ERN1 inhibition modifies hypoxic regulation of EGLN1 expression in U87MG glioblastoma cells

Oleh V. Halkin; Dmytro O. Minchenko; Yuliia M. Viletska; Myroslava Y. Sliusar; Olena O. Khita; Taia V. Feldman; Oleksandr H. Minchenko

doi:10.2478/enr-2026-0018

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ERN1 inhibition modifies hypoxic regulation of EGLN1 expression in U87MG glioblastoma cells

Endocrine Regulations

Volume 60 (2026): Issue 1 (January 2026)

By: Oleh V. Halkin, Dmytro O. Minchenko, Yuliia M. Viletska, Myroslava Y. Sliusar, Olena O. Khita, Taia V. Feldman and Oleksandr H. Minchenko

Open Access

|Aug 2026

Abstract

Objective. Glioblastomas are the most common and highly aggressive malignant brain tumors that are difficult to treat. Hypoxia is a significant factor in glioblastoma growth. Previous studies have shown that the inhibition of ERN1 (endoplasmic reticulum to nucleus signaling 1) significantly suppresses the glioblastoma cell proliferation and modifies the hypoxic regulation. The present study aimed to investigate the impact of hypoxia on the expression of endoplasmic reticulum stress-dependent genes including EGLN1 in U87MG glioblastoma cells in relation to ERN1 inhibition to reveal a possible role of the ERN1 signaling pathway in the hypoxic regulation of these genes’ expression.

Methods. The control U87MG glioblastoma cells (transfected by an empty vector) and ERN1 knockdown cells with inhibited ERN1 endoribonuclease and protein kinase (dnERN1) were used. Hypoxia was induced at normoxic conditions with dimethyloxalylglycine (0.5 mM for 4 h). RNA was extracted and reverse transcribed. The expression levels of EGLN1, HSPA5, DDIT4, PA2G4, XIAP, and MTOR genes were studied by real-time qPCR and normalized to ACTB mRNA.

Results. It was established that hypoxia increases the expression level of EGLN1, HSPA5, DDIT4, and XIAP in glioblastoma cells with native ERN1. However, the expression levels of two other stress-dependent genes (PA2G4 and MTOR) were reduced in these glioblastoma cells under hypoxia. Furthermore, inhibition of ERN1 enzymatic activity attenuated the effects of hypoxia on the expression of most of the genes studied in glioblastoma cells, while XIAP gene expression was dramatically increased.

Conclusion. The results of this study showed that hypoxia differentially affects the expression of genes related to endoplasmic reticulum stress in glioblastoma cells and is dependent on ERN1 activity. This reflected the ERN1-mediated reprogramming of hypoxic regulation of gene expression.

