scholarly journals Trim39 regulates neuronal apoptosis by acting as a SUMO-targeted E3 ubiquitin-ligase for the transcription factor NFATc3

2020 ◽  
Author(s):  
Meenakshi Basu Shrivastava ◽  
Barbara Mojsa ◽  
Stéphan Mora ◽  
Ian Robbins ◽  
Guillaume Bossis ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Ubiquitin Ligase ◽  
Protein Level ◽  
Neuronal Apoptosis ◽  
Transcriptional Activity ◽  
E3 Ubiquitin Ligase ◽  
Deficient Mutant ◽  
Ubiquitin Ligase Activity ◽  
Apoptotic Activity

AbstractNFATc3 is the predominant member of the NFAT family of transcription factor in neurons, where it plays a pro-apoptotic role. Mechanisms controlling NFAT protein stability are poorly understood. Here we identify Trim39 as an E3 ubiquitin-ligase of NFATc3. Indeed, Trim39 ubiquitinates NFATc3 in vitro and in cells, whereas silencing of endogenous Trim39 decreases NFATc3 ubiquitination. We also show that Trim17 inhibits Trim39-mediated ubiquitination of NFATc3 by reducing both the E3 ubiquitin-ligase activity of Trim39 and the NFATc3/Trim39 interaction. Moreover, mutation of SUMOylation sites in NFATc3 or SUMO-interacting motif in Trim39 reduces the NFATc3/Trim39 interaction and Trim39-induced ubiquitination of NFATc3. As a consequence, silencing of Trim39 increases the protein level and transcriptional activity of NFATc3, resulting in enhanced neuronal apoptosis. Likewise, a SUMOylation-deficient mutant of NFATc3 exhibits increased stability and pro-apoptotic activity. Taken together, these data indicate that Trim39 modulates neuronal apoptosis by acting as a SUMO-targeted E3 ubiquitin-ligase for NFATc3.

scholarly journals E3 ubiquitin ligase MARCHF5 controls BAK apoptotic activity independently of BH3-only proteins

2022 ◽  
Author(s):  
Grant Dewson ◽  
Alan Shuai Huang ◽  
Hui San Chin ◽  
Boris Reljic ◽  
Tirta M Djajawi ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Apoptotic Cell ◽  
E3 Ubiquitin Ligase ◽  
Ubiquitin Ligase Activity ◽  
A Genome ◽  
Library Screen ◽  
Important Regulator ◽  
Apoptotic Activity ◽  
Bh3 Mimetic ◽  
Survival Proteins

Intrinsic apoptosis is principally governed by the BCL-2 family of proteins, but some non-BCL-2 proteins are also critical to control this process. To identify novel apoptosis regulators, we performed a genome-wide CRISPR-Cas9 library screen, and identified the mitochondrial E3 ubiquitin ligase MARCHF5/MITOL/RNF153 as an important regulator of BAK apoptotic function. Deleting MARCHF5 in diverse cell lines dependent on BAK conferred profound resistance to BH3-mimetic drugs. The loss of MARCHF5 or its E3 ubiquitin ligase activity surprisingly drove BAK to adopt an activated conformation, with resistance to BH3-mimetics afforded by the formation of inhibitory complexes with pro-survival proteins MCL-1 and BCL-XL. Importantly, these changes to BAK conformation and pro-survival association occurred independently of BH3-only proteins and influence on pro-survival proteins. This study identifies a new mechanism by which MARCHF5 regulates apoptotic cell death and provides new insight into how cancer cells respond to BH3-mimetic drugs. These data also highlight the emerging role of ubiquitin signalling in apoptosis that may be exploited therapeutically.

scholarly journals TRIB3 promotes MYC-associated lymphoma development through suppression of UBE3B-mediated MYC degradation

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Ke Li ◽  
Feng Wang ◽  
Zhao-na Yang ◽  
Ting-ting Zhang ◽  
Yu-fen Yuan ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Ubiquitin Ligase ◽  
Transcriptional Activity ◽  
Therapeutic Option ◽  
E3 Ubiquitin Ligase ◽  
Chromosomal Translocation ◽  
Tumor Burden ◽  
Self Renewal ◽  
Human Lymphoma ◽  
Almost All

