The role of the HIF-1α/ALYREF/PKM2 axis in glycolysis and tumorigenesis of bladder cancer
Jing-Zi Wang
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorWei Zhu
Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorJie Han
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorXiao Yang
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorRui Zhou
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorHong-Cheng Lu
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorHao Yu
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorWen-Bo Yuan
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorPeng-Chao Li
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorJun Tao
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorCorresponding Author
Qiang Lu
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Correspondence
Qiang Lu and Haiwei Yang, Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China
Email: [email protected]; [email protected]
Ji-Fu Wei, Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210000, P. R. China
Email: [email protected]
Search for more papers by this authorCorresponding Author
Ji-Fu Wei
Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Correspondence
Qiang Lu and Haiwei Yang, Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China
Email: [email protected]; [email protected]
Ji-Fu Wei, Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210000, P. R. China
Email: [email protected]
Search for more papers by this authorCorresponding Author
Haiwei Yang
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Correspondence
Qiang Lu and Haiwei Yang, Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China
Email: [email protected]; [email protected]
Ji-Fu Wei, Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210000, P. R. China
Email: [email protected]
Search for more papers by this authorJing-Zi Wang
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorWei Zhu
Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorJie Han
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorXiao Yang
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
These authors contributed equally to this study
Search for more papers by this authorRui Zhou
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorHong-Cheng Lu
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorHao Yu
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorWen-Bo Yuan
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorPeng-Chao Li
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorJun Tao
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Search for more papers by this authorCorresponding Author
Qiang Lu
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Correspondence
Qiang Lu and Haiwei Yang, Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China
Email: [email protected]; [email protected]
Ji-Fu Wei, Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210000, P. R. China
Email: [email protected]
Search for more papers by this authorCorresponding Author
Ji-Fu Wei
Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Correspondence
Qiang Lu and Haiwei Yang, Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China
Email: [email protected]; [email protected]
Ji-Fu Wei, Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210000, P. R. China
Email: [email protected]
Search for more papers by this authorCorresponding Author
Haiwei Yang
Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, 210000 P. R. China
Correspondence
Qiang Lu and Haiwei Yang, Department of Urology, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210000, Jiangsu, P. R. China
Email: [email protected]; [email protected]
Ji-Fu Wei, Research Division of Clinical Pharmacology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210000, P. R. China
Email: [email protected]
Search for more papers by this authorAbstract
Background
As a rate-limiting enzyme of glycolysis, pyruvate kinase muscle isozyme M2 (PKM2) participates in tumor metabolism and growth. The regulatory network of PKM2 in cancer is complex and has not been fully studied in bladder cancer. The 5-methylcytidine (m5C) modification in PKM2 mRNA might participate in the pathogenesis of bladder cancer and need to be further clarified. This study aimed to investigate the biological function and regulatory mechanism of PKM2 in bladder cancer.
Methods
The expression of PKM2 and Aly/REF export factor (ALYREF) was measured by Western blotting, qRT-PCR, and immunohistochemistry. The bioprocesses of bladder cancer cells were demonstrated by a series of experiments in vitro and in vivo. RNA immunoprecipitation, RNA-sequencing, and dual-luciferase reporter assays were conducted to explore the potential regulatory mechanisms of PKM2 in bladder cancer.
Results
In bladder cancer, we first demonstrated that ALYREF stabilized PKM2 mRNA and bound to its m5C sites in 3′-untranslated regions. Overexpression of ALYREF promoted bladder cancer cell proliferation by PKM2-mediated glycolysis. Furthermore, high expression of PKM2 and ALYREF predicted poor survival in bladder cancer patients. Finally, we found that hypoxia-inducible factor-1alpha (HIF-1α) indirectly up-regulated the expression of PKM2 by activating ALYREF in addition to activating its transcription directly.
Conclusions
The m5C modification in PKM2 mRNA in the HIF-1α/ALYREF/PKM2 axis may promote the glucose metabolism of bladder cancer, providing a new promising therapeutic target for bladder cancer.
CONFLICT OF INTEREST
The authors declared no conflicts.
Supporting Information
| Filename | Description |
|---|---|
| cac212158-sup-0001-SuppMat.docx3.1 MB | Supporing Infromation |
| cac212158-sup-0002-SuppMat.pdf2.1 MB | Supporing Infromation |
| cac212158-sup-0003-SuppMat.pdf2 MB | Supporing Infromation |
Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
REFERENCES
- 1Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2018.
- 2Feng RM, Zong YN, Cao SM, Xu RH. Current cancer situation in China: good or bad news from the 2018 Global Cancer Statistics? Cancer Communications (London, England). 2019; 39(1): 22.
