Molecular characterization and expression regulation of the factor-inhibiting HIF-1 (FIH-1) gene under hypoxic stress in bighead carp (Aristichthys nobilis)

  • Xiu Feng
  • Xiaomu Yu
  • Meixia Pang
  • Jingou TongEmail author


Factor-inhibiting HIF-1 (FIH-1) is an asparagine hydroxylase that interacts with hypoxia-inducible factor 1α (HIF-1α) to regulate transcriptional activity of HIF-1. Few studies of fish FIH-1 have been reported to date. In this study, the cDNA of FIH-1 gene was cloned and characterized for bighead carp, Aristichthys nobilis (AnFIH-1). The AnFIH-1 cDNA is 2065 bp in length, encoding a protein of 357 amino acid (aa) residues, which contains a JmjC homology region of the jumonji transcription factors. AnFIH-1 shares high identities with other vertebrate FIH-1 (79.1–96.4%), especially in the JmjC homology region, suggesting its conserved function. During the embryonic stages of A. nobilis, AnFIH-1 had significantly high expression levels in unfertilized egg and blastula. In healthy tissues, its predominant mRNA expression was detected in muscle. The mRNA levels of AnFIH-1 were significantly upregulated in the liver, gill, hypothalamus, and spleen after hypoxic treatment, and then decreased to pretreatment levels after 6-h re-oxygenation. However, in the muscle, continual increasing of mRNA expression was observed after hypoxic shock and re-oxygenation. These results indicate that FIH-1 may play an important role in physiological regulation for adapting to hypoxia stress in A. nobilis.


Aristichthys nobilis Factor-inhibiting HIF-1 Gene expression Hypoxia 



This study was supported by grants from the NSFC (31272647, 31472268) of China. We thank Xinhua Wang, Haiyang Liu, and Xueli Liu for their assistaence in sample preparation and laboratory work.


