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Molecular characteristics and induction profiles of hypoxia-inducible factor-1α and other basic helix–loop–helix and Per–Arnt–Sim domain-containing proteins identified in a carcinogenic liver fluke Clonorchis sinensis

Published online by Cambridge University Press:  02 August 2018

Seon-Hee Kim
Department of Microbiology, Gachon University College of Medicine, 191 Hambakmoe-ro, Yeonsu-gu, Incheon 21936, Republic of Korea
Gyu-Seok Oh
Department of Microbiology, Gachon University College of Medicine, 191 Hambakmoe-ro, Yeonsu-gu, Incheon 21936, Republic of Korea
Woon-Mok Sohn
Department of Parasitology, Institute of Health Sciences, Gyeongsang National University College of Medicine, Jinju 52727, Republic of Korea
Kihyun Lee
Department of Systems Biotechnology, Center for Antibiotic Resistome, Chung-Ang University, Anseong 17546, Republic of Korea
Hyun-Jong Yang
Department of Parasitology, Ewha Womans University School of Medicine, Seoul 03760, Republic of Korea
Young-An Bae*
Department of Microbiology, Gachon University College of Medicine, 191 Hambakmoe-ro, Yeonsu-gu, Incheon 21936, Republic of Korea
Author for correspondence: Young-An Bae, E-mail:


Clonorchis sinensis (C. sinensis), a trematode parasite that invades the hypoxic hepatobiliary tract of vertebrate hosts requires a considerable amount of oxygen for its sexual reproduction and energy metabolism. However, little is known regarding the molecular mechanism of C. sinensis involved in the adaptation to the hypoxic environments. In this study, we investigated the molecular structures and induction patterns of hypoxia-inducible factor-1α (HIF-1α) and other basic helix–loop–helix and Per–Arnt–Sim (bHLH–PAS) domain-containing proteins such as HIF-1β, single-minded protein and aryl hydrocarbon receptor, which might prompt adaptive response to hypoxia, in C. sinensis. These proteins possessed various bHLH–PAS family-specific domains. Expression of C. sinensis HIF-1α (CsHIF-1α) was highly induced in worms which were either exposed to a hypoxic condition or co-incubated with human cholangiocytes. In addition to oxygen, nitric oxide and nitrite affected the CsHIF-1α expression depending on the surrounding oxygen concentration. Treatment using a prolyl hydroxylase-domain protein inhibitor under 20%-oxygen condition resulted in an increase in the CsHIF-1α level. Conversely, the other bHLH–PAS genes were less responsive to these exogenous stimuli. We suggest that nitrite and nitric oxide, as well as oxygen, coordinately involve in the regulation of HIF-1α expression to adapt to the hypoxic host environments in C. sinensis.

Research Article
Copyright © Cambridge University Press 2018 

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Adachi, J, Mori, Y, Matsui, S, Takigami, H, Fujino, J, Kitagawa, H, Miller, CA III, Kato, T, Saeki, K and Matsuda, T (2001) Indirubin and indigo are potent aryl hydrocarbon receptor ligands present in human urine. The Journal of Biological Chemistry 276, 3147531478.Google Scholar
Bae, YA, Ahn, DW, Lee, EG, Kim, SH, Cai, GB, Kang, I, Sohn, WM and Kong, Y (2013 a) Differential activation of diverse glutathione transferases of Clonorchis sinensis in response to the host bile and oxidative stressors. PLoS Neglected Tropical Diseases 7, e2211.Google Scholar
Bae, YA, Cai, GB, Kim, SH, Sohn, WM and Kong, Y (2013 b) Expression pattern and substrate specificity of Clonorchis sinensis tyrosinases. International Journal for Parasitology 43, 891900.Google Scholar
Bailey-Serres, J, Fukao, T, Gibbs, DJ, Holdsworth, MJ, Lee, SC, Licausi, F, Perata, P, Voesenek, ACJ and van Dongen, JT (2012) Making sense of low oxygen sensing. Trends in Plant Science 17, 129138.Google Scholar
