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A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma

BACKGROUND & AIMS: N(6)-methyladenosine (m(6)A) modification plays a critical role in progression of hepatocellular carcinoma (HCC), and aerobic glycolysis is a hallmark of cancer including HCC. However, the role of YTHDF3, one member of the core readers of the m(6)A pathway, in aerobic glycolys...

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Autores principales: Zhou, Rong, Ni, Wen, Qin, Chao, Zhou, Yunxia, Li, Yuqing, Huo, Jianping, Bian, Lijuan, Zhou, Aijun, Li, Jianming
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9724358/
https://www.ncbi.nlm.nih.gov/pubmed/36471428
http://dx.doi.org/10.1186/s13046-022-02538-4
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author Zhou, Rong
Ni, Wen
Qin, Chao
Zhou, Yunxia
Li, Yuqing
Huo, Jianping
Bian, Lijuan
Zhou, Aijun
Li, Jianming
author_facet Zhou, Rong
Ni, Wen
Qin, Chao
Zhou, Yunxia
Li, Yuqing
Huo, Jianping
Bian, Lijuan
Zhou, Aijun
Li, Jianming
author_sort Zhou, Rong
collection PubMed
description BACKGROUND & AIMS: N(6)-methyladenosine (m(6)A) modification plays a critical role in progression of hepatocellular carcinoma (HCC), and aerobic glycolysis is a hallmark of cancer including HCC. However, the role of YTHDF3, one member of the core readers of the m(6)A pathway, in aerobic glycolysis and progression of HCC is still unclear. METHODS: Expression levels of YTHDF3 in carcinoma and surrounding tissues of HCC patients were evaluated by immunohistochemistry. Loss and gain-of-function experiments in vitro and in vivo were used to assess the effects of YTHDF3 on HCC cell proliferation, migration and invasion. The role of YTHDF3 in hepatocarcinogenesis was observed in a chemically induced HCC model with Ythdf3(−/−) mice. Untargeted metabolomics and glucose metabolism phenotype assays were performed to evaluate relationship between YTHDF3 and glucose metabolism. The effect of YTHDF3 on PFKL was assessed by methylated RNA immunoprecipitation assays (MeRIP). Co-immunoprecipitation and immunofluorescence assays were performed to investigate the connection between YTHDF3 and PFKL. RESULTS: We found YTHDF3 expression was greatly upregulated in carcinoma tissues and it was correlated with poor prognosis of HCC patients. Gain-of-function and loss-of-function assays demonstrated YTHDF3 promoted proliferation, migration and invasion of HCC cells in vitro, and YTHDF3 knockdown inhibited xenograft tumor growth and lung metastasis of HCC cells in vivo. YTHDF3 knockout significantly suppressed hepatocarcinogenesis in chemically induced mice model. Mechanistically, YTHDF3 promoted aerobic glycolysis by promoting phosphofructokinase PFKL expression at both mRNA and protein levels. MeRIP assays showed YTHDF3 suppressed PFKL mRNA degradation via m(6)A modification. Surprisingly, PFKL positively regulated YTHDF3 protein expression, not as a glycolysis rate-limited enzyme, and PFKL knockdown effectively rescued the effects of YTHDF3 overexpression on proliferation, migration and invasion ability of Sk-Hep-1 and HepG2 cells. Notably, co-immunoprecipitation assays demonstrated PFKL interacted with YTHDF3 via EFTUD2, a core subunit of spliceosome involved in pre-mRNA splicing process, and ubiquitination assays showed PFKL could positively regulate YTHDF3 protein expression via inhibiting ubiquitination of YTHDF3 protein by EFTUD2. CONCLUSIONS: our study uncovers the key role of YTHDF3 in HCC, characterizes a positive functional loop between YTHDF3 and phosphofructokinase PFKL in glucose metabolism of HCC, and suggests the connection between pre-mRNA splicing process and m(6)A modification. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13046-022-02538-4.