  1. Bracken CP, Whitelaw ML, Peet DJ (2003) The hypoxia-inducible factors: key transcriptional regulators of hypoxic responses. Cell Mol Life Sci 60:1376–1393CrossRefGoogle Scholar
  2. Brahimi-Horn MC, Pouyssegur J (2009) HIF at a glance. J Cell Sci 122:1055–1057CrossRefGoogle Scholar
  3. Chen N, Chen LP, Zhang J, Chen C, Wei XL, Gul Y, Wang WM, Wang HL (2012) Molecular characterization and expression analysis of three hypoxia-inducible factor alpha subunits, HIF-1α/2α/3α of the hypoxia-sensitive freshwater species, Chinese sucker. Gene 498:81–90CrossRefGoogle Scholar
  4. Chi W, Gan XN, Xiao WH, Wang W, He SP (2013) Different evolutionary patterns of hypoxia-inducible factor α (HIF-α) isoforms in the basal branches of Actinopterygii and Sarcopterygii. FEBS Open Bio 3:479–483CrossRefGoogle Scholar
  5. Clissold PM, Ponting CP (2001) JmjC: cupin metalloenzyme-like domains in jumonji, hairless and phospholipase A2β. Trends Biochem Sci 26:7–9CrossRefGoogle Scholar
  6. Cockman ME, Masson N, Mole DR, Jaakkola P, Chang GW, Clifford SC, Maher ER, Pugh CW, Ratcliffe PJ, Maxwell PH (2000) Hypoxia inducible factor-α binding and ubiquitylation by the von Hippel-Lindau tumor suppressor protein. J Biol Chem 275:25733–25741CrossRefGoogle Scholar
  7. Couvelard A, Deschamps L, Rebours V, Sauvanet A, Gatter K, Pezzella F, Ruszniewski P, Bedossa P (2008) Overexpression of the oxygen sensors PHD-1, PHD-2, PHD-3, and FIH is associated with tumor aggressiveness in pancreatic endocrine tumors. Clin Cancer Res 14:6634–6639CrossRefGoogle Scholar
  8. Elkins JM, Hewitson KS, McNeill LA, Seibel JF, Schlemminger I, Pugh CW, Ratcliffe PJ, Schofield CJ (2003) Structure of factor-inhibiting hypoxia-inducible factor (HIF) reveals mechanism of oxidative modification of HIF-1α. J Biol Chem 278:1802–1806CrossRefGoogle Scholar
  9. Geng X, Feng J, Liu S, Wang Y, Arias C, Liu Z (2014) Transcriptional regulation of hypoxia inducible factors alpha (HIF-α) and their inhibiting factor (FIH-1) of channel catfish (Ictalurus punctatus) under hypoxia. Comp Biochem Physiol B 169:38–50CrossRefGoogle Scholar
  10. Ivan M, Kondo K, Yang HF, Kim W, Valiando J, Ohh M, Salic A, Asara JM, Lane WS, Kaelin WG Jr (2001) HIFα targeted for VHL-mediated destruction by proline hydroxylation: implications for O2 sensing. Science 292:464–468CrossRefGoogle Scholar
  11. Koivunen P, Hirsila M, Gunzler V, Kivirikko KI, Myllyharju J (2004) Catalytic properties of the asparaginyl hydroxylase (FIH) in the oxygen sensing pathway are distinct from those of its prolyl 4-hydroxylases. J Biol Chem 279:9899–9904CrossRefGoogle Scholar
  12. Kuzmanov A, Wielockx B, Rezaei M, Kettelhake A, Breier G (2012) Overexpression of factor inhibiting HIF-1 enhances vessel maturation and tumor growth via platelet-derived growth factor-C. Int J Cancer 131:E603–EE13CrossRefGoogle Scholar
  13. Kwasek K, Rimoldi S, Cattaneo AG, Parker T, Dabrowski K, Terova G (2017) The expression of hypoxia-inducible factor-1α gene is not affected by low-oxygen conditions in yellow perch (Perca flavescens) juveniles. Fish Physiol Biochem 43:849–862CrossRefGoogle Scholar
  14. Lando D, Peet DJ, Gorman JJ, Whelan DA, Whitelaw ML, Bruick RK (2002) FIH-1 is an asparaginyl hydroxylase enzyme that regulates the transcriptional activity of hypoxia-inducible factor. Genes Dev 16:1466–1471CrossRefGoogle Scholar
  15. Law SHW, Wu RSS, Ng PKS, Yu RMK, Kong RYC (2006) Cloning and expression analysis of two distinct HIF-alpha isoforms - gcHIF-1alpha and gcHIF-4alpha- from the hypoxia-tolerant grass carp, Ctenopharyngodon idellus. BMC Mol Biol 7(15)Google Scholar
  16. Lee Y, Song AJ, Baker R, Micales B, Conway SJ, Lyons GE (2000) Jumonji, a nuclear protein that is necessary for normal heart development. Circ Res 86:932–938CrossRefGoogle Scholar
  17. Lee C, Kim SJ, Jeong DG, Lee SM, Ryu SE (2003) Structure of human FIH-1 reveals a unique active site pocket and interaction sites for HIF-1 and von Hippel-Lindau. J Biol Chem 278:7558–7563CrossRefGoogle Scholar
  18. Li HL, Gu XH, Li BJ, Chen X, Lin HR, Xia JH (2017) Characterization and functional analysis of hypoxia-inducible factor HIF1α and its inhibitor HIF1αn in tilapia. PLoS One 12:e0173478CrossRefGoogle Scholar
  19. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method. Methods 25:402–408CrossRefGoogle Scholar
