Transduction pathways involved in Hypoxia-Inducible Factor-1 phosphorylation and activation

doi:10.1016/S0891-5849(01)00657-8

Free Radical Biology and Medicine

Volume 31, Issue 7, 1 October 2001, Pages 847-855

https://doi.org/10.1016/S0891-5849(01)00657-8 Get rights and content

Abstract

Hypoxia-Inducible Factor-1 (HIF-1) is a transcription factor which is activated by hypoxia and involved in the adaptative response of the cell to oxygen deprivation. During hypoxic stress, HIF-1 triggers the overexpression of genes coding for glycolytic enzymes and angiogenic factors. To be active HIF-1 must be phosphorylated. HIF-1 is a substrate for various kinase pathways including PI-3K and the MAP kinases ERK and p38. Several transduction pathways have been proposed which act downstream of putative oxygen sensors and lead to the activation of these kinases. In this review, we summarize some of the latest advances describing the possible signaling pathways leading to HIF-1 phosphorylation and subsequent activation. The physiological relevance of these regulations is also discussed.

Introduction

Vascular injuries can lead to an inadequate flow of the blood, which results in a decrease in the delivery of oxygen and nutrients to the tissues [1]. These ischemic conditions can lead to tissue necrosis, cardiovascular and cerebral strokes, as well as thrombosis [1]. Mammalian cells have developed adaptative systems allowing them to survive to moderate or even severe hypoxia. This process involves an increase in the expression of genes coding for proteins responsible for the anaerobic production of ATP [2], [3], [4], [5], namely aldolase A, enolase-α, lactate dehydrogenase, pyruvate kinase, and glucose transporter-1. Hypoxic cells also secrete vascular endothelial growth factor (VEGF) [4], [6], an angiogenic growth factor allowing neovascularization of the hypoxic tissue.

Several transcription factors have been reported to be involved in the response to hypoxic stress (AP-1, NF-κB, HIF-1) [7]. Among these transcription factors Hypoxia-Inducible Factor-1 (HIF-1) is the most potent inducer of the expression of genes such as those coding for glycolytic enzymes, VEGF and erythropoietin (EPO) [2], [3], [4], [5]. HIF-1 is a heterodimer composed of the HIF-1α and ARNT-1 subunits [8]. Both subunits belong to the basic Helix-Loop-Helix-Per/ARNT/AhR/Sim (bHLH-PAS) transcription factor family [8]. HIF-1α as well as ARNT-1 are constitutively expressed [9]. However, HIF-1α appears to be the HIF-1 subunit regulated by hypoxia as under this condition, HIF-1α degradation is inhibited [10], [11], [12], whereas in contrast, it is rapidly degraded by the ubiquitin-proteasome system under normoxic conditions [10], [11], [12]. Under hypoxia, HIF-1α is seen to dissociate from the chaperone protein heat shock protein 90 (Hsp90) [13] and then translocates into the nucleus [14], where it dimerizes with the nuclear protein ARNT-1 to form the HIF-1 complex. To be fully activated, HIF-1 requires suitable redox conditions [15] as well as interaction with coactivators, namely CRB binding protein/p300 (CBP/p300) and steroid receptor coactivator-1 (SRC-1) [14].

The first evidence indicating that HIF-1 is a phospho-protein was produced by Wang and Semenza [16]. Using electrophoretic mobility shift assay (EMSA) they showed that when nuclear extracts of hypoxic Hep3B cells were treated with phosphatase, the HIF-1/DNA complex is disrupted [16]. They went on to demonstrate that the serine/threonine kinase inhibitor 2-aminopurine, the tyrosine kinase inhibitor genistein, as well as the serine threonine phosphatase inhibitor NaF, are able to inhibit HIF-1-DNA-binding activity and HIF-1α stabilization in under hypoxic conditions [17]. Moreover, the MAP kinase inhibitor, PD98059, is able to inhibit HIF-1 transcriptional activity [18]. These data suggest that phosphorylation as well as dephosphorylation could both be involved in HIF-1 activation. Similar results have already been described for c-jun, which needs to be dephosphorylated in its DNA-binding domain and phosphorylated in its transactivation domain to be active [19].