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spelling pubmed-97243582022-12-07 A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma Zhou, Rong Ni, Wen Qin, Chao Zhou, Yunxia Li, Yuqing Huo, Jianping Bian, Lijuan Zhou, Aijun Li, Jianming J Exp Clin Cancer Res Research BACKGROUND & AIMS: N(6)-methyladenosine (m(6)A) modification plays a critical role in progression of hepatocellular carcinoma (HCC), and aerobic glycolysis is a hallmark of cancer including HCC. However, the role of YTHDF3, one member of the core readers of the m(6)A pathway, in aerobic glycolysis and progression of HCC is still unclear. METHODS: Expression levels of YTHDF3 in carcinoma and surrounding tissues of HCC patients were evaluated by immunohistochemistry. Loss and gain-of-function experiments in vitro and in vivo were used to assess the effects of YTHDF3 on HCC cell proliferation, migration and invasion. The role of YTHDF3 in hepatocarcinogenesis was observed in a chemically induced HCC model with Ythdf3(−/−) mice. Untargeted metabolomics and glucose metabolism phenotype assays were performed to evaluate relationship between YTHDF3 and glucose metabolism. The effect of YTHDF3 on PFKL was assessed by methylated RNA immunoprecipitation assays (MeRIP). Co-immunoprecipitation and immunofluorescence assays were performed to investigate the connection between YTHDF3 and PFKL. RESULTS: We found YTHDF3 expression was greatly upregulated in carcinoma tissues and it was correlated with poor prognosis of HCC patients. Gain-of-function and loss-of-function assays demonstrated YTHDF3 promoted proliferation, migration and invasion of HCC cells in vitro, and YTHDF3 knockdown inhibited xenograft tumor growth and lung metastasis of HCC cells in vivo. YTHDF3 knockout significantly suppressed hepatocarcinogenesis in chemically induced mice model. Mechanistically, YTHDF3 promoted aerobic glycolysis by promoting phosphofructokinase PFKL expression at both mRNA and protein levels. MeRIP assays showed YTHDF3 suppressed PFKL mRNA degradation via m(6)A modification. Surprisingly, PFKL positively regulated YTHDF3 protein expression, not as a glycolysis rate-limited enzyme, and PFKL knockdown effectively rescued the effects of YTHDF3 overexpression on proliferation, migration and invasion ability of Sk-Hep-1 and HepG2 cells. Notably, co-immunoprecipitation assays demonstrated PFKL interacted with YTHDF3 via EFTUD2, a core subunit of spliceosome involved in pre-mRNA splicing process, and ubiquitination assays showed PFKL could positively regulate YTHDF3 protein expression via inhibiting ubiquitination of YTHDF3 protein by EFTUD2. CONCLUSIONS: our study uncovers the key role of YTHDF3 in HCC, characterizes a positive functional loop between YTHDF3 and phosphofructokinase PFKL in glucose metabolism of HCC, and suggests the connection between pre-mRNA splicing process and m(6)A modification. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13046-022-02538-4. BioMed Central 2022-12-06 /pmc/articles/PMC9724358/ /pubmed/36471428 http://dx.doi.org/10.1186/s13046-022-02538-4 Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research
Zhou, Rong
Ni, Wen
Qin, Chao
Zhou, Yunxia
Li, Yuqing
Huo, Jianping
Bian, Lijuan
Zhou, Aijun
Li, Jianming
A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma
title A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma
title_full A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma
title_fullStr A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma
title_full_unstemmed A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma
title_short A functional loop between YTH domain family protein YTHDF3 mediated m(6)A modification and phosphofructokinase PFKL in glycolysis of hepatocellular carcinoma
title_sort functional loop between yth domain family protein ythdf3 mediated m(6)a modification and phosphofructokinase pfkl in glycolysis of hepatocellular carcinoma
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9724358/
https://www.ncbi.nlm.nih.gov/pubmed/36471428
http://dx.doi.org/10.1186/s13046-022-02538-4
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