Activating transcription factor 4 regulates hypoxia inducible factor 1α in chronic hypoxia in pancreatic cancer cells

Chee,Nancy T.; Carriere,Candace H.; Miller,Zachary; Welford,Scott; Brothers,Shaun P.

doi:10.3892/or.2022.8451

Activating transcription factor 4 regulates hypoxia inducible factor 1α in chronic hypoxia in pancreatic cancer cells

Authors:
- Nancy T. Chee
- Candace H. Carriere
- Zachary Miller
- Scott Welford
- Shaun P. Brothers
View Affiliations

Affiliations: Center for Therapeutic Innovation, Miller School of Medicine, University of Miami, Miami, FL 33136, USA, Department of Radiation Oncology, Molecular Therapeutics Shared Resource, Sylvester Comprehensive Cancer Center, University of Miami, Miami, FL 33136, USA
Published online on: November 23, 2022 https://doi.org/10.3892/or.2022.8451
Article Number: 14
Copyright: © Chee et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and difficult to treat cancers with tumors typically exhibiting high levels of chronic hypoxia. Hypoxia activates hypoxia-inducible factors (HIFs) that mediate cellular responses to adapt to low oxygen environments. Hypoxia also causes endoplasmic reticulum (ER) stress, increasing activating transcription factor 4 (ATF4), a master regulator of the unfolded protein response (UPR) pathway that mediates cellular response to ER stress. ATF4 is overexpressed in PDAC and is associated with poor prognoses. While ATF4 promotes cell proliferation and tumorigenesis, most studies have been conducted under normoxia or acute hypoxia. The functions of ATF4 in chronic hypoxia remain largely unexplored. Using siRNA knockdown experiments of healthy skin fibroblast cells WS1 and PDAC cell lines PANC-1 and Mia-PaCa2 to analyze mRNA and protein expression levels, a novel ATF4 function was identified, in which it decreases HIF2α mRNA and increases HIF1α mRNA in chronic hypoxia while having no effect in acute hypoxia. A scratch assay was used to show that ATF4 decreases cell migration in chronic hypoxia as opposed to the increase in cell migration ATF4 imparts in acute hypoxia. Colony formation assay and cell viability assay showed that ATF4 promotes colony formation and cell viability in both chronic and acute hypoxia. In addition to the differential response of ATF4 in chronic hypoxia compared with acute hypoxia, this is the first time ATF4 has been implicated in regulation of response to hypoxia via interaction with HIF proteins in PDAC.

