Dataset Information

Trabid inhibits hepatocellular carcinoma growth and metastasis by cleaving RNF8-induced K63 ubiquitination of Twist1.

ABSTRACT: TRAF-binding domain (Trabid), one of deubiquitination enzymes, was recently reported to activate Wnt/ β-catenin signaling pathway. However, the role of Trabid in tumors including hepatocellular carcinoma (HCC) and the underlying mechanisms controlling its activity remain poorly understood. Here, we report that Trabid is significantly downregulated in HCC tumor samples and cell lines compared with normal controls and that its expression level is negatively correlated with HCC pathological grading, recurrence, and metastasis. The reintroduction of Trabid expression in tumor cells significantly decreases HCC progression as well as pulmonary metastasis. The effect of Trabid on HCC development occurs at least partially through regulation of Twist1 activity. Mechanistically, Trabid forms a complex with Twist1 and specifically cleaves RNF8-induced K63-linked poly-ubiquitin chains from Twist1, which enhances the association of Twist1 with β-TrCP1 and allows for subsequent K48-linked ubiquitination of Twist1. Knockdown of Trabid increases K63-linked ubiquitination, but abrogates K48-linked ubiquitination and degradation of Twist1, thus enhancing HCC growth and metastasis. Interestingly, Twist1 negatively regulates the promoter activity of Trabid, indicating that a double-negative feedback loop exists. Our findings also identify an essential role for activation of Trabid by AKT-mediated phosphorylation at Ser78/Thr117 in negatively regulating Twist1 signaling, which further provides insights into the mechanisms by which Trabid regulates Twist1 ubiquitination. Our results reveal that Trabid is a previously unrecognized inhibitor of HCC progression and metastasis, which sheds light on new strategies for HCC treatment.

SUBMITTER: Zhu Y

PROVIDER: S-EPMC6329825 | biostudies-other | 2019 Jan

REPOSITORIES: biostudies-other

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Trabid inhibits hepatocellular carcinoma growth and metastasis by cleaving RNF8-induced K63 ubiquitination of Twist1.

Zhu Yuekun Y Qu Chao C Hong Xuehui X Jia Yanyan Y Lin Meihua M Luo Yunmei Y Lin Fengqin F Xie Xiaolong X Xie Xiaoqi X Huang Juan J Wu Qin Q Qiu Xingfeng X Piao Daxun D Xing Yanwei Y Yu Tian T Lu Yuanfu Y Huang Qiang Q Yu Changyin C Jin Junfei J Zhang Zhiyong Z Zhang Zhiyong Z

Cell death and differentiation 20180510 2

TRAF-binding domain (Trabid), one of deubiquitination enzymes, was recently reported to activate Wnt/ β-catenin signaling pathway. However, the role of Trabid in tumors including hepatocellular carcinoma (HCC) and the underlying mechanisms controlling its activity remain poorly understood. Here, we report that Trabid is significantly downregulated in HCC tumor samples and cell lines compared with normal controls and that its expression level is negatively correlated with HCC pathological grading ...[more]

