol 3-kinase complex I; PtdIns3K-C2, class III phosphatidylinositol 3-kinase complex II; PKB, protein kinase B; PRKA, protein kinase, AMP-activated; PRKD1, protein kinase D1; PRKDC, protein kinase, DNA-activated, catalytic polypeptide; PtdIns5P, phosphatidylinositol 5-phosphate; PtdIns4P, phosphatidylinositol 4-phosphate; PtdIns(4,5)P 2 , phosphatidylinositol 4,5-bisphosphate; PtdIns3P, phosphatidylinositol 3-phosphate; PtdIns(3,5)P 2 , phosphatidylinositol 3,5-bisphosphate; PtdIns(3,4,5)P 3 , phosphatidylinositol 3,4,5-trisphosphate; RHEB, Ras homolog enriched in brain; RPS6KB1, ribosomal protein S6 kinase, 70kDa, polypeptide 1; SQSTM1, sequestosome 1; TECPR1, tectonin b-propeller repeat containing 1; TFEB, transcription factor EB; TRIM28, tripartite motif containing 28; TSC, tuberous sclerosis; UV, ultraviolet; UVRAG, UV radiation resistance associated; WDFY3, WD repeat and FYVE domain containing 3; WIPI, WD repeat domain, phosphoinositide interacting; ZFYVE1, zinc finger, FYVE domain containing 1.Autophagy is an evolutionarily conserved and exquisitely regulated self-eating cellular process with important biological functions. Phosphatidylinositol 3-kinases (PtdIns3Ks) and phosphoinositide 3-kinases (PI3Ks) are involved in the autophagic process. Here we aim to recapitulate how 3 classes of these lipid kinases differentially regulate autophagy. Generally, activation of the class I PI3K suppresses autophagy, via the well-established PI3K-AKT-MTOR (mechanistic target of rapamycin) complex 1 (MTORC1) pathway. In contrast, the class III PtdIns3K catalytic subunit PIK3C3/Vps34 forms a protein complex with BECN1 and PIK3R4 and produces phosphatidylinositol 3-phosphate (PtdIns3P), which is required for the initiati...

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Atg38 is required for autophagy-specific phosphatidylinositol 3-kinase complex integrity

Abstract: Atg38 provides a physical linkage between the Vps15–Vps34 and Atg14–Vps30 subcomplexes to facilitate PI3-kinase complex I formation.

Cited by 104 publications

References 53 publications

Dynamics and architecture of the NRBF2-containing phosphatidylinositol 3-kinase complex I of autophagy

Dynamics and architecture of the NRBF2-containing phosphatidylinositol 3-kinase complex I of autophagy

The machinery of macroautophagy

Differential regulatory functions of three classes of phosphatidylinositol and phosphoinositide 3-kinases in autophagy

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