View Figures

View References

1	Siegel RL, Miller KD and Jemal A: Cancer statistics, 2018. CA Cancer J Clin. 68:7–30. 2018. View Article : Google Scholar : PubMed/NCBI
2	Hessmann E, Buchholz SM, Demir IE, Singh SK, Gress TM, Ellenrieder V and Neesse A: Microenvironmental determinants of pancreatic cancer. Physiol Rev. 100:1707–1751. 2020. View Article : Google Scholar : PubMed/NCBI
3	Orth M, Metzger P, Gerum S, Mayerle J, Schneider G, Belka C, Schnurr M and Lauber K: Pancreatic ductal adenocarcinoma: Biological hallmarks, current status, and future perspectives of combined modality treatment approaches. Radiat Oncol. 14:1412019. View Article : Google Scholar : PubMed/NCBI
4	Yuen A and Díaz B: The impact of hypoxia in pancreatic cancer invasion and metastasis. Hypoxia (Auckl). 2:91–106. 2014.PubMed/NCBI
5	Buttgereit F and Brand MD: A hierarchy of ATP-consuming processes in mammalian cells. Biochem J. 312:163–167. 1995. View Article : Google Scholar : PubMed/NCBI
6	Fähling M: Surviving hypoxia by modulation of mRNA translation rate. J Cell Mol Med. 13:2770–2779. 2009. View Article : Google Scholar : PubMed/NCBI
7	Wortel IMN, van der Meer LT, Kilberg MS and van Leeuwen FN: Surviving stress: Modulation of ATF4-mediated stress responses in normal and malignant cells. Trends Endocrinol Metab. 28:794–806. 2017. View Article : Google Scholar : PubMed/NCBI
8	B'chir W, Maurin AC, Carraro V, Averous J, Jousse C, Muranishi Y, Parry L, Stepien G, Fafournoux P and Bruhat A: The eIF2α/ATF4 pathway is essential for stress-induced autophagy gene expression. Nucleic Acids Res. 41:7683–7699. 2013. View Article : Google Scholar : PubMed/NCBI
9	Guo X, Aviles G, Liu Y, Tian R, Unger BA, Lin YT, Wiita AP, Xu K, Correia MA and Kampmann M: Mitochondrial stress is relayed to the cytosol by an OMA1-DELE1-HRI pathway. Nature. 579:427–432. 2020. View Article : Google Scholar : PubMed/NCBI
10	Pathria G, Scott DA, Feng Y, Sang Lee J, Fujita Y, Zhang G, Sahu AD, Ruppin E, Herlyn M, Osterman AL and Ronai ZA: Targeting the Warburg effect via LDHA inhibition engages ATF4 signaling for cancer cell survival. EMBO J. 37:e997352018. View Article : Google Scholar : PubMed/NCBI
11	Fernandez MR and Cleveland JL: ATF4-amino acid circuits: A recipe for resistance in melanoma. EMBO J. 37:e1006002018. View Article : Google Scholar : PubMed/NCBI
12	Chiou SH, Risca VI, Wang GX, Yang D, Grüner BM, Kathiria AS, Ma RK, Vaka D, Chu P, Kozak M, et al: BLIMP1 induces transient metastatic heterogeneity in pancreatic cancer. Cancer Discov. 7:1184–1199. 2017. View Article : Google Scholar : PubMed/NCBI
13	Mesclon F, Lambert-Langlais S, Carraro V, Parry L, Hainault I, Jousse C, Maurin AC, Bruhat A, Fafournoux P and Averous J: Decreased ATF4 expression as a mechanism of acquired resistance to long-term amino acid limitation in cancer cells. Oncotarget. 8:27440–27453. 2017. View Article : Google Scholar : PubMed/NCBI
14	Palam LR, Gore J, Craven KE, Wilson JL and Korc M: Integrated stress response is critical for gemcitabine resistance in pancreatic ductal adenocarcinoma. Cell Death Dis. 6:e19132015. View Article : Google Scholar : PubMed/NCBI
15	Ait Ghezala H, Jolles B, Salhi S, Castrillo K, Carpentier W, Cagnard N, Bruhat A, Fafournoux P and Jean-Jean O: Translation termination efficiency modulates ATF4 response by regulating ATF4 mRNA translation at 5′ short ORFs. Nucleic Acids Res. 40:9557–9570. 2012. View Article : Google Scholar : PubMed/NCBI
16	Blais JD, Filipenko V, Bi M, Harding HP, Ron D, Koumenis C, Wouters BG and Bell JC: Activating transcription factor 4 is translationally regulated by hypoxic stress. Mol Cell Biol. 24:7469–7482. 2004. View Article : Google Scholar : PubMed/NCBI
17	Chee NT, Lohse I and Brothers SP: mRNA-to-protein translation in hypoxia. Mol Cancer. 18:492019. View Article : Google Scholar : PubMed/NCBI
18	Logsdon DP, Shah F, Carta F, Supuran CT, Kamocka M, Jacobsen MH, Sandusky GE, Kelley MR and Fishel ML: Blocking HIF signaling via novel inhibitors of CA9 and APE1/Ref-1 dramatically affects pancreatic cancer cell survival. Sci Rep. 8:137592018. View Article : Google Scholar : PubMed/NCBI