Bartels, K, Grenz, A and Eltzschig, HK (2013) Hypoxia and inflammation are two sides of the same coin. Proceedings of the National Academy of Sciences of the United States of America 110, 1835118352.Google Scholar
Bertout, JA, Patel, SA and Simon, MC (2008) The impact of O2 availability on human cancer. Nature Reviews. Cancer 8, 967975.Google Scholar
Biddlestone, J, Bandarra, D and Rocha, S (2015) The role of hypoxia in inflammatory disease (review). International Journal of Molecular Medicine 35, 859869.Google Scholar
Bouvard, V, Baan, R, Straif, K, Grosse, Y, Secretan, B, El Ghissassi, F, Benbrahim-Tallaa, L, Guha, N, Freeman, C, Galichet, L, Cogliano, V and WHO International Agency for Research on Cancer Monograph Working Group (2009) A review of human carcinogens – part B: biological agents. The Lancet Oncology 10, 321322.Google Scholar
Brook, I (1989) Aerobic and anaerobic microbiology of biliary tract disease. Journal of Clinical Microbiology 27, 23732375.Google Scholar
Chandel, NS and Budinger, GR (2007) The cellular basis for diverse responses to oxygen. Free Radical Biology and Medicine 42, 165174.Google Scholar
Chandrasekharan, JA and Sharma-Walia, N (2015) Lipoxins: nature's way to resolve inflammation. Journal of Inflammation Research 30, 181192.Google Scholar
Cordingley, JS (1987) Trematode eggshells: novel protein biopolymers. Parasitology Today 3, 341344.Google Scholar
Cossins, A and Berenbrink, M (2008) Physiology: myoglobin's new clothes. Nature 454, 416417.Google Scholar
Crews, ST (1998) Control of cell lineage-specific development and transcription by bHLH-PAS proteins. Genes & Development 12, 607620.Google Scholar
Dennis, EA and Norris, PC (2015) Eicosanoid storm in infection and inflammation. Nature Reviews. Immunology 15, 511523.Google Scholar
Dyson, A and Singer, M (2011) Tissue oxygen tension monitoring: will it fill the void? Current Opinion in Critical Care 17, 281289.Google Scholar
Edgar, RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research 32, 17921797.Google Scholar
Fritsch, P, de Saint Blanquat, G and Klein, D (1985) Excretion of nitrates and nitrites in saliva and bile in the dog. Food and Chemical Toxicology 23, 655659.Google Scholar
Goto, M, Amino, H, Nakajima, M, Tsuji, N, Sakamoto, K and Kita, K (2013) Cloning and characterization of hypoxia-inducible factor-1 subunits from Ascaris suum – a parasitic nematode highly adapted to change of oxygen conditions during its life cycle. Gene 516, 3947.Google Scholar
Gunda, VG and Janapala, VR (2009) Effects of dissolved oxygen levels on survival and growth of Haliclona pigmentifera (Demospongiae). Cell and Tissue Research 337, 527535.Google Scholar
Huang, J, Zhao, Q, Mooney, SM and Lee, FS (2002) Sequence determinants in hypoxia-inducible factor-1 alpha for hydroxylation by the prolyl hydroxylases PHD1, PHD2, and PHD3. The Journal of Biological Chemistry 277, 3979239800.Google Scholar
Huang, X and Madan, A (1999) CAP3: a DNA sequence assembly program. Genome Research 9, 868877.Google Scholar
Huang, Y, Chen, W, Wang, X, Liu, H, Chen, Y, Guo, L, Luo, F, Sun, J, Mao, Q, Liang, P, Xie, Z, Zhou, C, Tian, Y, Lv, X, Huang, L, Zhou, J, Hu, Y, Li, R, Zhang, F, Lei, H, Li, W, Hu, X, Liang, C, Xu, J, Li, X and Yu, X (2013) The carcinogenic liver fluke, Clonorchis sinensis: new assembly, reannotation and analysis of the genome and characterization of tissue transcriptomes. PLoS ONE 8, e54732.Google Scholar
Jiang, H, Guo, R and Powell-Coffman, J (2001) The Caenorhabditis elegans hif-1 gene encodes a bHLH-PAS protein that is required for adaptation to hypoxia. Proceedings of the National Academy of Sciences of the United States of America 98, 79167921.Google Scholar
Jürgens, KD, Papadopoulos, S, Peters, T and Gros, G (2000) Myoglobin: just an oxygen store or also an oxygen transporter? News in Physiological Sciences 15, 269274.Google Scholar