AbstractThe transcription factor MYC is deregulated in almost all human cancers, especially in aggressive lymphomas, through chromosomal translocation, amplification, and transcription hyperactivation. Here, we report that high expression of tribbles homologue 3 (TRIB3) positively correlates with elevated MYC expression in lymphoma specimens; TRIB3 deletion attenuates the initiation and progression of MYC-driven lymphoma by reducing MYC expression. Mechanistically, TRIB3 interacts with MYC to suppress E3 ubiquitin ligase UBE3B-mediated MYC ubiquitination and degradation, which enhances MYC transcriptional activity, causing high proliferation and self-renewal of lymphoma cells. Use of a peptide to disturb the TRIB3-MYC interaction together with doxorubicin reduces the tumor burden in MycEμ mice and patient-derived xenografts. The pathophysiological relevance of UBE3B, TRIB3 and MYC is further demonstrated in human lymphoma. Our study highlights a key mechanism for controlling MYC expression and a potential therapeutic option for treating lymphomas with high TRIB3-MYC expression.

scholarly journals The multi-subunit GID/CTLH E3 ubiquitin ligase promotes cell proliferation and targets the transcription factor Hbp1 for degradation

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Fabienne Lampert ◽  
Diana Stafa ◽  
Algera Goga ◽  
Martin Varis Soste ◽  
Samuel Gilberto ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Hierarchical Organization ◽  
Negative Regulator ◽  
Cell Cycle Exit ◽  
Cellular Analysis ◽  
Ubiquitin Ligase Activity

In yeast, the glucose-induced degradation-deficient (GID) E3 ligase selectively degrades superfluous gluconeogenic enzymes. Here, we identified all subunits of the mammalian GID/CTLH complex and provide a comprehensive map of its hierarchical organization and step-wise assembly. Biochemical reconstitution demonstrates that the mammalian complex possesses inherent E3 ubiquitin ligase activity, using Ube2H as its cognate E2. Deletions of multiple GID subunits compromise cell proliferation, and this defect is accompanied by deregulation of critical cell cycle markers such as the retinoblastoma (Rb) tumor suppressor, phospho-Histone H3 and Cyclin A. We identify the negative regulator of pro-proliferative genes Hbp1 as a bonafide GID/CTLH proteolytic substrate. Indeed, Hbp1 accumulates in cells lacking GID/CTLH activity, and Hbp1 physically interacts and is ubiquitinated in vitro by reconstituted GID/CTLH complexes. Our biochemical and cellular analysis thus demonstrates that the GID/CTLH complex prevents cell cycle exit in G1, at least in part by degrading Hbp1.

scholarly journals The E3 Ubiquitin Ligase TBK1 Mediates the Degradation of Multiple Picornavirus VP3 Proteins by Phosphorylation and Ubiquitination

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Dan Li ◽  
Wenping Yang ◽  
Jingjing Ren ◽  
Yi Ru ◽  
Keshan Zhang ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
E3 Ubiquitin Ligase ◽  
Adaptor Protein ◽  
Ubiquitin Ligase Activity ◽  
Lysine Residues ◽  
Innate Antiviral Immunity

ABSTRACT TANK-binding kinase 1 (TBK1) is essential for interferon beta (IFN-β) production and innate antiviral immunity. However, other, additional functions of TBK1 have remained elusive. Here, we showed that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. Further evidence showed that TBK1 could also be self-ubiquitylated in vivo. Importantly, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Mechanistically, TBK1 phosphorylated multiple picornavirus VP3 proteins at serine residues and ubiquitinated them via K63-linked ubiquitination at lysine residues. In addition, the C426 and C605 residues of TBK1 were not essential for TBK1 innate immunity activity; however, these residues were required for degradation of multiple picornavirus VP3 proteins and for its E3 ubiquitin ligase activity. Hence, our findings identified a novel role of TBK1 in regulating the virus life cycle and provided new insights into the molecular mechanisms of TBK1-mediated antiviral response. IMPORTANCE TBK1 is an important adaptor protein required for innate immune response to viruses, but its other functions were unknown. In this study, we found that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. In addition, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Our report provides evidence that TBK1 plays a role in viral protein degradation.

scholarly journals The RBCC GeneRFP2(Leu5) Encodes a Novel Transmembrane E3 Ubiquitin Ligase Involved in ERAD

2007 ◽  
Vol 18 (5) ◽  
pp. 1670-1682 ◽  
Author(s):  
Mikael Lerner ◽  
Martin Corcoran ◽  
Diana Cepeda ◽  
Michael L. Nielsen ◽  
Roman Zubarev ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Fluorescent Protein ◽  
Transmembrane Domain ◽  
Yellow Fluorescent Protein ◽  
E3 Ubiquitin Ligase ◽  
Coiled Coil ◽  
Ring Finger ◽  
Ubiquitin Ligase Activity