19	Zhang S, Zhao L, Wang J, Chen N, Yan J and Pan X: HIF-2α and Oct4 have synergistic effects on survival and myocardial repair of very small embryonic-like mesenchymal stem cells in infarcted hearts. Cell Death Dis. 8:e25482017. View Article : Google Scholar : PubMed/NCBI
20	Uniacke J, Holterman CE, Lachance G, Franovic A, Jacob MD, Fabian MR, Payette J, Holcik M, Pause A and Lee S: An oxygen-regulated switch in the protein synthesis machinery. Nature. 486:126–129. 2012. View Article : Google Scholar : PubMed/NCBI
21	Ho JJD, Wang M, Audas TE, Kwon D, Carlsson SK, Timpano S, Evagelou SL, Brothers S, Gonzalgo ML, Krieger JR, et al: Systemic reprogramming of translation efficiencies on oxygen stimulus. Cell Rep. 14:1293–1300. 2016. View Article : Google Scholar : PubMed/NCBI
22	Uniacke J, Perera JK, Lachance G, Francisco CB and Lee S: Cancer cells exploit eIF4E2-directed synthesis of hypoxia response proteins to drive tumor progression. Cancer Res. 74:1379–1389. 2014. View Article : Google Scholar : PubMed/NCBI
23	Rzymski T, Milani M, Pike L, Buffa F, Mellor HR, Winchester L, Pires I, Hammond E, Ragoussis I and Harris AL: Regulation of autophagy by ATF4 in response to severe hypoxia. Oncogene. 29:4424–4435. 2010. View Article : Google Scholar : PubMed/NCBI
24	Koumenis C and Wouters BG: ‘Translating’ tumor hypoxia: Unfolded protein response (UPR)-dependent and UPR-independent pathways. Mol Cancer Res. 4:423–436. 2006. View Article : Google Scholar : PubMed/NCBI
25	Suarez-Arnedo A, Torres Figueroa F, Clavijo C, Arbeláez P, Cruz JC and Muñoz-Camargo C: An image J plugin for the high throughput image analysis of in vitro scratch wound healing assays. PLoS One. 15:e02325652020. View Article : Google Scholar : PubMed/NCBI
26	Wei L, Lin Q, Lu Y, Li G, Huang L, Fu Z, Chen R and Zhou Q: Cancer-associated fibroblasts-mediated ATF4 expression promotes malignancy and gemcitabine resistance in pancreatic cancer via the TGF-β1/SMAD2/3 pathway and ABCC1 transactivation. Cell Death Dis. 12:3342021. View Article : Google Scholar : PubMed/NCBI
27	Koong AC, Mehta VK, Le QT, Fisher GA, Terris DJ, Brown JM, Bastidas AJ and Vierra M: Pancreatic tumors show high levels of hypoxia. Int J Radiat Oncol Biol Phys. 48:919–922. 2000. View Article : Google Scholar : PubMed/NCBI
28	Graffman S, Björk P, Ederoth P and Ihse I: Polarographic pO2 measurements of intra-abdominal adenocarcinoma in connection with intraoperative radiotherapy before and after change of oxygen concentration of anaesthetic gases. Acta Oncol. 40:105–107. 2001. View Article : Google Scholar : PubMed/NCBI
29	McKeown SR: Defining normoxia, physoxia and hypoxia in tumours-implications for treatment response. Br J Radiol. 87:201306762014. View Article : Google Scholar : PubMed/NCBI
30	Saxena K and Jolly MK: Acute vs chronic vs cyclic hypoxia: Their differential dynamics, molecular mechanisms, and effects on tumor progression. Biomolecules. 9:3392019. View Article : Google Scholar : PubMed/NCBI
31	Koh MY, Lemos R Jr, Liu X and Powis G: The hypoxia-associated factor switches cells from HIF-1α- to HIF-2α-dependent signaling promoting stem cell characteristics, aggressive tumor growth and invasion. Cancer Res. 71:4015–4027. 2011. View Article : Google Scholar : PubMed/NCBI
32	Koh MY and Powis G: Passing the baton: The HIF switch. Trends Biochem Sci. 37:364–372. 2012. View Article : Google Scholar : PubMed/NCBI
33	Onnis B, Rapisarda A and Melillo G: Development of HIF-1 inhibitors for cancer therapy. J Cell Mol Med. 13:2780–2786. 2009. View Article : Google Scholar : PubMed/NCBI
34	Downes NL, Laham-Karam N, Kaikkonen MU and Ylä-Herttuala S: Differential but complementary HIF1α and HIF2α transcriptional regulation. Mol Ther. 26:1735–1745. 2018. View Article : Google Scholar : PubMed/NCBI
35	Holmquist-Mengelbier L, Fredlund E, Löfstedt T, Noguera R, Navarro S, Nilsson H, Pietras A, Vallon-Christersson J, Borg A, Gradin K, et al: Recruitment of HIF-1alpha and HIF-2alpha to common target genes is differentially regulated in neuroblastoma: HIF-2alpha promotes an aggressive phenotype. Cancer Cell. 10:413–423. 2006. View Article : Google Scholar : PubMed/NCBI