  20. Mahon PC, Hirota K, Semenza GL (2001) FIH-1: a novel protein that interacts with HIF-1α and VHL to mediate repression of HIF-1 transcriptional activity. Genes Dev 15:2675–2686CrossRefGoogle Scholar
  21. Maxwell PH, Wiesener MS, Chang GW, Clifford SC, Vaux EC, Cockman ME, Wykoff CC, Pugh CW, Maher ER, Ratcliffe PJ (1999) The tumour suppressor protein VHL targets hypoxia-inducible factors for oxygen-dependent proteolysis. Nature 399:271–275CrossRefGoogle Scholar
  22. Min JH, Yang HF, Ivan M, Gertler F, Kaelin WG, Pavletich NP (2002) Structure of an HIF-1α-pVHL complex: hydroxyproline recognition in signaling. Science 296:1886–1889CrossRefGoogle Scholar
  23. Mohindra V, Tripathi RK, Singh A, Singh B (2013) Molecular characterization and expression analysis of a novel cystatin-like gene in a hypoxia-tolerant Indian catfish, Clarias batrachus [Linnaeus, 1758]. Fish Shellfish Immun 34:683–687CrossRefGoogle Scholar
  24. Ni M, Wen H, Li J, Chi M, Bu Y, Ren Y, Zhang M, Song Z, Ding H (2014) The physiological performance and immune responses of juvenile Amur sturgeon (Acipenser schrenckii) to stocking density and hypoxia stress. Fish Shellfish Immun 36:325–335CrossRefGoogle Scholar
  25. Nikinmaa M, Rees BB (2005) Oxygen-dependent gene expression in fishes. Am J Phys Regul Integr Comp Phys 288:R1079–R1090Google Scholar
  26. Quan FB, Kenigfest NB, Mazan S, Tostivint H (2013) Molecular cloning of the cDNAs encoding three somatostatin variants in the dogfish (Scylorhinus canicula). Gen Comp Endocrinol 180:1–6CrossRefGoogle Scholar
  27. Querat B, Sellouk A, Salmon C (2000) Phylogenetic analysis of the vertebrate glycoprotein hormone family including new sequences of sturgeon (Acipenser baeri) β subunits of the two gonadotropins and the thyroid-stimulating hormone. Biol Reprod 63:222–228CrossRefGoogle Scholar
  28. Rahman MS, Thomas P (2007) Molecular cloning, characterization and expression of two hypoxia-inducible factor alpha subunits, HIF-1α and HIF-2α, in a hypoxia-tolerant marine teleost, Atlantic croaker (Micropogonias undulatus). Gene 396:273–282CrossRefGoogle Scholar
  29. Rimoldi S, Terova G, Ceccuzzi P, Marelli S, Antonini M, Saroglia M (2012) HIF-1α mRNA levels in Eurasian perch (Perca fluviatilis) exposed to acute and chronic hypoxia. Mol Biol Rep 39:4009–4015CrossRefGoogle Scholar
  30. Semenza GL (1999) Regulation of mammalian O2 homeostasis by hypoxia-inducible factor 1. Annu Rev Cell Dev Biol 15:551–578CrossRefGoogle Scholar
  31. Shen RJ, Jiang XY, Pu JW, Zou SM (2010) HIF-1α and -2α genes in a hypoxia-sensitive teleost species Megalobrama amblycephala: cDNA cloning, expression and different responses to hypoxia. Comp Biochem Physiol B 157:273–280CrossRefGoogle Scholar
  32. Soilleux EJ, Turley H, Tian YM, Pugh CW, Gatter KC, Harris AL (2005) Use of novel monoclonal antibodies to determine the expression and distribution of the hypoxia regulatory factors PHD-1, PHD-2, PHD-3 and FIH in normal and neoplastic human tissues. Histopathology 47:602–610CrossRefGoogle Scholar
  33. Soitamo AJ, Råbergh CM, Gassmann M, Sistonen L, Nikinmaa M (2001) Characterization of a hypoxia-inducible factor (HIF-1alpha) from rainbow trout: accumulation of protein occurs at normal venous oxygen tension. J Biol Chem 276:19699–19705CrossRefGoogle Scholar
  34. Stolze IP, Tian YM, Appelhoff RJ, Turley H, Wykoff CC, Gleadle JM, Ratcliffe PJ (2004) Genetic analysis of the role of the asparaginyl hydroxylase factor inhibiting hypoxia-inducible factor (HIF) in regulating HIF transcriptional target genes. J Biol Chem 279:42719–42725CrossRefGoogle Scholar
  35. Su AI, Wiltshire T, Batalov S, Lapp H, Ching KA, Block D, Zhang J, Soden R, Hayakawa M, Kreiman G, Cooke MP, Walker JR, Hogenesch JB (2004) A gene atlas of the mouse and human protein-encoding transcriptomes. Proc Natl Acad Sci U S A 101:6062–6067CrossRefGoogle Scholar
  36. Terova G, Rimoldi S, Corà S, Bernardini G, Gornati R, Saroglia M (2008) Acute and chronic hypoxia affects HIF-1α mRNA levels in sea bass (Dicentrarchus labrax). Aquaculture 279:150–159CrossRefGoogle Scholar

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© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of HydrobiologyChinese Academy of SciencesWuhanChina
  2. 2.School of Applied Chemistry and Biological TechnologyShenzhen PolytechnicShenzhenChina

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