- 3Massari F, Ciccarese C, Santoni M, Iacovelli R, Mazzucchelli R, Piva F, et al. Metabolic phenotype of bladder cancer. Cancer Treat Rev. 2016; 45: 46-57.
- 4Vander Heiden MG, Cantley LC, Thompson CB. Understanding the Warburg effect: the metabolic requirements of cell proliferation. Science. 2009; 324(5930): 1029-33.
- 5Icard P, Shulman S, Farhat D, Steyaert JM, Alifano M, Lincet H. How the Warburg effect supports aggressiveness and drug resistance of cancer cells? Drug Resist Updat. 2018; 38: 1-11.
- 6Dong G, Mao Q, Xia W, Xu Y, Wang J, Xu L, et al. PKM2 and cancer: The function of PKM2 beyond glycolysis. Oncol Lett. 2016; 11(3): 1980-86.
- 7Hu H, Tu W, Chen Y, Zhu M, Jin H, Huang T, et al. The combination of PKM2 overexpression and M2 macrophages infiltration confers a poor prognosis for PDAC patients. J Cancer. 2020; 11(8): 2022-31.
- 8Li TE, Wang S, Shen XT, Zhang Z, Chen M, Wang H, et al. PKM2 Drives Hepatocellular Carcinoma Progression by Inducing Immunosuppressive Microenvironment. Front Immunol. 2020; 11:589997.
- 9Papadaki C, Manolakou S, Lagoudaki E, Pontikakis S, Ierodiakonou D. Correlation of PKM2 and CD44 Protein Expression with Poor Prognosis in Platinum-Treated Epithelial Ovarian Cancer: A Retrospective Study. Cancers. 2020; 12(4).
- 10Xiao H, Zhang L, Chen Y, Zhou C, Wang X, Wang D. PKM2 Promotes Breast Cancer Progression by Regulating Epithelial Mesenchymal Transition. Analytical Cellular Pathology (Amsterdam). 2020; 2020:8396023.
10.1155/2020/8396023 Google Scholar
- 11Bian Z, Zhang J, Li M, Feng Y, Wang X, Zhang J, et al. LncRNA-FEZF1-AS1 Promotes Tumor Proliferation and Metastasis in Colorectal Cancer by Regulating PKM2 Signaling. Clin Cancer Res. 2018; 24(19): 4808-19.
- 12Dayton TL, Gocheva V, Miller KM, Israelsen WJ, Bhutkar A, Clish CB, et al. Germline loss of PKM2 promotes metabolic distress and hepatocellular carcinoma. Genes Dev. 2016; 30(9): 1020-33.
10.1101/gad.278549.116 Google Scholar
- 13Morita M, Sato T, Nomura M, Sakamoto Y, Inoue Y, Tanaka R, et al. PKM1 Confers Metabolic Advantages and Promotes Cell-Autonomous Tumor Cell Growth. Cancer Cell. 2018; 33(3): 355-67.e7.
- 14Huang C, Huang Z, Bai P, Luo G, Zhao X, Wang X. Expression of pyruvate kinase M2 in human bladder cancer and its correlation with clinical parameters and prognosis. Onco Targets Therapy. 2018; 11: 2075-82.
- 15Wang Y, Hao F, Nan Y, Qu L, Na W, Jia C, Chen X. PKM2 Inhibitor Shikonin Overcomes the Cisplatin Resistance in Bladder Cancer by Inducing Necroptosis. Int J Biol Sci.. 2018; 14(13): 1883-91.
- 16Wang X, Zhang F, Wu XR. Inhibition of Pyruvate Kinase M2 Markedly Reduces Chemoresistance of Advanced Bladder Cancer to Cisplatin. Sci Rep. 2017; 7:45983.
- 17Zhu Q, Hong B, Zhang L, Wang J. Pyruvate kinase M2 inhibits the progression of bladder cancer by targeting MAKP pathway. J Cancer Res Ther. 2018; 14(Supplement): S616-s21.
- 18Yang X, Cheng Y, Li P, Tao J, Deng X, Zhang X, et al. A lentiviral sponge for miRNA-21 diminishes aerobic glycolysis in bladder cancer T24 cells via the PTEN/PI3K/AKT/mTOR axis. Tumour Biol. 2015; 36(1): 383-91.
10.1007/s13277-014-2617-2 Google Scholar
- 19Boriack-Sjodin PA, Ribich S, Copeland RA. RNA-modifying proteins as anticancer drug targets. Nat Rev Drug Discov. 2018; 17(6): 435-53.
- 20Luo W, Hu H, Chang R, Zhong J, Knabel M, O'Meally R, et al. Pyruvate kinase M2 is a PHD3-stimulated coactivator for hypoxia-inducible factor 1. Cell. 2011; 145(5): 732-44.
