scholarly journals Nedd4-dependent lysine-11-linked polyubiquitination of the tumour suppressor Beclin 1

2011 ◽  
Vol 441 (1) ◽  
pp. 399-406 ◽  
Author(s):  
Harald W. Platta ◽  
Hilde Abrahamsen ◽  
Sigrid B. Thoresen ◽  
Harald Stenmark
Keyword(s):  
Tumour Suppressor ◽  
Beclin 1 ◽  
Class Iii ◽  
Phosphatidylinositol 3 Kinase ◽  
Interacting Protein ◽  
Polyubiquitin Chains ◽  
Protein Levels ◽  
Regulatory Link ◽  
A Subunit ◽  
The Stability

Beclin 1, a subunit of the class III phosphatidylinositol 3-kinase complex, is a tumour suppressor with a central role in endocytic trafficking, cytokinesis and the cross-regulation between autophagy and apoptosis. Interestingly, not only reduced expression but also overexpression of Beclin 1 is correlated with cancer development and metastasis. Thus it seems necessary for the cell to balance the protein levels of Beclin 1. In the present study we describe a regulatory link between Beclin 1 and the ubiquitin ligase Nedd4 (neural-precursor-cell-expressed developmentally down-regulated 4). We establish Nedd4 as a novel binding partner of Beclin 1 and demonstrate that Nedd4 polyubiquitinates Beclin 1 with Lys11- and Lys63-linked chains. Importantly, Nedd4 expression controls the stability of Beclin 1, and depletion of the Beclin 1-interacting protein VPS34 causes Nedd4-mediated proteasomal degradation of Beclin 1 via Lys11-linked polyubiquitin chains. Beclin 1 is thus the first tumour suppressor reported to be controlled by Lys11-linked polyubiquitination.

scholarly journals Beclin 1 Forms Two Distinct Phosphatidylinositol 3-Kinase Complexes with Mammalian Atg14 and UVRAG

2008 ◽  
Vol 19 (12) ◽  
pp. 5360-5372 ◽  
Author(s):  
Eisuke Itakura ◽  
Chieko Kishi ◽  
Kinji Inoue ◽  
Noboru Mizushima
Keyword(s):  
Membrane Trafficking ◽  
Mammalian Cells ◽  
Coiled Coil ◽  
Pi3 Kinase ◽  
Beclin 1 ◽  
Class Iii ◽  
Phosphatidylinositol 3 Kinase ◽  
Interacting Protein ◽  
Vacuolar Protein ◽  
Phosphatidylinositol 3

Class III phosphatidylinositol 3-kinase (PI3-kinase) regulates multiple membrane trafficking. In yeast, two distinct PI3-kinase complexes are known: complex I (Vps34, Vps15, Vps30/Atg6, and Atg14) is involved in autophagy, and complex II (Vps34, Vps15, Vps30/Atg6, and Vps38) functions in the vacuolar protein sorting pathway. Atg14 and Vps38 are important in inducing both complexes to exert distinct functions. In mammals, the counterparts of Vps34, Vps15, and Vps30/Atg6 have been identified as Vps34, p150, and Beclin 1, respectively. However, orthologues of Atg14 and Vps38 remain unknown. We identified putative mammalian homologues of Atg14 and Vps38. The Vps38 candidate is identical to UV irradiation resistance-associated gene (UVRAG), which has been reported as a Beclin 1-interacting protein. Although both human Atg14 and UVRAG interact with Beclin 1 and Vps34, Atg14, and UVRAG are not present in the same complex. Although Atg14 is present on autophagic isolation membranes, UVRAG primarily associates with Rab9-positive endosomes. Silencing of human Atg14 in HeLa cells suppresses autophagosome formation. The coiled-coil region of Atg14 required for binding with Vps34 and Beclin 1 is essential for autophagy. These results suggest that mammalian cells have at least two distinct class III PI3-kinase complexes, which may function in different membrane trafficking pathways.

scholarly journals Vaccinia-Related Kinase 2 Controls the Stability of the Eukaryotic Chaperonin TRiC/CCT by Inhibiting the Deubiquitinating Enzyme USP25

2015 ◽  
Vol 35 (10) ◽  
pp. 1754-1762 ◽  
Author(s):  
Sangjune Kim ◽  
Dohyun Lee ◽  
Juhyun Lee ◽  
Haengjin Song ◽  
Hyo-Jin Kim ◽  
...  
Keyword(s):  
Protein Aggregation ◽  
Ubiquitin Proteasome System ◽  
Mutant Protein ◽  
Critical Pathway ◽  
Interacting Protein ◽  
Protein Levels ◽  
Functional Regulation ◽  
The Stability ◽  
And Function ◽  
Polyglutamine Protein

