Oxygen sensing in yeast: Evidence for the involvement of the respiratory chain in regulating the transcription of a subset of hypoxic genes

Abstract

Oxygen availability affects the transcription of a number of genes in nearly all organisms. Although the molecular mechanisms for sensing oxygen are not precisely known, heme is thought to play a pivotal role. Here, we address the possibility that oxygen sensing in yeast, as in mammals, involves a redox-sensitive hemoprotein. We have found that carbon monoxide (CO) completely blocks the anoxia-induced expression of two hypoxic genes, OLE1 and CYC7, partially blocks the induction of a third gene, COX5b, and has no effect on the expression of other hypoxic or aerobic genes. In addition, transition metals (Co and Ni) induce the expression of OLE1 and CYC7 in a concentration-dependent manner under aerobic conditions. These findings suggest that the redox state of an oxygen-binding hemoprotein is involved in controlling the expression of at least two hypoxic yeast genes. By using mutants deficient in each of the two major yeast CO-binding hemoproteins (cytochrome c oxidase and f lavohemoglobin), respiratory inhibitors, and cob1 and 0 mutants, we have found that the respiratory chain is involved in the anoxic induction of these two genes and that cytochrome c oxidase is likely the hemoprotein ''sensor.'' Our findings also indicate that there are at least two classes of hypoxic genes in yeast (CO sensitive and CO insensitive) and imply that multiple pathways͞mechanisms are involved in modulating the expression of hypoxic yeast genes.

Oxygen-regulated isoforms of cytochrome c oxidase have differential effects on its nitric oxide production and on hypoxic signaling

“…COX5a is switched off and COX5b is switched on when the O 2 concentration drops below a threshold of 0.5 M O 2 . Insofar as the full induction of COX5b under hypoxic conditions depends on a respiratory chain, which contains the Cco Va isozyme (14) when cells are shifted from normoxic to anoxic conditions, it appears that the induction of COX5b and expression of its product, Vb, constitutes a positive feedback mechanism in which hypoxia induces the expression of a gene whose product is incorporated into Cco to make it better able to produce NO. It will be interesting to determine whether a similar link can be established for the oxygen-regulated isoforms of mammalian Cco (30) and hypoxic gene induction.…”

“…This NO production is likely to be important for physiological adaptation to hypoxia, which requires the induction of several hypoxic nuclear genes (68,69). In support of NO involvement in hypoxic signaling are the findings that: (i) the respiratory chain and Cco have been implicated in the induction of hypoxic genes in both yeast and mammalian cells (13,14); (ii) NO production and tyrosine protein nitration increase in hypoxic yeast cells (21) in a respiration-dependent fashion; (iii) NO has a stabilizing effect on HIF-1␣ in mammalian cells (70), and (iv) some hypoxic yeast genes in normoxic cells are induced by exogenous NO donors (21). The finding here that the two subunit V isoforms have differential effects on the Cco NO production and on the expression of two different hypoxic genes that have been shown previously to be under the control of the respiratory chain provide further evidence for a link between Cco and hypoxic gene induction.…”

“…Previously, it has been demonstrated that induction of the hypoxic nuclear genes CYC7 and OLE1 in yeast cells requires Cco and the respiratory chain (14). It has also been shown that DETA-NO induces the expression of CYC7 in normoxic cells and that the kinetics of induction of CYC7 in hypoxic cells is accelerated when YHB1 is deleted (21).…”

“…These protein modifications may involve tyrosine nitration or the nitrosation of one or more proteins (e.g., HIF-1␣). Support for the involvement of mitochondrially produced NO in hypoxic signaling in yeast comes from the finding that: (i) an NO donor, 2,2'-(hydroxynitrohydrazono)bisethanimine (DETA-NO), induces the expression of yeast CYC7, a hypoxic nuclear gene, in normoxic cells, (ii) CYC7 is induced earlier and to higher levels in yhb1 Ϫ cells than in YHB1 wild-type cells, (iii) there is an increase in protein-specific tyrosine nitration levels in cells exposed to hypoxia (21), and (iv) neither NO production nor the hypoxic induction of respiration-dependent hypoxic genes occurs in mutants that lack Cco (14,21). This model also receives support from recent mammalian cells studies that have shown that cytochrome c, which is required for mitochondrial NO 2 Ϫ -dependent NO production, functions to stabilize HIF-1␣ in murine cell lines exposed to hypoxia (27).…”

“…The mitochondrial respiratory chain and cytochrome c oxidase (Cco) have been implicated in the induction of some hypoxic nuclear genes (hypoxic signaling) in both yeast and mammalian cells (13)(14)(15)(16) exposed to reduced oxygen levels, and it has been proposed that mitochondrially generated ROS are involved (17)(18)(19)(20). However, recent studies have revealed that ROS are not sufficient for the induction of hypoxic nuclear genes (R. Dirmeier and R.O.P., unpublished work), suggesting that the respiratory chain has an additional role in hypoxic signaling (21).…”