- 21Kleszcz R, Paluszczak J, Krajka-Kuźniak V, Baer-Dubowska W. The inhibition of c-MYC transcription factor modulates the expression of glycolytic and glutaminolytic enzymes in FaDu hypopharyngeal carcinoma cells. Adv Clin Exp Med. 2018; 27(6): 735-42.
- 22Yang W, Xia Y, Ji H, Zheng Y, Liang J, Huang W, et al. Nuclear PKM2 regulates β-catenin transactivation upon EGFR activation. Nature. 2011; 480(7375): 118-22.
- 23Xu Q, Liu LZ, Yin Y, He J, Li Q, Qian X,et al. Regulatory circuit of PKM2/NF-κB/miR-148a/152-modulated tumor angiogenesis and cancer progression. Oncogene. 2015; 34(43): 5482-93.
10.1038/onc.2015.6 Google Scholar
- 24Guo M, Zhao X, Yuan X, Jiang J, Li P. MiR-let-7a inhibits cell proliferation, migration, and invasion by down-regulating PKM2 in cervical cancer. Oncotarget. 2017; 8(17): 28226-36.
- 25Hu CY, Chen J, Qin XH, You P, Ma J, Zhang J, et al.: Long non-coding RNA NORAD promotes the prostate cancer cell extracellular vesicle release via microRNA-541-3p-regulated PKM2 to induce bone metastasis of prostate cancer. J Exp Clin Cancer Res. 2021; 40(1): 98.
- 26Zahra K, Dey T, Ashish, Mishra SP, Pandey U. Pyruvate Kinase M2 and Cancer: The Role of PKM2 in Promoting Tumorigenesis. Front Oncol. 2020; 10: 159.
- 27Chen X, Li A, Sun BF, Yang Y, Han YN, Yuan X, et al.: 5-methylcytosine promotes pathogenesis of bladder cancer through stabilizing mRNAs. Nat Cell Biol. 2019; 21(8): 978-90.
- 28Cheng M, Sheng L, Gao Q, Xiong Q, Zhang H, Wu M, et al. The m(6)A methyltransferase METTL3 promotes bladder cancer progression via AFF4/NF-κB/MYC signaling network. Oncogene. 2019; 38(19): 3667-80.
- 29Han J, Wang JZ, Yang X, Yu H, Zhou R, Lu HC, et al. METTL3 promote tumor proliferation of bladder cancer by accelerating pri-miR221/222 maturation in m6A-dependent manner. Mol Cancer. 2019; 18(1): 110.
- 30McCloskey A, Taniguchi I, Shinmyozu K, Ohno M. hnRNP C tetramer measures RNA length to classify RNA polymerase II transcripts for export. Science. 2012; 335(6076): 1643-6.
- 31Meyer KD, Jaffrey SR. The dynamic epitranscriptome: N6-methyladenosine and gene expression control. Nat Rev Mol Cell Biol. 2014; 15(5): 313-26.
- 32Boccaletto P, Machnicka MA, Purta E, Piatkowski P, Baginski B, Wirecki TK, et al. MODOMICS: a database of RNA modification pathways. 2017 update. Nucleic Acids Res. 2018; 46(D1): D303-d07.
- 33Yu H, Yang X, Tang J, Si S, Zhou Z, Lu J, et al. ALKBH5 Inhibited Cell Proliferation and Sensitized Bladder Cancer Cells to Cisplatin by m6A-CK2α-Mediated Glycolysis. Molecular Therapy Nucleic Acids. 2021; 23: 27-41.
10.1016/j.omtn.2020.10.031 Google Scholar
- 34Wang Q, Chen C, Ding Q, Zhao Y, Wang Z, Chen J, et al. METTL3-mediated m(6)A modification of HDGF mRNA promotes gastric cancer progression and has prognostic significance. Gut. 2020; 69(7): 1193-205.
- 35Shen C, Xuan B, Yan T, Ma Y, Xu P, Tian X, et al. m(6)A-dependent glycolysis enhances colorectal cancer progression. Mol Cancer. 2020; 19(1): 72.
- 36Schaefer M, Pollex T, Hanna K, Lyko F. RNA cytosine methylation analysis by bisulfite sequencing. Nucleic Acids Res. 2009; 37(2):e12.
- 37Agris PF. Bringing order to translation: the contributions of transfer RNA anticodon-domain modifications. EMBO Rep. 2008; 9(7): 629-35.
- 38Helm M. Post-transcriptional nucleotide modification and alternative folding of RNA. Nucleic Acids Res. 2006; 34(2): 721-33.
