Acute and chronic hypoxia affects HIF-1α mRNA levels in sea bass (Dicentrarchus labrax) Genciana Terova ⁎, Simona Rimoldi, Samuela Corà, Giovanni Bernardini, Rosalba Gornati, Marco Saroglia Department of Biotechnology and Molecular Sciences (DBSM) University of Insubria, Via J.H.Dunant, 321100 Varese, Italy ABSTRACT ARTICLE INFO Article history: Received 3 October 2007 Received in revised form 17 March 2008 Accepted 22 March 2008 Keywords: Sea bass Aquaculture Hypoxia Real-time PCR Gene expression Aquatic hypoxia is a frequent event and in fish a complex set of physiological and biochemical alterations are employed to cope with this environmental stress. Many of these adjustments depend to a large extent on changes in the expression of genes that encode for physiologically relevant proteins. Genes that are induced by hypoxia appear to share a common mode of transcriptional regulation. This induction depends upon activation of a transcription factor, the hypoxia-inducible factor-1 (HIF-1), which is composed of α and β subunits. In this study we report first on the molecular cloning and characterization of HIF-1α in sea bass (Dicentrarchus labrax). The full-length sea bass cDNA for HIF-1α was isolated and deposited in the GenBank with accession no. DQ171936. It consists of 3317 base pairs (bp) carrying a single open-reading frame that encompasses 2265 bp of the coding region and 1052 bp of the 3′ UTR. The predicted sea bass HIF-1α protein (755 amino acids) shows a high level of conservation at the bHLH (basic helix–loop–helix domain), PAS A/B (Per-ARNT-Sim A/B domain), N-TAD (N-terminal transactivation domain), and C-TAD (C-terminal transacti- vation domain), whereas only few variations are found at the ODD (oxygen-dependent degradation domain) regions among vertebrates. We then utilized the real-time PCR technology to monitor dynamic changes in levels of HIF-1α mRNA in response to acute and chronic hypoxic stress. The number of HIF-1α mRNA copies was significantly increased in liver tissue in response to both acute (1.9 mg/L, dissolved oxygen for 4 h) and chronic (4.3 mg/l, DO for 15 days) hypoxia in sea bass whereas it remained unchanged in fish exposed to hyperoxic (DO 13.5±1.2 mg/L, 155% saturation) conditions. This is the first study to investigate the behaviour of HIF-1α gene transcripts during hypoxia in a representative of marine, hypoxia-sensitive species. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Oxygen is essential to life for most organisms, but changes in the environment can reduce the availability of oxygen. In general, hypoxia even for brief periods can be detrimental or fatal to humans and most mammals as they possess only little tolerance to anoxia and their tissues are normally debilitated by any prolonged lack of O 2 (van der Meer et al., 2005). However, certain vertebrate species have evolved to subsist on low amounts of oxygen for prolonged periods owing to a range of specially developed physiological mechanisms known as the “hypoxia response” (for a review see Nikinmaa, 2002; Nikinmaa and Rees, 2005). Fish species that live and survive in environments with low and/or variable oxygen levels represent an example of this. Thus, it can be expected that these animals have highly evolved mechanisms for surviving aquatic hypoxia that are perhaps more sophisticated than those of the more extensively studied terrestrial mammals. In fish a complex set of physiological and biochemical alterations are employed to cope with this environmental stress. These strategies include decreased metabolic rate (DallaVia et al., 1994), increased ventilation rate, hematocrit and haemoglobin O 2 affinity (Jensen et al., 1993), and increased anaerobic respiration (Virani and Rees, 2000). Many of these adjustments depend to a large extent on changes in the expression of genes that encode diverse groups of physiologically relevant proteins. Gracey et al. (2001) recently identified alterations in the expression of over 120 genes in hypoxic fish (Gillichthys mirabilis), whereas in mammals, hypoxia-induced changes in the expression of a wide range of genes were reported (for a review see Bruick, 2003), including numerous genes homologues to those observed in fish. Examples include genes involved in glycolysis, gluconeogenesis, iron metabolism, cell survival and proliferation, translational machinery, and muscle contraction (Semenza, 1999; Wenger, 2002; Gracey et al., 2001). In mammals, where HIF has been extensively studied, genes that are induced by hypoxia appear to share a common mode of transcriptional regulation. This induction depends upon activation of a transcription factor, the hypoxia-inducible factor-1 (HIF-1). Under hypoxic conditions, the HIF-1 binds specifically to a consensus sequence (5′- RCGTG - 3′) known as the hypoxia-responsive element (HRE) in the promoter or enhancer of various hypoxia-inducible genes Aquaculture 279 (2008) 150–159 ⁎ Corresponding author. Tel.: +39 0332 421 428; fax: +390332 421 500. E-mail address: genciana.terova@uninsubria.it (G. Terova). 0044-8486/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.aquaculture.2008.03.041 Contents lists available at ScienceDirect Aquaculture journal homepage: www.elsevier.com/locate/aqua-online