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O-GlcNAcylation of SPOP promotes carcinogenesis in hepatocellular carcinoma

Oncogene volume 42, pages 725–736 (2023)Cite this article

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Abstract

Aberrantly elevated O-GlcNAcylation level is commonly observed in human cancer patients, and has been proposed as a potential therapeutic target. Speckle-type POZ protein (SPOP), an important substrate adaptor of cullin3-RING ubiquitin ligase, plays a key role in the initiation and development of various cancers. However, the regulatory mechanisms governing SPOP and its function during hepatocellular carcinoma (HCC) progression remain unclear. Here, we show that, in HCC, SPOP is highly O-GlcNAcylated by O-GlcNAc transferase (OGT) at Ser96. In normal liver cells, the SPOP protein mainly localizes in the cytoplasm and mediates the ubiquitination of the oncoprotein neurite outgrowth inhibitor-B (Nogo-B) (also known as reticulon 4 B) by recognizing its N-terminal SPOP-binding consensus (SBC) motifs. However, O-GlcNAcylation of SPOP at Ser96 increases the nuclear positioning of SPOP in hepatoma cells, alleviating the ubiquitination of the Nogo-B protein, thereby promoting HCC progression in vitro and in vivo. In addition, ablation of O-GlcNAcylation by an S96A mutation increased the cytoplasmic localization of SPOP, thereby inhibiting the Nogo-B/c-FLIP cascade and HCC progression. Our findings reveal a novel post-translational modification of SPOP and identify a novel SPOP substrate, Nogo-B, in HCC. Intervention with the hyper O-GlcNAcylation of SPOP may provide a novel strategy for HCC treatment.

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Fig. 1: SPOP is dynamically O-GlcNAcylated at Ser96.
Fig. 2: O-GlcNAcylation promotes the nuclear localization of SPOP in HCC cells.
Fig. 3: SPOP O-GlcNAcylation promotes HCC progression in vitro and in vivo.
Fig. 4: SPOP-Nogo-B interaction mediates the ubiquitination of Nogo-B.
Fig. 5: SPOP ubiquitinates Nogo-B through its N-terminal domain.
Fig. 6: SPOP O-GlcNAcylation modulates the ubiquitination of Nogo-B and tumor growth.
Fig. 7: O-GlcNAcylation of SPOP strengthens Nogo-B function and correlates with HCC progression.
Fig. 8: O-GlcNAcylation of SPOP promotes HCC progression.

Data availability

The original mass spectrometry analysis data of SPOP-interacting proteins were uploaded to integrated proteome resource (iProX, PXD038208). All relevant data are available from the corresponding author on reasonable request.

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Acknowledgements

The authors acknowledge Prof. Dr. T.-C He (University of Chicago, USA) and Prof. Ding Xue (Tsinghua University, China) for providing the pAdEasy and CRISPR/Cas9 system. We also thank Prof. Bing Sun (Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, China) for kindly providing pLL3.7 and pMSCV2.2-IRES-GFP vectors.

Funding

This work was supported by the National Natural Science Foundation of China (Grant No. U20A20392, AH; 82072286, NT; 82073251, KW), the 111 Project (No. D20028, AH), the Innovative and Entrepreneurial Team of Chongqing Talents Plan, Natural Science Foundation Project of Chongqing (cstc2019jscx-dxwtBX0019, NT), Chongqing Medical Scientific Research Project (Joint project of Chongqing Health Commission and Science and Technology Bureau, 2023DBXM007, NT), Kuanren Talents Program of the Second Affiliated Hospital of Chongqing Medical University, Program for Youth Innovation in Future Medicine, Chongqing Medical University (W0036, NT; W0101, KW), and Science and Technology Research Program of Chongqing Municipal Education Commission grants HZ2021006 (NT) and JZD-M202000401 (NT).

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  1. These authors contributed equally: Peng Zhou, Wen-yi Chang, De-ao Gong.

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  1. Key Laboratory of Molecular Biology for Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing, China

    Peng Zhou, Wen-yi Chang, De-ao Gong, Lu-yi Huang, Rui Liu, Yi Liu, Jie Xia, Kai Wang, Ni Tang & Ai-long Huang

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Contributions

NT, KW and ALH designed the experiments; PZ, WYC and DAG performed and analyzed the data; PZ, WYC, DAG. LYH, RL, YL and JX contributed materials and data, and assisted in data analysis; PZ, WYC, KW and NT wrote and edited the paper.

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Zhou, P., Chang, Wy., Gong, Da. et al. O-GlcNAcylation of SPOP promotes carcinogenesis in hepatocellular carcinoma. Oncogene 42, 725–736 (2023). https://doi.org/10.1038/s41388-022-02589-z

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