Several kinases are known to be activated under hypoxia and are therefore possible candidates for HIF-1 activation. This is the case for some of the MAP kinase family members and for PI-3 kinase.

Section snippets

Kinases activated during hypoxia

Under hypoxic conditions, the activation of several MAP kinases has been demonstrated in different cell types and is associated with the activation of transcription factors such as AP-1 (activated protein-1, c-Jun, c-Jos, Fra-1, ATF-2), elk-1, and nuclear factor-κB (NF-κB) [7]. However, all the kinases and the transcription factors that are involved, as well as the target genes that are over-expressed, could differ according to the cell type investigated (Table 1). One of the most

HIF-1 activation depends on several kinase pathways

As previously described, HIF-1 is a phosphorylated protein and its phosphorylation is involved in HIF-1α subunit stabilization as well as in the regulation of HIF-1 transcriptional activity [16], [17], [18]. Here, we describe the role played by PI-3K/Akt and ERK in HIF-1 stabilization and activation under hypoxic conditions.

Physiological relevance of HIF-1 phosphorylation

Although other transcription factors such as AP-1, Egr-1, or NF-κB mediate the hypoxia-inducible expression of specific genes in specific cell types [9], HIF-1 appears to be unique with respect to its function as a global regulator of oxygen homeostasis. The increase in the expression of glycolytic enzymes, the glucose transporter GLUT-1, or VEGF all contribute towards the survival of cells and tissues undergoing oxygen deficiency [51]. In the process of HIF-1 activation by hypoxia,

Conclusion

The activation of HIF-1 is a multi-step process requiring HIF-1α subunit stabilization [10], [11], [12], redox-regulation [15], phosphorylation, and interaction with Hsp90 [13] and coactivator(s) [14]. Here, we present some of the recent data highlighting the activation of several kinases in hypoxic cells and their possible role in the activation of the HIF-1 transcription factor. Firstly, the PI-3 kinase/Akt pathway is involved in HIF-1α stabilization under hypoxia [30], [33], [34], [37], but

Acknowledgements

C. Michiels is Research Associate and E. Minet Research Assistant of the National Funds for Scientific Research (FNRS, Belgium). G. Michel and D. Mottet are fellows of FRIA (Fonds pour la Recherche dans I’Industrie et I’Agriculture, Belgium). This text presents results of the Belgian Programme on Interuniversity Poles of Attraction initiated by the Belgian State, Prime Minister’s Office, Science Policy Programming. The scientific responsability is assumed by its authors. This work was partly

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¹: Emmanuel Minet has obtained his Ph.D. thesis at the University of Namur. This work was focused on the transcriptional and post-translational regulation of HIF-1. He is now a post-doctoral fellow at the Dana Farber Cancer Institute in Boston.

²: Gaetan Michel is a Ph.D. student in chemistry. He has developed a 3D model of the DNA-binding domain of HIF-1 when bound to HRE. He is now studying the effect of mutations within the DNA-binding domain of HIF-1α.

³: Denis Mottet is a Ph.D. student in Biology. His research is aimed at identifying new proteins interacting with HIF-1.

⁴: Martine Raes obtained her doctorate in biological sciences in 1983 at the University of Namur, where she is now in charge of the Laboratory of Biochemistry and Cellular Biology. Her research is focused on signal transduction in inflammatory conditions and during the development of atherosclerosis.

⁵: Carine Michiels was awarded her doctorate on the protective effect of antioxidant enzymes against oxidative stress in 1989. Since then, her research interests include understanding the effects of hypoxia on cells at the transcriptional as well as on biochemical levels. She is also currently engaged in the development of new anti-ischemic molecules.

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Review articleTransduction pathways involved in Hypoxia-Inducible Factor-1 phosphorylation and activation

Abstract

Introduction

Section snippets

Kinases activated during hypoxia

HIF-1 activation depends on several kinase pathways

Physiological relevance of HIF-1 phosphorylation

Conclusion

Acknowledgements

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Review article
Transduction pathways involved in Hypoxia-Inducible Factor-1 phosphorylation and activation

Cellular and developmental control of O₂ homeostasis by hypoxia-inducible factor 1 alpha

Hypoxia-Inducible Factor-1 is a basic-helix-loop-helix-PAS hetrodimer regulated by cellular O₂ tension