References

Acosta-Alvear D, Harnoss JM, Walter P, Ashkenazi A. Homeostasis control in health and disease by the unfolded protein response. Nat Rev Mol Cell Biol 26, 193–212, 2025.
Search in Google Scholar Back to article
Almanza A, Carlesso A, Chintha C, Creedican S, Doultsinos D, Leuzzi B, Luis A, McCarthy N, Montibeller L, More S, Papaioannou A, Puschel F, Sassano ML, Skoko J, Agostinis P, de Belleroche J, Eriksson LA, Fulda S, Gorman AM, Healy S, Kozlov A, Munoz-Pinedo C, Rehm M, Chevet E, Samali A. Endoplasmic reticulum stress signalling - from basic mechanisms to clinical applications. FEBS J 286, 241–278, 2019.
Search in Google Scholar Back to article
Auf G, Jabouille A, Guerit S, Pineau R, Delugin M, Bouchecareilh M, Favereaux A, Maitre M, Gaiser T, von Deimling A, Czabanka M, Vajkoczy P, Chevet E, Bikfalvi A, Moenner M. A shift from an angiogenic to invasive phenotype induced in malignant glioma by inhibition of the unfolded protein response sensor IRE1. Proc Natl Acad Sci USA 107, 15553–15558, 2010.
Search in Google Scholar Back to article
Auf G, Jabouille A, Delugin M, Guerit S, Pineau R, North S, Platonova N, Maitre M, Favereaux A, Vajkoczy P, Seno M, Bikfalvi A, Minchenko D, Minchenko O, Moenner M. High epiregulin expression in human U87 glioma cells relies on IRE1alpha and promotes autocrine growth through EGF receptor. BMC Cancer 13, 597, 2013.
Search in Google Scholar Back to article
Bobarykina AY, Minchenko DO, Opentanova IL, Moenner M, Caro J, Esumi H, Minchenko OH. Hypoxic regulation of PFKFB-3 and PFKFB-4 gene expression in gastric and pancreatic cancer cell lines and expression of PFKFB genes in gastric cancers. Acta Biochim Pol 53, 789–799, 2006.
Search in Google Scholar Back to article
Casillas AL, Chauhan SS, Toth RK, Sainz AG, Clements AN, Jensen CC, Langlais PR, Miranti CK, Cress AE, Warfel NA. Direct phosphorylation and stabilization of HIF-1alpha by PIM1 kinase drives angiogenesis in solid tumors. Oncogene 40, 5142–5152, 2021.
Search in Google Scholar Back to article
Chan MC, Ilott NE, Schodel J, Sims D, Tumber A, Lippl K, Mole DR, Pugh CW, Ratcliffe PJ, Ponting CP, Schofield CJ. Tuning the transcriptional response to hypoxia by inhibiting hypoxia-inducible factor (HIF) prolyl and asparaginyl hydroxylases. J Biol Chem 291, 20661–20673, 2016.
Search in Google Scholar Back to article
Chen X, Iliopoulos D, Zhang Q, Tang Q, Greenblatt MB, Hatziapostolou M, Lim E, Tam WL, Ni M, Chen Y, Mai J, Shen H, Hu DZ, Adoro S, Hu B, Song M, Tan C, Landis MD, Ferrari M, Shin SJ, Brown M, Chang JC, Liu XS, Glimcher LH. XBP1 promotes triple-negative breast cancer by controlling the HIF-1α pathway. Nature 508, 103–107, 2014.
Search in Google Scholar Back to article
Chen Y, Shen H, Wang Z, Huang C, Zhang H, Shao Y, Tong Y, Xu L, Lu Y, Fu Z. Recruitment of USP10 by GCS1 to deubiquitinate GRP78 promotes the progression of colorectal cancer via alleviating endoplasmic reticulum stress. J Exp Clin Cancer Res 43, 261, 2024.
Search in Google Scholar Back to article
Chevet E, Hetz C, Samali A. Endoplasmic reticulum stress-activated cell reprogramming in oncogenesis. Cancer Discov 5, 586–597, 2015.
Search in Google Scholar Back to article
Dai B, Xu L, Rong S, Song M, Lan Z, Chen W, Zhang L, Liu Y, Wang L, Li J, Chen J, Wu Z. YTHDF2 promotes ana-plastic thyroid cancer progression by activating the DDIT4/AKT/mTOR signaling pathway. Biol Direct 19, 122, 2024.
Search in Google Scholar Back to article
Danilovskyi SV, Minchenko DO, Karbovskyi LL, Moliavko OS, Kovalevska OV, Minchenko OH. ERN1 knockdown modifies the hypoxic regulation of TP53, MDM2, USP7 and PERP gene expressions in U87 glioma cells. Ukr Biochem J 86, 90–102, 2014.
Search in Google Scholar Back to article
Denko NC. Hypoxia, HIF1 and glucose metabolism in the solid tumor. Nat Rev Cancer 8, 705–713, 2008.
Search in Google Scholar Back to article
Doultsinos D, Avril T, Lhomond S, Dejeans N, Guedat P, Chevet E. Control of the unfolded protein response in health and disease. SLAS Discov 22, 787–800, 2017.
Search in Google Scholar Back to article
