ReviewHypoxia-dependent activation of HIF into a transcriptional regulator
Introduction
Throughout evolution, it is evident that most higher eukaryotes have developed a variety of strategies to adapt to the increase in animal size, complexity and metabolic functions, in an atmosphere with increasing oxygen levels (up to the 21% O2 observed in modern times). The necessity to maintain a careful balance between the amount of oxygen to be delivered to a specific cell or tissue and the need to avoid generation of toxic reactive oxygen species by excess oxygen supply, has driven the development of several efficient mechanisms of homeostasis, both at the systemic and cellular level. Oxygen is used by cells not only for mitochondrial ATP production but also as a substrate in a large number of enzymatic reactions that can be strongly affected by local changes in pO2 [1]. For these reasons, in a poorly oxygenated environment (hypoxia), cell and tissue viability depends on the activation of several molecular processes that will ultimately lead to changes in protein activity and gene expression [2]. One of the primary targets for oxygen-dependent regulation is the transcriptional activator hypoxia-inducible factor-1 (HIF-1) [3], [4]. This transcriptional activator is tightly regulated by cellular oxygen levels and is strictly necessary for the activation of a network of genes that encode for proteins needed to maintain tissue viability by adjusting cell metabolism (glucose transporters, glycolytic enzymes), erythropoiesis (erythropoietin), vasomotor control (endothelin-1), angiogenesis (vascular endothelial growth factor (VEGF)) and tissue remodeling (placental growth factor (PLGF)) [5]. The activation of HIF to its fully competent form, able to bind specific DNA consensus sequences (hypoxia-response elements or HREs) present in the regulatory regions of target genes and recruit the necessary coactivator proteins to initiate transcription, is achieved through a series of events that will be the focus of this review.
In this step-wise activation process, several oxygen-dependent enzymatic activities have been identified that act on specific HIF-1 amino acid residues (summarized in Table 1). These post-translational modifications dictate the formation of different HIF-1-associated protein complexes involved in the control of protein stability, subcellular compartmentalization and transactivation function of this transcription factor.
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HIF-1 subunit composition, structure and function
HIF-1 is a heterodimer formed by two members of the basic helix–loop–helix (bHLH)/PER–ARNT–SIM (PAS) family of transcription factors: HIF-1α and ARNT (also termed HIF-1β) (Fig. 1). ARNT is the previously characterized aryl hydrocarbon receptor nuclear translocator that can serve as DNA-binding partner for other bHLH/PAS proteins such as the aryl hydrocarbon receptor (AhR) or the single minded proteins (SIM-1 and SIM-2) [6]. HIF-1α is the oxygen-regulated subunit, which has been the subject of
HIF-1 regulation by control of protein stability
Oxygen levels do not seem to significantly affect the activity of ARNT as a transcription factor, which is constitutively expressed and localized to the nuclear compartment due to an N-terminal nuclear localization signal (NLS) [17]. Hypoxia may, however indirectly, limit the availability of ARNT as a dimerization partner in the cell nucleus, since at low oxygen concentrations the ARNT–HIF-α combination seems to be favored above all others due to the dominating affinity of HIF-1α for ARNT [18].
HIF-1-mediated activation of transcription
Stabilization of HIF-1α is the first step in a cascade of events that will ultimately lead to the recruitment of a variety of proteins involved in the activation of transcription of hypoxia-inducible genes, the products of which will participate in the adaptation to the hypoxic environment. In eukaryotic cells, transcriptional activation mechanisms are complex events that depend on the modulation of the amount, activity and localization of many proteins through post-translational modification,
Subcellular compartmentalization and HIF regulation
The use of fluorescence microscopy together with green fluorescent protein (GFP) and GFP-related technologies has led to the identification of another important level of regulation in the HIF signaling pathway. Several studies focused on the subcellular distribution of proteins involved in HIF regulation have shown that compartmentalization of several HIF partner proteins (e.g. HIF-1α, PHDs, FIH-1) participates in the modulation of their overall activity. The intracellular redistribution of
HIF-1-dependent transactivation in a cell- or promoter-specific context
Although HIF-1 is ubiquitously expressed and activated by hypoxia [80], the pattern of tissue-specific expression of HIF-1 target genes, such as erythropoietin (Epo), is not altered upon activation. Epo is expressed in the kidney and liver in a hypoxia-inducible manner, which is mediated by an enhancer situated in the 3′ region of the gene [81]. Early observations determined that binding of HIF-1 to an HRE present in the 3′ enhancer was critical for Epo induction under hypoxia [3]. It was
Acknowledgement
We thank the members of the Poellinger lab for critical review and suggestions.
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