RFP2, a gene frequently lost in various malignancies, encodes a protein with RING finger, B-box, and coiled-coil domains that belongs to the RBCC/TRIM family of proteins. Here we demonstrate that Rfp2 is an unstable protein with auto-polyubiquitination activity in vivo and in vitro, implying that Rfp2 acts as a RING E3 ubiquitin ligase. Consequently, Rfp2 ubiquitin ligase activity is dependent on an intact RING domain, as RING deficient mutants fail to drive polyubiquitination in vitro and are stabilized in vivo. Immunopurification and tandem mass spectrometry enabled the identification of several putative Rfp2 interacting proteins localized to the endoplasmic reticulum (ER), including valosin-containing protein (VCP), a protein indispensable for ER-associated degradation (ERAD). Importantly, we also show that Rfp2 regulates the degradation of the known ER proteolytic substrate CD3-δ, but not the N-end rule substrate Ub-R-YFP (yellow fluorescent protein), establishing Rfp2 as a novel E3 ligase involved in ERAD. Finally, we show that Rfp2 contains a C-terminal transmembrane domain indispensable for its localization to the ER and that Rfp2 colocalizes with several ER-resident proteins as analyzed by high-resolution immunostaining. In summary, these data are all consistent with a function for Rfp2 as an ERAD E3 ubiquitin ligase.

scholarly journals Liganded ERα Stimulates the E3 Ubiquitin Ligase Activity of UBE3C to Facilitate Cell Proliferation

2015 ◽  
Vol 29 (11) ◽  
pp. 1646-1657 ◽  
Author(s):  
Maiko Okada ◽  
Fumiaki Ohtake ◽  
Hiroyuki Nishikawa ◽  
Wenwen Wu ◽  
Yasushi Saeki ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Cyclin B1 ◽  
Dependent Manner ◽  
Ubiquitin Ligase Activity ◽  
Dependent Cell ◽  
Estrogen Antagonist

Abstract Estrogen receptor (ER)α is a well-characterized ligand-dependent transcription factor. However, the global picture of its nongenomic functions remains to be illustrated. Here, we demonstrate a novel function of ERα during mitosis that facilitates estrogen-dependent cell proliferation. An E3 ubiquitin ligase, UBE3C, was identified in an ERα complex from estrogen-treated MCF-7 breast cancer cells arrested at mitosis. UBE3C interacts with ERα during mitosis in an estrogen-dependent manner. In vitro, estrogen dramatically stimulates the E3 activity of UBE3C in the presence of ERα. This effect was inhibited by the estrogen antagonist tamoxifen. Importantly, estrogen enhances the ubiquitination of cyclin B1 (CCNB1) and destabilizes CCNB1 during mitosis in a manner dependent on endogenous UBE3C. ERα, UBE3C, and CCNB1 colocalize in prophase nuclei and at metaphase spindles before CCNB1 is degraded in anaphase. Depletion of UBE3C attenuates estrogen-dependent cell proliferation without affecting the transactivation function of ERα. Collectively, these results demonstrate a novel ligand-dependent action of ERα that stimulates the activity of an E3 ligase. The mitotic role of estrogen may contribute to its effects on proliferation in addition to its roles in target gene expression.

scholarly journals E3 Ubiquitin Ligase SPL2 Is a Lanthanide-Binding Protein

2021 ◽  
Vol 22 (11) ◽  
pp. 5712
Author(s):  
Michał Tracz ◽  
Ireneusz Górniak ◽  
Andrzej Szczepaniak ◽  
Wojciech Białek
Keyword(s):  
Ubiquitin Ligase ◽  
Conformational Changes ◽  
Protein Interactions ◽  
E3 Ubiquitin Ligase ◽  
Lanthanide Ions ◽  
E3 Ligases ◽  
Fluorescence Measurements ◽  
Partial Unfolding

The SPL2 protein is an E3 ubiquitin ligase of unknown function. It is one of only three types of E3 ligases found in the outer membrane of plant chloroplasts. In this study, we show that the cytosolic fragment of SPL2 binds lanthanide ions, as evidenced by fluorescence measurements and circular dichroism spectroscopy. We also report that SPL2 undergoes conformational changes upon binding of both Ca2+ and La3+, as evidenced by its partial unfolding. However, these structural rearrangements do not interfere with SPL2 enzymatic activity, as the protein retains its ability to auto-ubiquitinate in vitro. The possible applications of lanthanide-based probes to identify protein interactions in vivo are also discussed. Taken together, the results of this study reveal that the SPL2 protein contains a lanthanide-binding site, showing for the first time that at least some E3 ubiquitin ligases are also capable of binding lanthanide ions.

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