Fels DR, Koumenis C. The PERK/eIF2a/ATF4 module of the UPR in hypoxia resistance and tumor growth. Cancer Biol Ther 5, 723–728, 2006.
Search in Google Scholar Back to article
He SW, Liang YL, Zhang Y, Liu X, Gong S, Ye ML, Huang SY, Tan XR, Zhou SQ, Zhao Y, Liu N, Li YQ. LINC00173 facilitates tumor progression by stimulating RAB1B-mediated PA2G4 and SDF4 secretion in nasopharyngeal carcinoma. Mol Oncol 17, 518–533, 2023.
Search in Google Scholar Back to article
Hetz C, Zhang K, Kaufman RJ. Mechanisms, regulation and functions of the unfolded protein response. Nat Rev Mol Cell Biol 21, 421–438, 2020.
Search in Google Scholar Back to article
Hiramatsu N, Messah C, Han J, Lavail MM, Kaufman RJ, Lin JH. Translational and post-translational regulation of XIAP by eIF2α and ATF4 promotes ER stress-induced cell death during the unfolded protein response. Mol Biol Cell 25, 1411–1420, 2014.
Search in Google Scholar Back to article
Huang X, Wang XN, Yuan XD, Wu WY, Lobie PE, Wu Z. XIAP facilitates breast and colon carcinoma growth via promotion of p62 depletion through ubiquitination-dependent proteasomal degradation. Oncogene 38, 1448–1460, 2019.
Search in Google Scholar Back to article
Jiang W, Zhang M, Gao C, Yan C, Gao R, He Z, Wei X, Xiong J, Ruan Z, Yang Q, Li J, Li Q, Zhong Z, Zhang M, Yuan Q, Hu H, Wang S, Hu MM, Cai C, Wu GS, Jiang C, Zhang YL, Zhang CS, Zhang J. A mitochondrial EglN1-AMPKalpha axis drives breast cancer progression by enhancing metabolic adaptation to hypoxic stress. EMBO J 42, e113743, 2023.
Search in Google Scholar Back to article
Knaup KX, Monti J, Hackenbeck T, Jobst-Schwan T, Klanke B, Schietke RE, Wacker I, Behrens J, Amann K, Eckardt KU, Warnecke C, Wiesener MS. Hypoxia regulates the sperm associated antigen 4 (SPAG4) via HIF, which is expressed in renal clear cell carcinoma and promotes migration and invasion in vitro. Mol Carcinog 53, 970–978, 2014.
Search in Google Scholar Back to article
Krasnytska DA, Khita OO, Viletska YM Minchenko DO, Halkin OV, Rudnytska OV, Hoian SL, Minchenko OH. ERN1 knockdown modifies the hypoxic regulation of homeobox gene expression in U87MG glioblastoma cells. Endocr Regul 58, 47–56, 2024.
Search in Google Scholar Back to article
Kumar S, Shuaib M, AlAsmari AF, Alqahtani F, Gupta S. GNL3 and PA2G4 as prognostic biomarkers in prostate cancer. Cancers (Basel) 15, 2723, 2023.
Search in Google Scholar Back to article
Lin X, Yoshikawa N, Liu W, Matsukawa T, Nakamura K, Yoshihara M, Koya Y, Sugiyama M, Tamauchi S, Ikeda Y, Yokoi A, Shimizu Y, Kajiyama H. DDIT4 facilitates lymph node metastasis via the activation of NF-kappaB pathway and epithelial-mesenchymal transition. Reprod Sci 30, 2829–2841, 2023.
Search in Google Scholar Back to article
Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2⁻ΔΔCt method. Methods 25, 402–408, 2001.
Search in Google Scholar Back to article
Liu Z, Liu G, Ha DP, Wang J, Xiong M, Lee AS. ER chaperone GRP78/BiP translocates to the nucleus under stress and acts as a transcriptional regulator. Proc Natl Acad Sci USA 120, e2303448120, 2023.
Search in Google Scholar Back to article
Logue SE, McGrath EP, Cleary P, Greene S, Mnich K, Almanza A, Chevet E, Dwyer RM, Oommen A, Legembre P, Godey F, Madden EC, Leuzzi B, Obacz J, Zeng Q, Patterson JB, Jager R, Gorman AM, Samali A. Inhibition of IRE1 RNase activity modulates the tumor cell secretome and enhances response to chemotherapy. Nat Commun 9, 3267, 2018.
Search in Google Scholar Back to article
Losman JA, Koivunen P, Kaelin WG Jr. 2-Oxoglutarate-dependent dioxygenases in cancer. Nat Rev Cancer 20, 710–726, 2020.
Search in Google Scholar Back to article
Marafie SK, Al-Mulla F, Abubaker J. mTOR: its critical role in metabolic diseases, cancer, and the aging process. Int J Mol Sci 25, 6141, 2024.
Search in Google Scholar Back to article
Minchenko A, Caro J. Regulation of endothelin-1 gene expression in human microvascular endothelial cells by hypoxia and cobalt: role of hypoxia responsible element. Mol Cell Biochem 208, 53–62, 2000.
Search in Google Scholar Back to article
