Cross-talk between the Aryl Hydrocarbon Receptor and Hypoxia Inducible Factor Signaling Pathways

Abstract

The aryl hydrocarbon receptor (AHR) and the ␣-class hypoxia inducible factors (HIF1␣, HIF2␣, and HIF3␣) are basic helix-loop-helix PAS (bHLH-PAS) proteins that heterodimerize with ARNT. In response to 2,3,7,8-tetrachlorodibenzo-p-dioxin, the AHR⅐ARNT complex binds to "dioxin responsive enhancers" (DREs) and activates genes involved in the metabolism of xenobiotics, e.g. cytochrome P4501A1 (Cyp1a1). The HIF1␣⅐ARNT complex binds to "hypoxia responsive enhancers" and activates the transcription of genes that regulate adaptation to low oxygen, e.g. erythropoietin (Epo). We postulated that activation of one pathway would inhibit the other due to competition for ARNT or other limiting cellular factors. Using pathway specific reporters in transient transfection assays, we observed that DRE driven transcription was markedly inhibited by hypoxia and that hypoxia responsive enhancer driven transcription was inhibited by AHR agonists. When we attempted to support this cross-talk model using endogenous loci, we observed that activation of the hypoxia pathway inhibited Cyp1a1 up-regulation, but that activation of the AHR actually enhanced the induction of Epo by hypoxia. To explain this unexpected additivity, we examined the Epo gene and found that its promoter harbors DREs immediately upstream of its transcriptional start site. These experiments outline conditions where inhibitory and additive cross-talk occur between the hypoxia and dioxin signal transduction pathways and identify Epo as an AHR-regulated gene. The AHR1 regulates a variety of biological responses to environmentally ubiquitous polycyclic aromatic hydrocarbons and dioxins (1,2). In what can be defined as an adaptive pathway, the AHR up-regulates a battery of XMEs that often metabolize many of these agonists to more soluble and excretable products. A classic example of this pathway is observed upon exposure to benzo [a]pyrene. This chemical binds to the AHR leading to the up-regulation of a battery of genes including Cyp1a1, Cyp1a2, and Cyp1b1 (3). The enzymes encoded by these loci have metabolic activity toward benzo[a]pyrene and thus play an important role in its elimination (4). At present, we understand many of the molecular events in what appears to be an adaptive response to polycyclic aromatic hydrocarbon exposure. In brief, the up-regulation of genes like Cyp1a1 are regulated by an agonist-induced heterodimerization between two bHLH-PAS proteins, the AHR and ARNT (5, 6). This heterodimeric pair interacts with DREs upstream of the regulated promoters leading to an increase in their transcription rate and a resultant increase in XME activity (7).Although we have developed models to describe how the AHR regulates the expression of XMEs, we still have very little knowledge about how this protein mediates the toxicity of potent agonists like dioxin. The molecular mechanisms of dioxin-induced effects like lymphoid involution, epithelial hyperplasia, tumor promotion, teratogenesis, or even death remain unclear. Moreover, although genetic studies indicate the ...

Cobaltous chloride and hypoxia inhibit aryl hydrocarbon receptor-mediated responses in breast cancer cells

“…Hypoxia also induces proteasomedependent degradation of ERα resulting in decreased E2-responsiveness ; however, we also observed that inhibitory AhR-ERα crosstalk was significantly decreased in breast cancer cells grown under hypoxic conditions (Stoner, 2002), and this is consistent with other reports showing that hypoxia decreases Ah-responsiveness (Chan et al, 1999;Gradin et al, 1996;Kim et al, 2000;Pollenz et al, 1999;Prasch et al, 2004). However, the mechanisms of hypoxia-dependent loss of Ah-responsiveness are unclear.…”

“…It has previously been suggested that HIF-1α or other proteins that competitively displace the AhR from binding Arnt may that decrease Ah-responsiveness in cells grown under hypoxia (Chan, Yao, Gu, and Bradfield, 1999;Gradin, McGuire, Wenger, Kvietikova, fhitelaw, Toftgard, Tora, Gassmann, and Poellinger, 1996;Kim and Sheen, 2000;Pollenz, Davarinos, and Shearer, 1999;Prasch, Andreasen, Peterson, and Heideman, 2004). However, we show that overexpression of Arnt in transfection assays (Fig.…”

“…Previous studies have also reported that hypoxia decreases Ah-responsiveness (Chan, Yao, Gu, and Bradfield, 1999;Gradin, McGuire, Wenger, Kvietikova, fhitelaw, Toftgard, Tora, Gassmann, and Poellinger, 1996;Kim and Sheen, 2000;Pollenz, Davarinos, and Shearer, 1999;Prasch, Andreasen, Peterson, and Heideman, 2004), and it has been suggested (Kim and Sheen, 2000) that this may be due to overexpression of HIF-1α which, like the AhR, competitively binds Arnt. Overexpression of HIF1α under hypoxia may competitively decrease availability of Arnt and thereby decrease AhR complex formation.…”

“…Since hypoxic conditions could limit the effectiveness of SAhRMs, we further investigated the mechanisms of hypoxia-induced modulation of Ahresponsiveness in breast cancer cells. Previous studies have also reported the effects of hypoxia on AhR function and their results are both complementary and different (Chan, Yao, Gu, and Bradfield, 1999;Gradin, McGuire, Wenger, Kvietikova, fhitelaw, Toftgard, Tora, Gassmann, and Poellinger, 1996;Kim and Sheen, 2000;Pollenz, Davarinos, and Shearer, 1999;Prasch, Andreasen, Peterson, and Heideman, 2004). Some data suggest that overexpression of HIF-1α under hypoxic conditions may decrease Ah-responsiveness through competition with the AhR for binding Arnt (Kim and Sheen, 2000).…”

“…Some data suggest that overexpression of HIF-1α under hypoxic conditions may decrease Ah-responsiveness through competition with the AhR for binding Arnt (Kim and Sheen, 2000). There is also evidence that overexpression of HIF-1α can decrease AhR:Arnt binding to DNA in a gel mobility shift assay (Chan, Yao, Gu, and Bradfield, 1999). In this study, we also show that hypoxia clearly decreases induction of CYP1A1, EROD activity, and AhRR by TCDD and blocks inhibitory AhR-ERα/Sp1 crosstalk in cells transfected with pVEGF1 ( Figs.…”