Keiser, J and Utzinger, J (2009) Food-borne trematodiases. Clinical Microbiology Reviews 22, 466483.Google Scholar
Kiger, L, Rashid, AK, Griffon, N, Haque, M, Moens, L, Gibson, QH, Poyart, C and Marden, MC (1998) Trematode hemoglobins show exceptionally high oxygen affinity. Biophysical Journal 75, 990998.Google Scholar
Kim, TI, Yoo, YG, Kwak, BK, Seok, JW and Hong, SJ (2011) Tracing of the bile-chemotactic migration of juvenile Clonorchis sinensis in rabbits by PET-CT. PLoS Neglected Tropical Diseases 5, e1414.Google Scholar
Kiriakidis, S, Esteban, MA and Maxwell, PH (2007) Genetic insights into the hypoxia-inducible factor (HIF) pathway. Advances in Enzyme Regulation 47, 288306.Google Scholar
Li, KL, Lu, TM and Yu, JK (2014) Genome-wide survey and expression analysis of the bHLH-PAS genes in the amphioxus Branchiostoma floridae reveal both conserved and diverged expression patterns between cephalochordates and vertebrates. EvoDevo 5, 20.Google Scholar
Livak, KJ and Schmittgen, TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔC T method. Methods 25, 402408.Google Scholar
Loenarz, C, Coleman, ML, Boleininger, A, Schierwater, B, Holland, PWH, Ratcliffe, PJ and Schofield, CJ (2011) The hypoxia-inducible transcription factor pathway regulates oxygen sensing in the simplest animal, Trichoplax adherens. EMBO Reports 12, 6370.Google Scholar
Lun, ZR, Gasser, RB, Lai, DH, Li, AX, Zhu, XQ, Yu, XB and Fang, YY (2005) Clonorchiasis: a key foodborne zoonosis in China. The Lancet. Infectious Diseases 5, 3141.Google Scholar
Majmundar, AJ, Wong, WJ and Simon, MC (2010) Hypoxia-inducible factors and the response to hypoxic stress. Molecular Cell 40, 294309.Google Scholar
Mao, Q, Xie, Z, Wang, X, Chen, W, Ren, M, Shang, M, Lei, H, Tian, Y, Li, S, Liang, P, Chen, T, Liang, C, Xu, J, Li, X, Huang, Y and Yu, X (2015) Clonorchis sinensis ferritin heavy chain triggers free radicals and mediates inflammation signaling in human hepatic stellate cells. Parasitology Research 114, 659670.Google Scholar
Mei, Q and Dvornyk, V (2014) Evolution of PAS domains and PAS-containing genes in eukaryotes. Chromosoma 123, 385405.Google Scholar
Olson, N and van der Vliet, A (2011) Interactions between nitric oxide and hypoxia-inducible factor signaling pathways in inflammatory disease. Nitric Oxide 25, 125137.Google Scholar
Page, RD (1996) Tree view: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences 12, 357358.Google Scholar
Powell-Coffman, J, Bradfield, CA and Wood, WB (1998) Caenorhabditis elegans orthologs of the aryl hydrocarbon receptor and its heterodimerization partner the aryl hydrocarbon nuclear translocator. Proceedings of the National Academy of Sciences of the United States of America 95, 28442849.Google Scholar
Qian, MB, Utzinger, J, Keiser, J and Zhou, XN (2016) Clonorchiasis. Lancet 387, 800810.Google Scholar
Rytkönen, KT, Williams, TA, Renshaw, GM, Primmer, CR and Nikinmaa, M (2011) Molecular evolution of the metazoan PHD-HIF oxygen-sensing system. Molecular Biology and Evolution 28, 19131926.Google Scholar
Sardiello, M, Palmieri, M, di Ronza, A, Medina, DL, Valenza, M, Gennarino, VA, Di Malta, C, Donaudy, F, Embrione, V, Polishchuk, RS, Banfi, S, Parenti, G, Cattaneo, E and Ballabio, A (2009) A gene network regulating lysosomal biogenesis and function. Science 325, 473477.Google Scholar
Schultz, J, Milpetz, F, Bork, P and Ponting, CP (1998) SMART, a simple modular architecture research tool: identification of signaling domains. Proceedings of the National Academy of Sciences of the United States of America 95, 58575864.Google Scholar
Seidel, SD, Winters, GM, Rogers, WJ, Ziccardi, MH, Li, V, Keser, B and Denison, MS (2001) Activation of the Ah receptor signaling pathway by prostaglandins. Journal of Biochemical and Molecular Toxicology 15, 187196.Google Scholar