Minchenko A, Bauer T, Salceda S, Caro J. Hypoxic stimulation of vascular endothelial growth factor expression in vitro and in vivo. Lab Invest 71, 374–379, 1994.
Search in Google Scholar Back to article
Minchenko A, Leshchinsky I, Opentanova I, Sang N, Srinivas V, Armstead V, Caro J. Hypoxia-inducible factor-1-mediated expression of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) gene. Its possible role in the Warburg effect. J Biol Chem 277, 6183–6187, 2002.
Search in Google Scholar Back to article
Minchenko O, Opentanova I, Caro J. Hypoxic regulation of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene family (PFKFB-1-4) expression in vivo. FEBS Lett 554, 264–270, 2003.
Search in Google Scholar Back to article
Minchenko O, Opentanova I, Minchenko D, Ogura T, Esumi H. Hypoxia induces transcription of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 4 gene via hypoxia-inducible factor-1alpha activation. FEBS Lett 576, 14–20, 2004.
Search in Google Scholar Back to article
Minchenko OH, Ogura T, Opentanova IL, Minchenko DO, Esumi H. Splice isoform of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-4: expression and hypoxic regulation. Mol Cell Biochem 280, 227–234, 2005.
Search in Google Scholar Back to article
Minchenko DO, Kharkova AP, Hubenia OV, Minchenko OH. Insulin receptor, IRS1, IRS2, INSIG1, INSIG2, RRAD, and BAIAP2 gene expressions in glioma U87 cells with ERN1 loss of function: effect of hypoxia and glutamine or glucose deprivation. Endocr Regul 47, 15–26, 2013.
Search in Google Scholar Back to article
Minchenko OH, Tsuchihara K, Minchenko DO, Bikfalvi A, Esumi H. Mechanisms of regulation of PFKFB expression in pancreatic and gastric cancer cells. World J Gastroenterol 20, 13705–13717, 2014. Minchenko DO, Tsymbal DO, Riabovol OO, Viletska YM, Lahanovska YO, Sliusar MY, Bezrodnyi BH,
Search in Google Scholar Back to article
Minchenko OH. Hypoxic regulation of EDN1, EDNRA, EDNRB, and ECE1 gene expressions in IRE1 knockdown U87 glioma cells. Endocr Regul 53, 250–262, 2019.
Search in Google Scholar Back to article
Minchenko OH, Tsymbal DO, Khita OO, Minchenko DO. Inhibition of ERN1 signaling is important for the suppression of tumor growth. Clin Cancer Drugs 8, 27–38, 2021.
Search in Google Scholar Back to article
Minchenko OH, Khita OO, Krasnytska DA, Viletska YM, Rudnytska OV, Hnatiuk OS, Minchenko DO. ERN1 knockdown affects the expression of TGIF1 and other homeobox genes in U87MG glioblastoma cells. Arch Biochem Biophys 758, 110073, 2024a.
Search in Google Scholar Back to article
Minchenko OH, Sliusar MY, Khikhlo YP, Halkin OV, Viletska YM, Khita OO, Minchenko DO. Knockdown of ERN1 disturbs the expression of phosphoserine aminotransferase 1 and related genes in glioblastoma cells. Arch Biochem Biophys 759, 110104, 2024b.
Search in Google Scholar Back to article
Minchenko OH, Sliusar MY, Khita OO, Viletska YM, Luzina OY, Danilovskyi SV, Minchenko DO. Endoplasmic reticulum stress-dependent regulation of the expression of serine hydroxymethyltransferase 2 in glioblastoma cells. Endocr Regul 58, 144–152, 2024c.
Search in Google Scholar Back to article
Minchenko OH, Hrebennykova VO, Viletska YM, Hnatiuk OS, Sliusar MY, Kozynkevych HE, Minchenko DO. The ERN1 signaling pathway of unfolded protein response controls the expression of EDEM1 and their hypoxic regulation in glioblastoma cells. Endocr Regul 59, 1–9, 2025.
Search in Google Scholar Back to article
Mossmann D, Park S, Hall MN. mTOR signalling and cellular metabolism are mutual determinants in cancer. Nat Rev Cancer 18, 744–757, 2018.
Search in Google Scholar Back to article
Murugan AK. mTOR: Role in cancer, metastasis and drug resistance. Semin Cancer Biol 59, 92–111, 2019.
Search in Google Scholar Back to article
Papaioannou A, Chevet E. Driving cancer tumorigenesis and metastasis through UPR signaling. Curr Top Microbiol Immunol 414, 159–192, 2018.
Search in Google Scholar Back to article
Pei M, Zhang J, Yu Z, Peng Y, Chen Y, Peng S, Wei X, Wu J, Huang X, Xie Y, Yang P, Hong L, Huang X, Wu X, Tang W, Chen Y, Liu S, Lin J, Xiang L, Wang J. LINC02139 interacts with and stabilizes XIAP to regulate cell proliferation and apoptosis in gastric cancer. Commun Biol 7, 1497, 2024.
