Nrbf2 Protein Suppresses Autophagy by Modulating Atg14L Protein-containing Beclin 1-Vps34 Complex Architecture and Reducing Intracellular Phosphatidylinositol-3 Phosphate Levels

Zhong, Yu; Morris, Deanna H.; Jin, Lin; Patel, Mittul S.; Karunakaran, Senthil; Fu, You‐Jun; Matuszak, Emily A.; Weiss, Heidi L.; Chait, Brian T.; Wang, Qing Jun

doi:10.1074/jbc.m114.561134

Cited by 64 publications

(91 citation statements)

References 66 publications

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“…The human ortholog of Atg38 is NRBF2 (nuclear receptor binding factor 2). In parallel with the observations for Atg38 and yeast PI3KC3-C1, NRBF2 is an interactor and/or component of the mammalian PI3KC3-C1 complex (15)(16)(17). Cells from NRBF2 knockout mice show reduced autophagy, reduced PI3KC3 activity, and reduced stability of PI3KC3-C1 (15).…”

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confidence: 79%

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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mentioning

confidence: 79%

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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“…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%

“…61 NRBF2 was identified as a protein involved in autophagy, which is present only in the ATG14-containing class III PtdIns3K complex in mammalian cells. [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.…”

Section: Regulation Of Autophagy By Pik3c3 Via Other Interacting Partmentioning

confidence: 99%

“…[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%

“…2). The stratification can be viewed based on their interaction and stability: PIK3C3-PIK3R4-BECN1, the most stable and prevalent complex, constitutes the core; while ATG14 and UVRAG (UV radiation resistance associated) are combined with the core in PtdIns3K complex I (PtdIns3K-C1; ATG14-containing PtdIns3K complex) and PtdIns3K complex II (PtdIns3K-C2; UVRAG-containing PtdIns3K complex), respectively; 75-78 KIAA0226/Rubicon is a regulatory subunit that is thought to stably bind to UVRAGcontaining complexes; 77,78 AMBRA1, NRBF2 (nuclear receptor binding factor 2) and other accessory factors transiently or unstably interact with either BECN1 (e.g., AMBRA1) 79,80 or ATG14 (e.g., NRBF2), [81][82][83] thus comprising the peripheral components. 84,85 UVRAG also exists in a separate complex excluding ATG14, which holds an important role in the regulation of autophagy at the maturation step.…”

Section: Regulation Of Autophagy By Class III Ptdins3kmentioning

confidence: 99%

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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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The interplay of ROS and the PI3K/Akt pathway in autophagy regulation

Kma

Baruah²

2021

Biotech and App Biochem

154

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Autophagy causes the breakdown of damaged proteins and organelles to their constituent components. The phosphatidylinositol 3-kinase (PI3K) pathway played an important role in regulating the autophagic response of cells in response to changing reactive oxygen species (ROS) levels. The PI3K α catalytic subunit inhibits autophagy, while its β catalytic subunit promotes autophagy in response to changes in ROS levels. The downstream Akt protein acts against autophagy initiation in response to increases in ROS levels under nutrient-rich conditions. Akt acts by activating a mechanistic target of the rapamycin complex 1 (mTORC1) and by arresting autophagic gene expression. The AMP-activated protein kinase (AMPK) protein counteracts the Akt actions. mTORC1 and mTORC2 inhibit autophagy under moderate ROS levels, but under high ROS levels, mTORC2 can promote cellular senescence via autophagy. Phosphatase and tensin homolog (PTEN) protein are the negative regulators of the PI3K pathway, and it has proautophagic activities. Studies conducted on cells treated with flavonoids and ionizing radiation showed that the moderate increase in ROS levels in the flavonoid-treated groups corresponded with higher PTEN levels and lowered Akt levels leading to a higher occurrence of autophagy. In contrast, higher ROS levels evoked by ionizing radiation caused a lowering of the incidence of autophagy.

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Nrbf2 Protein Suppresses Autophagy by Modulating Atg14L Protein-containing Beclin 1-Vps34 Complex Architecture and Reducing Intracellular Phosphatidylinositol-3 Phosphate Levels

Cited by 64 publications

References 66 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

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

The interplay of ROS and the PI3K/Akt pathway in autophagy regulation

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