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E3 ligase ZFP91 inhibits Hepatocellular Carcinoma Metabolism Reprogramming by regulating PKM splicing.

ABSTRACT: Rationale: Hepatocellular carcinoma (HCC) is one of the most lethal cancers, and few molecularly targeted anticancer therapies have been developed to treat it. Thus, the identification of new therapeutic targets is urgent. Metabolic reprogramming is an important hallmark of cancer. However, how ubiquitin ligases are involved in the regulation of cancer metabolism remains poorly understood. Methods: RT-PCR, western blot and IHC were used to determine ZFP91 expression. RNAi, cell proliferation, colony formation and transwell assays were used to determine the in vitro functions of ZFP91. Mouse xenograft models were used to study the in vivo effects of ZFP91. Co-IP together with mass spectrometry or western blot was utilized to investigate protein-protein interaction. Ubiquitination was analyzed using IP together with western blot. RNA splicing was assessed by using RT-PCR followed by restriction digestion. Lactate production and glucose uptake assays were used to analyze cancer metabolism. Results: We identified that an E3 ligase zinc finger protein 91 (ZFP91) suppressed HCC metabolic reprogramming, cell proliferation and metastasis in vitro and in vivo. Mechanistically, ZFP91 promoted the Lys48-linked ubiquitination of the oncoprotein hnRNP A1 at lysine 8 and proteasomal degradation, thereby inhibiting hnRNP A1-dependent PKM splicing, subsequently resulting in higher PKM1 isoform formation and lower PKM2 isoform formation and suppressing HCC glucose metabolism reprogramming, cell proliferation and metastasis. Moreover, HCC patients with lower levels of ZFP91 have poorer prognoses, and ZFP91 is an independent prognostic factor for patients with HCC. Conclusions: Our study identifies ZFP91 as a tumor suppressor of hepatocarcinogenesis and HCC metabolism reprogramming and proposes it as a novel prognostic biomarker and therapeutic target of HCC.

SUBMITTER: Chen

PROVIDER: S-EPMC7392027 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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E3 ligase ZFP91 inhibits Hepatocellular Carcinoma Metabolism Reprogramming by regulating PKM splicing.

Chen De Wang Yanjie Y Lu Ruixun R Jiang Xiaofeng X Chen Xinhui X Meng Nan N Chen Min M Xie Shan S Yan Guang-Rong GR

Theranostics 20200709 19

<b>Rationale:</b> Hepatocellular carcinoma (HCC) is one of the most lethal cancers, and few molecularly targeted anticancer therapies have been developed to treat it. Thus, the identification of new therapeutic targets is urgent. Metabolic reprogramming is an important hallmark of cancer. However, how ubiquitin ligases are involved in the regulation of cancer metabolism remains poorly understood. <b>Methods:</b> RT-PCR, western blot and IHC were used to determine ZFP91 expression. RNAi, cell pro ...[more]

PMID: 32754263

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Project description:BackgroundThe E2F family of transcription factors play a crucial role in the development of various cancers. However, E2F members lack targetable binding pockets and are typically considered "undruggable". Unlike canonical small-molecule therapeutics, molecular glues mediate new E3 ligase-protein interactions to induce selective proteasomal degradation, which represents an attractive option to overcome these limitations.MethodsHuman proteome microarray was utilized to identify a natural product-derived molecular glue for targeting E2F2 degradation. Co-IP analysis with stable isotope labeling of amino acids in cell culture (SILAC)-based quantitative proteomics was carried out to further explore the E3 ligase for E2F2 degradation.FindingsIn this study, we identified a molecular glue bufalin, which significantly promoted E2F2 degradation. Unexpectedly, E2F2 underwent ubiquitination and proteasomal degradation via a previously undisclosed atypical E3 ligase, zinc finger protein 91 (ZFP91). In particular, we observed that bufalin markedly promoted E2F2-ZFP91 complex formation, thereby leading to E2F2 polyubiquitination via K48-linked ubiquitin chains for degradation. E2F2 degradation subsequently caused transcriptional suppression of multiple oncogenes including c-Myc, CCNE1, CCNE2, MCM5 and CDK1, and inhibited hepatocellular carcinoma growth in vitro and in vivo.InterpretationCollectively, our findings open up a new direction for transcription factors degradation by targeting atypical E3 ligase ZFP91. Meanwhile, the chemical knockdown strategy with molecular glue may promote innovative transcription factor degrader development in cancer therapy.FundingThis work was financially supported by the National Key Research and Development Project of China (2022YFC3501601), National Natural Sciences Foundation of China (81973505, 82174008, 82030114), and China Postdoctoral Science Foundation (2019M650396), the Fundamental Research Funds for the Central Universities.

Dataset Information

E3 ligase ZFP91 inhibits Hepatocellular Carcinoma Metabolism Reprogramming by regulating PKM splicing.

Publications

E3 ligase ZFP91 inhibits Hepatocellular Carcinoma Metabolism Reprogramming by regulating PKM splicing.

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