Semenza, GL (2009) Regulation of oxygen homeostasis by hypoxia-inducible factor 1. Physiology 24, 97106.Google Scholar
Shin, HR, Oh, JK, Masuyer, E, Curado, MP, Bouvard, V, Fang, YY, Wianqnon, S, Sripa, B and Hong, ST (2010) Epidemiology of cholangiocarcinoma: an update focusing on risk factors. Cancer Science 101, 579585.Google Scholar
Sinal, CJ and Bend, JR (1997) Aryl hydrocarbon receptor-dependent induction of Cyp1a1 by bilirubin in mouse hepatoma hepa 1c1c7 cells. Molecular Pharmacology 52, 590599.Google Scholar
Smyth, JD and Halton, DW (1983) The Physiology of Trematodes, 2nd Edn. New York, NK: Cambridge University Press.Google Scholar
Sorci, G and Faivre, B (2009) Inflammation and oxidative stress in vertebrate host-parasite systems. Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences 364, 7183.Google Scholar
Tamura, K, Stecher, G, Peterson, D, Filipski, A and Kumar, S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Molecular Biology and Evolution 30, 27252729.Google Scholar
Tielens, AG, van den Heuvel, JM and van den Bergh, SG (1984) The energy metabolism of Fasciola hepatica during its development in the final host. Molecular and Biochemical Parasitology 13, 301307.Google Scholar
Wang, G, Yu, Z, Zhen, Y, Mi, T, Shi, Y, Wang, J, Wang, M and Sun, S (2014) Molecular characterization, evolution and expression of hypoxia-inducible factor in Aurelia sp.1. PLoS ONE 9, e100057.Google Scholar
Wang, GL and Semenza, GL (1993) General involvement of hypoxia-inducible factor 1 in transcriptional response to hypoxia. Proceedings of the National Academy of Sciences of the United States of America 90, 43044308.Google Scholar
Wang, GL, Jiang, BH, Rue, EA and Semenza, GL (1995) Hypoxia-inducible factor 1 is a basic helix-loop-helix-PAS heterodimer regulated by cellular O2 tension. Proceedings of the National Academy of Sciences of the United States of America 92, 55105514.Google Scholar
Wittenberg, JB (1959) Oxygen transport: a new function proposed for myoglobin. The Biological Bulletin 117, 402403.Google Scholar
Yang, QL, Shen, JQ, Xue, Y, Cheng, XB, Jiang, ZH, Yang, YC, Chen, YD and Zhou, XN (2015) Pathological lesions and inducible nitric oxide synthase expressions in the liver of mice experimentally infected with Clonorchis sinensis. Korean Journal of Parasitology 53, 777783.Google Scholar
Young, ND, Campbell, BE, Hall, RS, Jex, AR, Cantacessi, C, Laha, T, Sohn, WM, Sripa, B, Loukas, A, Brindley, PJ and Gasser, RB (2010) Unlocking the transcriptomes of two carcinogenic parasites, Clonorchis sinensis and Opisthorchis viverrini. PLoS Neglected Tropical Diseases 4, e719.Google Scholar
Young, ND, Nagarajan, N, Lin, SJ, Korhonen, PK, Jex, AR, Hall, RS, Safavi-Hemami, H, Kaewkong, W, Bertrand, D, Gao, S, Seet, Q, Wongkham, S, The, BT, Wongkham, C, Intapan, PM, Maleewong, W, Yang, X, Hu, M, Wang, Z, Hofmann, A, Sternberg, PW, Tan, P, Wang, J and Gasser, RB (2014) The Opisthorchis viverrini genome provides insights into life in the bile duct. Nature Communications 5, 4378.Google Scholar
Zhang, N, Fu, Z, Linke, S, Chicher, J, Gorman, JJ, Visk, D, Haddad, GG, Poellinger, L, Peet, DJ, Powell, F and Johnson, RS (2010) The asparaginyl hydroxylase factor inhibiting HIF-1α is an essential regulator of metabolism. Cell Metabolism 11, 364378.Google Scholar
Zhang, Y (2008) I-TASSER server for protein 3D structure prediction. BMC Bioinformatics 9, 40.Google Scholar
Zhdanov, AV, Okkelman, IA, Collins, FW, Melgar, S and Papkovsky, DB (2015) A novel effect of DMOG on cell metabolism: direct inhibition of mitochondrial function precedes HIF-target gene expression. Biochimica et Biophysica Acta 1847, 12541266.Google Scholar
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