Perspectives on Oxygen Sensing

Further progress in this field would be greatly facilitated by genetic approaches utilizing somatic cells and/or model organisms. This is the first major contribution of the recent study by Wingrove and O'Farrell 1999xWingrove, J.A and O'Farrell, P.H. Cell. 1999; 98: 105–114Abstract | Full Text | Full Text PDF | PubMed | Scopus (178)See all ReferencesWingrove and O'Farrell 1999 that utilized Drosophila melanogaster as a model organism to investigate molecular mechanisms of O2 homeostasis. The second major contribution is their demonstration of a role for cGMP-dependent protein kinase (PKG) and/or nitric oxide synthase (NOS), using genetic and pharmacologic approaches in a remarkable spectrum of behavioral, cellular, developmental, and survival responses to hypoxia in this organism. Their data indicate that both acute and chronic responses to hypoxia are impaired in flies carrying a mutation in the gene encoding PKG or in flies exposed to the NOS inhibitor L-NAME, and these responses are amplified in transgenic flies that overexpress NOS. These results will stimulate interest in the use of model organisms to genetically dissect hypoxia signal transduction pathways and will also focus attention on the role of NO in mammalian O2 sensing.Can any connections be made between available data from Drosophila and mammalian cells? Analysis of isolated rat liver mitochondria (Giulivi 1998xGiulivi, C. Biochem. J. 1998; 332: 673–679CrossRef | PubMedSee all ReferencesGiulivi 1998) and cultured endothelial cells (Clementi et al. 1999xClementi, E, Brown, G.C, Foxwell, N, and Moncada, S. Proc. Natl. Acad. Sci. USA. 1999; 96: 1559–1562CrossRef | PubMed | Scopus (242)See all ReferencesClementi et al. 1999) revealed that as O2 concentration was reduced, the rate of O2 consumption declined, whereas such an effect was not observed in the presence of a NOS inhibitor, suggesting that constitutive low-level mitochondrial production of NO may result in competitive inhibition of cytochrome oxidase at low O2 concentrations. This is intriguing given the discovery of a novel mitochondrial NOS (mtNOS) isoform in rat liver (Giulivi 1998xGiulivi, C. Biochem. J. 1998; 332: 673–679CrossRef | PubMedSee all ReferencesGiulivi 1998 and references therein). These results have obvious implications for the mitochondrial model of O2 sensing (Figure 1DFigure 1D). NO also binds to and activates the hemoprotein guanylate cyclase, which produces cGMP, the allosteric activator of PKG. In mammalian vascular smooth muscle cells, O2− produced by NADH oxidase inhibits activation of guanylate cyclase by NO (Gupte et al. 1999xGupte, S.A, Rupawalla, T, Mohazzab-H, K.M, and Wolin, M.S. Am. J. Physiol. 1999; 276: H1535–H1542PubMedSee all ReferencesGupte et al. 1999). The PKG-dependent and -independent effects of NO on O2 homeostasis in Drosophila (Wingrove and O'Farrell 1999xWingrove, J.A and O'Farrell, P.H. Cell. 1999; 98: 105–114Abstract | Full Text | Full Text PDF | PubMed | Scopus (178)See all ReferencesWingrove and O'Farrell 1999) may therefore result from NO binding to guanylate cyclase and cytochrome oxidase, respectively (Figure 2Figure 2). This model provides an interesting parallel to the two mammalian models (Figure 1CFigure 1C and Figure 1DFigure 1D).Figure 2A Model for Hypoxia Signal Transduction Pathways in DrosophilaAlthough both pathways are NOS dependent, one is PKG independent (left) whereas the other is PKG dependent (right). Both pathways can be inhibited by either L-NAME or O2. The two models are similar to those proposed for mammalian cells (Figure 1Figure 1): the PKG-independent and -dependent pathways involve cytochrome oxidase and NAD(P)H oxidase, respectively.View Large Image | View Hi-Res Image | Download PowerPoint SlideInhibitors of NOS, guanylate cyclase, and PKG have been shown to decrease EPO expression in hypoxic human cells (Ohigashi et al. 1993xOhigashi, T, Brookins, J, and Fisher, J.W. J. Clin. Invest. 1993; 92: 1587–1591CrossRef | PubMedSee all ReferencesOhigashi et al. 1993), but the effects were modest. However, the NO/CO/cGMP pathway intersects with the hypoxia signal transduction pathway at multiple levels in mammalian cells, which may confound such analyses. Physiologic levels of NO and CO inhibit HIF-1 DNA binding activity (13xLiu, Y, Christou, H, Morita, T, Laughner, E, Semenza, G.L, and Kourembanas, S. J. Biol. Chem. 1998; 273: 15257–15262CrossRef | PubMed | Scopus (189)See all References, 17xSogawa, K, Numayama-Tsuruta, K, Ema, M, Abe, M, Abe, H, and Fujii-Kuriyama, Y. Proc. Natl. Acad. Sci. USA. 1998; 95: 7368–7373CrossRef | PubMed | Scopus (180)See all References) and pharmacologic levels also inhibit induction of HIF-1α protein expression and transactivation function (Huang et al. 1999xHuang, L.E, Willmore, W.G, Gu, J, Goldberg, M.A, and Bunn, H.F. J. Biol. Chem. 1999; 274: 9038–9044CrossRef | PubMed | Scopus (238)See all ReferencesHuang et al. 1999) possibly via inhibition of ETC complex I. Finally, HIF-1 activates expression of genes encoding inducible NOS and heme oxygenase 1 resulting in the production of NO and CO, respectively (reviewed by Semenza 1998xSemenza, G.L. Curr. Opin. Genet. Dev. 1998; 8: 588–594CrossRef | PubMed | Scopus (678)See all ReferencesSemenza 1998). The further characterization and cloning of mammalian mtNOS and analysis of knockout mouse ES cells lacking expression of this or other NOS isoforms (or other components of the guanylate cyclase/PKG axis) may provide additional insights into mechanisms of O2 sensing.Whereas flies express only a single NOS isoform, mammals express three, and possibly four, different isoforms, which illustrates a potential limitation of utilizing invertebrates to understand mammalian biology. This difference in relative complexity is also observed with respect to development and physiology. Whereas Drosophila develop tracheal tubes that allow direct diffusion of O2 to all cells, the greatly increased size of mammals necessitated development of complex circulatory and respiratory systems for O2 delivery. It is likely that hypoxia signal transduction in mammals will also be more complex than in Drosophila. Just as we now appreciate that O2 sensing is not restricted to specialized chemoreceptor cell types, the paradigm of a monolithic O2 sensor responsible for initiating biological responses to hypoxia may represent an oversimplification. Given the fundamental importance of O2 homeostasis for survival, multiple hypoxia signal-transduction and modulatory cross-talk pathways may eventually be identified. For now, understanding at least one of these pathways remains a formidable challenge.