Molecular chaperones monitor the proper folding of misfolded proteins and function as the first line of defense against mutant protein aggregation in neurodegenerative diseases. The eukaryotic chaperonin TRiC is a potent suppressor of mutant protein aggregation and toxicity in early stages of disease progression. Elucidation of TRiC functional regulation will enable us to better understand the pathological mechanisms of neurodegeneration. We have previously shown that vaccinia-related kinase 2 (VRK2) downregulates TRiC protein levels through the ubiquitin-proteasome system by recruiting the E3 ligase COP1. However, although VRK2 activity was necessary in TRiC downregulation, the phosphorylated substrate was not determined. Here, we report that USP25 is a novel TRiC interacting protein that is also phosphorylated by VRK2. USP25 catalyzed deubiquitination of the TRiC protein and stabilized the chaperonin, thereby reducing accumulation of misfolded polyglutamine protein aggregates. Notably, USP25 deubiquitinating activity was suppressed when VRK2 phosphorylated the Thr680, Thr727, and Ser745residues. Impaired USP25 deubiquitinating activity after VRK2-mediated phosphorylation may be a critical pathway in TRiC protein destabilization.

scholarly journals The dynamic interaction of AMBRA1 with the dynein motor complex regulates mammalian autophagy

2010 ◽  
Vol 191 (1) ◽  
pp. 155-168 ◽  
Author(s):  
Sabrina Di Bartolomeo ◽  
Marco Corazzari ◽  
Francesca Nazio ◽  
Serafina Oliverio ◽  
Gaia Lisi ◽  
...  
Keyword(s):  
Beclin 1 ◽  
Core Complex ◽  
Autophagosome Formation ◽  
Regulatory Relationship ◽  
Interacting Protein ◽  
Dynein Motor ◽  
Regulatory Link ◽  
Dynein Light Chains ◽  
Cytoskeleton Dynamics ◽  
Catabolic Process

Autophagy is an evolutionary conserved catabolic process involved in several physiological and pathological processes such as cancer and neurodegeneration. Autophagy initiation signaling requires both the ULK1 kinase and the BECLIN 1–VPS34 core complex to generate autophagosomes, double-membraned vesicles that transfer cellular contents to lysosomes. In this study, we show that the BECLIN 1–VPS34 complex is tethered to the cytoskeleton through an interaction between the BECLIN 1–interacting protein AMBRA1 and dynein light chains 1/2. When autophagy is induced, ULK1 phosphorylates AMBRA1, releasing the autophagy core complex from dynein. Its subsequent relocalization to the endoplasmic reticulum enables autophagosome nucleation. Therefore, AMBRA1 constitutes a direct regulatory link between ULK1 and BECLIN 1–VPS34, which is required for core complex positioning and activity within the cell. Moreover, our results demonstrate that in addition to a function for microtubules in mediating autophagosome transport, there is a strict and regulatory relationship between cytoskeleton dynamics and autophagosome formation.

scholarly journals PtdIns(4,5)P2 signaling regulates ATG14 and autophagy

2016 ◽  
Vol 113 (39) ◽  
pp. 10896-10901 ◽  
Author(s):  
Xiaojun Tan ◽  
Narendra Thapa ◽  
Yihan Liao ◽  
Suyong Choi ◽  
Richard A. Anderson
Keyword(s):  
Endoplasmic Reticulum ◽  
Functional Significance ◽  
Mammalian Cells ◽  
Beclin 1 ◽  
Class Iii ◽  
Phosphatidylinositol 3 Kinase ◽  
Atg Proteins ◽  
Late Endosomes ◽  
Irradiation Resistance ◽  
Phosphatidylinositol 3

Autophagy is a regulated self-digestion pathway with fundamental roles in cell homeostasis and diseases. Autophagy is regulated by coordinated actions of a series of autophagy-related (ATG) proteins. The Barkor/ATG14(L)–VPS34 (a class III phosphatidylinositol 3-kinase) complex and its product phosphatidylinositol 3-phosphate [PtdIns(3)P] play key roles in autophagy initiation. ATG14 contains a C-terminal Barkor/ATG14(L) autophagosome-targeting sequence (BATS) domain that senses the curvature of PtdIns(3)P-containing membrane. The BATS domain also strongly binds PtdIns(4,5)P2, but the functional significance has been unclear. Here we show that ATG14 specifically interacts with type Iγ PIP kinase isoform 5 (PIPKIγi5), an enzyme that generates PtdIns(4,5)P2 in mammalian cells. Autophagosomes have associated PIPKIγi5 and PtdIns(4,5)P2 that are colocalized with late endosomes and the endoplasmic reticulum. PtdIns(4,5)P2 generation at these sites requires PIPKIγi5. Loss of PIPKIγi5 results in a loss of ATG14, UV irradiation resistance-associated gene, and Beclin 1 and a block of autophagy. PtdIns(4,5)P2 binding to the ATG14–BATS domain regulates ATG14 interaction with VPS34 and Beclin 1, and thus plays a key role in ATG14 complex assembly and autophagy initiation. This study identifies an unexpected role for PtdIns(4,5)P2 signaling in the regulation of ATG14 complex and autophagy.

scholarly journals Disrupted apolipoprotein L1-miR193a axis dedifferentiates podocytes through autophagy blockade in an APOL1 risk milieu