36	Koh MY, Darnay BG and Powis G: Hypoxia-associated factor, a novel E3-ubiquitin ligase, binds and ubiquitinates hypoxia-inducible factor 1alpha, leading to its oxygen-independent degradation. Mol Cell Biol. 28:7081–7095. 2008. View Article : Google Scholar : PubMed/NCBI
37	Serocki M, Bartoszewska S, Janaszak-Jasiecka A, Ochocka RJ, Collawn JF and Bartoszewski R: miRNAs regulate the HIF switch during hypoxia: A novel therapeutic target. Angiogenesis. 21:183–202. 2018. View Article : Google Scholar : PubMed/NCBI
38	Lu ZJ, Lu LG, Tao KZ, Chen DF, Xia Q, Weng JJ, Zhu F, Wang XP and Zheng P: MicroRNA-185 suppresses growth and invasion of colon cancer cells through inhibition of the hypoxia-inducible factor-2α pathway in vitro and in vivo. Mol Med Rep. 10:2401–2408. 2014. View Article : Google Scholar : PubMed/NCBI
39	Uchida T, Rossignol F, Matthay MA, Mounier R, Couette S, Clottes E and Clerici C: Prolonged hypoxia differentially regulates hypoxia-inducible factor (HIF)-1alpha and HIF-2alpha expression in lung epithelial cells: Implication of natural antisense HIF-1alpha. J Biol Chem. 279:14871–14878. 2004. View Article : Google Scholar : PubMed/NCBI
40	Nagelkerke A, Bussink J, Mujcic H, Wouters BG, Lehmann S, Sweep FC and Span PN: Hypoxia stimulates migration of breast cancer cells via the PERK/ATF4/LAMP3-arm of the unfolded protein response. Breast Cancer Res. 15:R22013. View Article : Google Scholar : PubMed/NCBI
41	Du J, Liu H, Mao X, Qin Y and Fan C: ATF4 promotes lung cancer cell proliferation and invasion partially through regulating Wnt/β-catenin signaling. Int J Med Sci. 18:1442–1448. 2021. View Article : Google Scholar : PubMed/NCBI
42	Dadey DYA, Kapoor V, Khudanyan A, Thotala D and Hallahan DE: PERK regulates glioblastoma sensitivity to ER stress although promoting radiation resistance. Mol Cancer Res. 16:1447–1453. 2018. View Article : Google Scholar : PubMed/NCBI
43	Zeng P, Sun S, Li R, Xiao ZX and Chen H: HER2 upregulates ATF4 to promote cell migration via activation of ZEB1 and downregulation of E-cadherin. Int J Mol Sci. 20:22232019. View Article : Google Scholar : PubMed/NCBI
44	González-González A, Muñoz-Muela E, Marchal JA, Cara FE, Molina MP, Cruz-Lozano M, Jiménez G, Verma A, Ramírez A, Qian W, et al: Activating transcription factor 4 modulates TGFβ-induced aggressiveness in triple-negative breast cancer via SMAD2/3/4 and mTORC2 signaling. Clin Cancer Res. 24:5697–5709. 2018. View Article : Google Scholar : PubMed/NCBI
45	Rundqvist H and Johnson RS: Tumour oxygenation: Implications for breast cancer prognosis. J Intern Med. 274:105–112. 2013. View Article : Google Scholar : PubMed/NCBI
46	Vaupel P, Schlenger K, Knoop C and Höckel M: Oxygenation of human tumors: Evaluation of tissue oxygen distribution in breast cancers by computerized O2 tension measurements. Cancer Res. 51:3316–3322. 1991.PubMed/NCBI
47	Lando D, Peet DJ, Whelan DA, Gorman JJ and Whitelaw ML: Asparagine hydroxylation of the HIF transactivation domain a hypoxic switch. Science. 295:858–861. 2002. View Article : Google Scholar : PubMed/NCBI
48	Gradiz R, Silva HC, Carvalho L, Botelho MF and Mota-Pinto A: MIA PaCa-2 and PANC-1-pancreas ductal adenocarcinoma cell lines with neuroendocrine differentiation and somatostatin receptors. Sci Rep. 6:216482016. View Article : Google Scholar : PubMed/NCBI
49	Yoshida T, Ri M, Kinoshita S, Narita T, Totani H, Ashour R, Ito A, Kusumoto S, Ishida T, Komatsu H and Iida S: Low expression of neural cell adhesion molecule, CD56, is associated with low efficacy of bortezomib plus dexamethasone therapy in multiple myeloma. PLoS One. 13:e01967802018. View Article : Google Scholar : PubMed/NCBI
50	Garcia-Carbonero N, Li W, Cabeza-Morales M, Martinez-Useros J and Garcia-Foncillas J: New hope for pancreatic ductal adenocarcinoma treatment targeting endoplasmic reticulum stress response: A systematic review. Int J Mol Sci. 19:24682018. View Article : Google Scholar : PubMed/NCBI