- 39Squires JE, Patel HR, Nousch M, Sibbritt T, Humphreys DT, Parker BJ, et al.: Widespread occurrence of 5-methylcytosine in human coding and non-coding RNA. Nucleic Acids Res. 2012; 40(11): 5023-33.
- 40Huang T, Chen W, Liu J, Gu N, Zhang R. Genome-wide identification of mRNA 5-methylcytosine in mammals. Nat Struct Mol Biol. 2019; 26(5): 380-88.
- 41Yang X, Yang Y, Sun BF, Chen YS, Xu JW, Lai WY, et al. 5-methylcytosine promotes mRNA export - NSUN2 as the methyltransferase and ALYREF as an m(5)C reader. Cell Res. 2017; 27(5): 606-25.
- 42Denko NC. Hypoxia, HIF1 and glucose metabolism in the solid tumour. Nat Rev Cancer. 2008; 8(9): 705-13.
- 43Semenza GL. HIF-1 mediates the Warburg effect in clear cell renal carcinoma. J Bioenerg Biomembr. 2007; 39(3): 231-4.
- 44Robey IF, Lien AD, Welsh SJ, Baggett BK, Gillies RJ. Hypoxia-inducible factor-1alpha and the glycolytic phenotype in tumors. Neoplasia (New York, NY). 2005; 7(4): 324-30.
10.1593/neo.04430 Google Scholar
- 45Lu H, Forbes RA, Verma A. Hypoxia-inducible factor 1 activation by aerobic glycolysis implicates the Warburg effect in carcinogenesis. J Biol Chem. 2002; 277(26): 23111-5.
- 46Yang C, Yuan W, Yang X, Li P, Wang J, Han J, et al. Circular RNA circ-ITCH inhibits bladder cancer progression by sponging miR-17/miR-224 and regulating p21, PTEN expression. Mol Cancer. 2018; 17(1): 19.
- 47Messeguer X, Escudero R, Farre D, Nunez O, Martinez J, Alba MM. PROMO: detection of known transcription regulatory elements using species-tailored searches. Bioinformatics. 2002; 18(2): 333-4.
- 48Khan A, Fornes O, Stigliani A, Gheorghe M, Castro-Mondragon JA, van der Lee R, et al. JASPAR 2018: update of the open-access database of transcription factor binding profiles and its web framework. Nucleic Acids Res. 2018; 46(D1): D260-d66.
- 49Tao T, Su Q, Xu S, Deng J, Zhou S, Zhuang Y, et al. Down-regulation of PKM2 decreases FASN expression in bladder cancer cells through AKT/mTOR/SREBP-1c axis. J Cell Physiol. 2019; 234(3): 3088-104.
- 50Liu Z, Zhang J. Human C-to-U Coding RNA Editing Is Largely Nonadaptive. Mol Biol Evol. 2018; 35(4): 963-69.
- 51Mei L, Shen C, Miao R, Wang JZ, Cao MD, Zhang YS, et al.: RNA methyltransferase NSUN2 promotes gastric cancer cell proliferation by repressing p57(Kip2) by an m(5)C-dependent manner. Cell Death Dis. 2020; 11(4): 270.
- 52Cheng Y, Yang X, Deng X, Zhang X, Li P, Tao J, et al.: Metabolomics in bladder cancer: a systematic review. Int J Clin Exp Med. 2015; 8(7): 11052-63.
- 53Dominguez-Sanchez MS, Saez C, Japon MA, Aguilera A, Luna R. Differential expression of THOC1 and ALY mRNP biogenesis/export factors in human cancers. BMC Cancer. 2011; 11: 77.
- 54Saito Y, Kasamatsu A, Yamamoto A, Shimizu T, Yokoe H, Sakamoto Y, et al.: ALY as a potential contributor to metastasis in human oral squamous cell carcinoma. J Cancer Res Clin Oncol. 2013; 139(4): 585-94.
- 55Dayton TL, Jacks T. Vander Heiden MG: PKM2, cancer metabolism, and the road ahead. EMBO Rep. 2016; 17(12): 1721-30.
- 56Mazurek S. Pyruvate kinase type M2: A key regulator of the metabolic budget system in tumor cells. Int J Biochem Cell Biol. 2011; 43(7): 969-80.
- 57Palsson-McDermott EM, Curtis AM, Goel G, Lauterbach MA, Sheedy FJ, Gleeson LE, et al. Pyruvate kinase M2 regulates Hif-1α activity and IL-1β induction and is a critical determinant of the warburg effect in LPS-activated macrophages. Cell Metab. 2015; 21(1): 65-80.