2019 ◽  
Vol 317 (2) ◽  
pp. C209-C225 ◽  
Author(s):  
Vinod Kumar ◽  
Kamesh Ayasolla ◽  
Alok Jha ◽  
Abheepsa Mishra ◽  
Himanshu Vashistha ◽  
...  
Keyword(s):  
Mammalian Target Of Rapamycin ◽  
Beclin 1 ◽  
Complex Iii ◽  
Class Iii ◽  
Phosphatidylinositol 3 Kinase ◽  
Protein Protein Interaction ◽  
Rich Domain ◽  
Risk Alleles ◽  
Enhanced Expression ◽  
Docking Protein

We hypothesized that a functional apolipoprotein LI (APOL1)-miR193a axis (inverse relationship) preserves, but disruption alters, the podocyte molecular phenotype through the modulation of autophagy flux. Podocyte-expressing APOL1G0 (G0-podocytes) showed downregulation but podocyte-expressing APOL1G1 (G1-podocytes) and APOL1G2 (G2-podocytes) displayed enhanced miR193a expression. G0-, G1-, and G2-podocytes showed enhanced expression of light chain (LC) 3-II and beclin-1, but a disparate expression of p62 (low in wild-type but high in risk alleles). G0-podocytes showed enhanced, whereas G1- and G2-podocytes displayed decreased, phosphorylation of Unc-51-like autophagy-activating kinase (ULK)1 and class III phosphatidylinositol 3-kinase (PI3KC3). Podocytes overexpressing miR193a (miR193a-podocytes), G1, and G2 showed decreased transcription of PIK3R3 (PI3KC3′s regulatory unit). Since 3-methyladenine (3-MA) enhanced miR193a expression but inhibited PIK3R3 transcription, it appears that 3-MA inhibits autophagy and induces podocyte dedifferentiation via miR193a generation. miR193a-, G1-, and G2-podocytes also showed decreased phosphorylation of mammalian target of rapamycin (mTOR) that could repress lysosome reformation. G1- and G2-podocytes showed enhanced expression of run domain beclin-1-interacting and cysteine-rich domain-containing protein (Rubicon); however, its silencing prevented their dedifferentiation. Docking, protein-protein interaction, and immunoprecipitation studies with antiautophagy-related gene (ATG)14L, anti-UV radiation resistance‐associated gene (UVRAG), or Rubicon antibodies suggested the formation of ATG14L complex I and UVRAG complex II in G0-podocytes and the formation of Rubicon complex III in G1- and G2-podocytes. These findings suggest that the APOL1 risk alleles favor podocyte dedifferentiation through blockade of multiple autophagy pathways.

scholarly journals The Class III PI3K/Beclin-1 Autophagic Pathway Participates in the mmLDL-Induced Upregulation of ETA Receptor in Mouse Mesenteric Arteries

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
Xi Xie ◽  
Chen Chen ◽  
Cang-Bao Xu ◽  
Jie Lin ◽  
Lei Cao ◽  
...  
Keyword(s):  
Cardiovascular Diseases ◽  
Mrna Expression ◽  
Protein Level ◽  
Beclin 1 ◽  
Mesenteric Arteries ◽  
Dose Effect ◽  
Class Iii ◽  
Protein Levels ◽  
Eta Receptor

Minimally modified low-density lipoprotein (mmLDL) is a risk factor for cardiovascular diseases. The current study explored the effect of mmLDL on the endothelin type A (ETA) receptor in mouse mesenteric arteries in vivo, as well as the role of autophagy in this process. mmLDL was injected via the caudal vein, and the Class III PI3K autophagic pathway inhibitor 3-methyladenine (3-MA) was injected intraperitoneally. The animals were divided into physiological saline (NS), mmLDL, and mmLDL + 3-MA groups. The dose-effect curve of endothelin-1- (ET-1-) induced mesenteric artery contraction was measured using myography, while ETA receptor mRNA expression was detected using real-time polymerase chain reactions, and the protein levels of the ETA receptor, class III PI3K, Beclin-1, LC3 II/I, p62, NF-κB, and p-NF-κB were observed using Western blot analysis. mmLDL significantly strengthened ET-1-induced contraction (the Emax value increased from 184.87 ± 7.46% in the NS group to 319.91 ± 20.31% in the mmLDL group (P<0.001), and the pEC50 value increased from 8.05 ± 0.05 to 9.11 ± 0.09 (P<0.01). In addition to upregulating the protein levels of Class III PI3K, Beclin-1, and LC3 II/I and downregulating that of p62, mmLDL significantly increased the mRNA expression and protein level of the ETA receptor and increased the protein level of p-NF-κB. However, these effects were significantly inhibited by 3-MA. mmLDL activates autophagy via the Class III PI3K/Beclin-1 pathway and upregulates the ETA receptor via the downstream NF-κB pathway. Understanding the effect of mmLDL on the ETA receptor and the underlying mechanisms may provide a new idea for the prevention and treatment of cardiovascular diseases.

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