PMID: 29748601

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Project description:UVRAG (UV radiation resistance associated) is an important regulator of mammalian macroautophagy/autophagy by interacting with BECN1, PIK3C3, and RUBCN. Phosphorylation of UVRAG by MTORC1 negatively regulates autophagosome maturation under nutrient-enriched conditions. However, how UVRAG ubiquitination is regulated is still unknown. Here we report that UVRAG is ubiquitinated by SMURF1 at lysine residues 517 and 559, which decreases the association of UVRAG with RUBCN and promotes autophagosome maturation. However, the deubiquitinase ZRANB1 specifically cleaves SMURF1-induced K29 and K33-linked polyubiquitin chains from UVRAG, thereby increasing the binding of UVRAG to RUBCN and inhibiting autophagy flux. We also demonstrate that CSNK1A1-mediated UVRAG phosphorylation at Ser522 disrupts the binding of SMURF1 to UVRAG through PPxY motif and blocks UVRAG ubiquitination-mediated autophagosome maturation. Interestingly, ZRANB1 is phosphorylated at Thr35, and Ser209 residues by CSNK1A1, and this phosphorylation activates its deubiquitinating activity. Importantly, we provide in vitro and in vivo evidence that UVRAG ubiquitination at lysine residues 517 and 559 or prevention of Ser522 phosphorylation by D4476, a CSNK1A1 inhibitor, enhances the lysosomal degradation of EGFR, which significantly inhibits hepatocellular carcinoma (HCC) growth. Furthermore, UVRAG S522 phosphorylation levels correlate with ZRANB1 T35/S209 phosphorylation levels and poor prognosis in HCC patients. These findings identify a novel molecular mechanism by which ubiquitination and phosphorylation of UVRAG regulate its function in autophagosome maturation and HCC growth, encouraging further study of their potential therapeutic implications. Abbreviations: ATG: autophagy related; BafA1: bafilomycin A1; BECN1: beclin 1; CHX: cycloheximide; CSNK1A1/CK1α: casein kinase 1 alpha 1; CQ: chloroquine; DUB: deubiquitinase; EBSS: Earle's balanced salt solution; EGF: epidermal growth factor; GFP: green fluorescent protein; GST: glutathione S-transferase; HBSS: Hanks balanced salts solution; HCC: hepatocellular carcinoma; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MEFs: mouse embryo fibroblasts; mRFP: monomeric red fluorescent protein; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PTMs: post-translational modifications; RUBCN: rubicon autophagy regulator; siRNA: small interfering RNA; SMURF1: SMAD specific E3 ubiquitin protein ligase 1; SQSTM1: sequestosome 1; Ub-AMC: ubiquitin-7-amido-4-methylcoumarin: a fluorogenic substrate; UVRAG: UV radiation resistance associated; ZRANB1/TRABID: zinc finger RANBP2-type containing 1.

Project description:BACKGROUND: Hepatocellular carcinoma (HCC) usually has a dismal prognosis because of its limited response to current pharmacotherapy and high metastatic rate. Sulfated oligosaccharide has been confirmed as having potent antitumor activities against solid tumors. Here, we explored the preclinical effects and molecular mechanisms of isomalto oligosaccharide sulfate (IMOS), another novel sulfated oligosaccharide, in HCC cell lines and a xenograft model. METHODS: The effects of IMOS on HCC proliferation, apoptosis, adhesion, migration, and invasiveness in vitro were assessed by cell counting, flow cytometry, adhesion, wound healing, and transwell assays, respectively. The roles of IMOS on HCC growth and metastasis in xenograft models were evaluated by tumor volumes and fluorescent signals. Total and phosphorylated protein levels of AKT, ERK, and JNK as well as total levels of c-MET were detected by Western blotting. IMOS-regulated genes were screened by quantitative reverse-transcription PCR (qRT-PCR) array in HCCLM3-red fluorescent protein (RFP) xenograft tissues and then confirmed by qRT-PCR in HepG2 and Hep3B cells. RESULTS: IMOS markedly inhibited cell proliferation and induced cell apoptosis of HCCLM3, HepG2, and Bel-7402 cells and also significantly suppressed cell adhesion, migration, and invasion of HCCLM3 in vitro. At doses of 60 and 90 mg/kg/d, IMOS displayed robust inhibitory effects on HCC growth and metastasis without obvious side effects in vivo. The levels of pERK, tERK, and pJNK as well as c-MET were significantly down-regulated after treatment with 16 mg/mL IMOS. No obvious changes were found in the levels of pAkt, tAkt, and tJNK. Ten differentially expressed genes were screened from HCCLM3-RFP xenograft tissues after treatment with IMOS at a dose of 90 mg/kg/d. Similar gene expression profiles were confirmed in HepG2 and Hep3B cells after treatment with 16 mg/mL IMOS. CONCLUSIONS: IMOS is a potential anti-HCC candidate through inhibition of ERK and JNK signaling independent of p53 and worth studying further in patients with HCC, especially at advanced stages.

Dataset Information

Trabid inhibits hepatocellular carcinoma growth and metastasis by cleaving RNF8-induced K63 ubiquitination of Twist1.

Publications

Trabid inhibits hepatocellular carcinoma growth and metastasis by cleaving RNF8-induced K63 ubiquitination of Twist1.

Similar Datasets

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