Search in Google Scholar Back to article
Pelizzari-Raymundo D, Doultsinos D, Pineau R, Sauzay C, Koutsandreas T, Langlais T, Carlesso A, Gkotsi E, Negroni L, Avril T, Chatziioannou A, Chevet E, Eriksson LA, Guillory X. A novel IRE1 kinase inhibitor for adjuvant glioblastoma treatment. iScience 26, 106687, 2023.
Search in Google Scholar Back to article
Rudnytska OV, Khita OO, Minchenko DO, Tsymbal DO, Yefimova YV, Sliusar MY, Minchenko OH. The low doses of SWCNTs exhibit a genotoxic effect on the normal human astrocytes by disrupting the functional integrity of the genome. Curr Res Toxicol 2, 64–71, 2021.
Search in Google Scholar Back to article
Schodel J, Grampp S, Maher ER, Moch H, Ratcliffe PJ, Russo P, Mole DR. Hypoxia, hypoxia-inducible transcription factors, and renal cancer. Eur Urol 69, 646–657, 2016.
Search in Google Scholar Back to article
Schofield CJ, Ratcliffe PJ. Oxygen sensing by HIF hydroxylases. Nat Rev Mol Cell Biol 5, 343–354, 2004.
Search in Google Scholar Back to article
Semenza GL. A compendium of proteins that interact with HIF-1α. Exp Cell Res 356, 128–135, 2017.
Search in Google Scholar Back to article
Sliusar MY, Minchenko DO, Khita OO, Tsymbal DO, Viletska YM, Luzina OY, Danilovskyi SV, Ratushna OO, Minchenko OH. Hypoxia controls the expression of genes responsible for serine synthesis in U87MG cells on ERN1-dependent manner. Endocr Regul 57, 252–261, 2023.
Search in Google Scholar Back to article
Tajik F, Fattahi F, Rezagholizadeh F, Bouzari B, Babaheidarian P, Baghai Wadji M, Madjd Z. Nuclear overexpression of DNA damage-inducible transcript 4 (DDIT4) is associated with aggressive tumor behavior in patients with pancreatic tumors. Sci Rep 13, 19403, 2023.
Search in Google Scholar Back to article
Viletska YM, Minchenko OH, Khita OO, Tsymbal DO, Sliusar MY, Halkin OV, Kozynkevych HE, Minchenko DO. ERN1-dependent regulation of BAG cochaperone 1 expression and the sensitivity to glutamine deprivation in U87MG glioblastoma cells. Endocr Regul 60, 37–47, 2026a.
Search in Google Scholar Back to article
Viletska YM, Minchenko OH, Khita OO, Sliusar MY, Halkin OV, Kozynkevych HE, Minchenko DO. Inhibition of ERN1 suppresses the expression of phosphoenolpyruvate carboxykinase 2 in U87MG glioblastoma cells and increases its sensitivity to glucose and glutamine deprivation. Endocr Regul 60, 61–71, 2026b.
Search in Google Scholar Back to article
Wang GL, Semenza GL. General involvement of hypoxia-inducible factor 1 in transcriptional response to hypoxia. Proc Natl Acad Sci U S A 90, 4304–4308, 1993.
Search in Google Scholar Back to article
Wenger RH. Cellular adaptation to hypoxia: O2-sensing protein hydroxylases, hypoxia-inducible transcription factors, and O2-regulated gene expression. FASEB J 16,1151–1162, 2002.
Search in Google Scholar Back to article
Yin Y, Chen C, Chen J, Zhan R, Zhang Q, Xu X, Li D, Li M. Cell surface GRP78 facilitates hepatoma cells proliferation and migration by activating IGF-IR. Cell Signal 35, 154–162, 2017.
Search in Google Scholar Back to article

Articles in this issue

DOI: https://doi.org/10.2478/enr-2026-0018 | Journal eISSN: 1336-0329 | Journal ISSN: 1210-0668

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Language: English

Page range: 179 - 189

Published on: Aug 31, 2026

Published by: Slovak Academy of Sciences, Institute of Experimental Endocrinology

In partnership with: Paradigm Publishing Services

Keywords:

EGLN1,

DDIT4,

HJSPA5,

mTOR,

hypoxia,

Related subjects:

Basic medical science,

Basic medical science, other,

Clinical medicine,

Internal medicine,

Endocrinology, diabetology

© 2026 Oleh V. Halkin, Dmytro O. Minchenko, Yuliia M. Viletska, Myroslava Y. Sliusar, Olena O. Khita, Taia V. Feldman, Oleksandr H. Minchenko, published by Slovak Academy of Sciences, Institute of Experimental Endocrinology
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 License.

Volume 60 (2026): Issue 1 (January 2026)