Timing is everything: Consequences of transient and sustained AhR activity

The aryl hydrocarbon receptor (AhR) was implicated as a mediator of xenobiotic toxicity over three decades ago. Although a complete picture continues to elude us, investigations by many laboratories during the ensuing period have revealed much about AhR biology in normal physiological processes, as well as the toxicities induced by the dioxins and related polychlorinated aromatic hydrocarbons. The findings are captured in numerous excellent reviews. This commentary attempts to inject a new perspective on some new as well as frequently overlooked observations in the context of established receptor properties. Specifically, we examine the impact of transient versus sustained receptor activation on AhR biology, and explore the potential role for cytochrome P450 expression in regulating AhR activity amongst various tissues. The growing recognition that AhR action functions through multiple mechanisms serves to further highlight the importance of limiting prolonged receptor activation.     

Reduction of androgen receptor expression by benzo[alpha]pyrene and 7,8-dihydro-9,10-epoxy-7,8,9,10-tetrahydrobenzo[alpha]pyrene in human lung cells

5Alpha-dihydrotestosterone significantly increased cell growth of lung adenocarcinoma cell line H1355. Benzo[alpha]pyrene (BaP) was a pulmonary carcinogen found in cigarette smoke. Treatment with 1microM BaP tremendously reduced constitutive androgen receptor (AR) expression, as determined with Western immunoblotting and the real-time RT-PCR assay, as well as testosterone-induced AR protein levels in H1355 cells. Similarly, 1microM BaP significantly reduced AR mRNA levels in human bronchial epithelial cells BEAS-2B. Although BaP, 2,3,7,8-tetrachlorodibenzo-p-dixin and polychlorinated biphenyl 126 activated aryl hydrocarbon receptor (AhR), which subsequently induced cytochrome P4501A1 (CYP1A1) and P4501B1 (CYP1B1) expression in H1355 cells, unexpectedly, neither TCDD nor PCB126 reduced AR expression. Antagonizing AhR activation and cytochrome P4501 activity with alpha-naphthoflavone, or inhibiting CYP1B1 activity with 2,4,3',5'-tetramethoxystilbene, however, prevented BaP-induced AR reduction. Furthermore, 7,8-dihydro-9,10-epoxy-7,8,9,10-tetrahydrobenzo[alpha]pyrene, a BaP carcinogenic metabolite catalyzed by CYP1A1 and CYP1B1, significantly reduced AR expression in H1355 cells and human lung fibroblasts WI-38. This was the first study that reports that BaP and BPDE reduced endogenous AR expression. These data suggest that metabolically activated BaP may disrupt androgen function by reducing AR levels in androgen-responsive organs.     

Association of Cullin1 haplotype variants with rheumatoid arthritis and response to methotrexate

Aberrations in ubiquitin pathway have been implicated in many diseases and drug response. In a previous study on rheumatoid arthritis (RA) in Japanese population, significant association of Cullin1 gene (CUL1), an ubiquitin E3 ligase, was observed. CUL1 also mediates degradation of IκBα and p27, levels of which has been associated with RA etiology and drug response, respectively. We carried out a replication study of association of CUL1 polymorphisms with RA in a north Indian population. Allelic, genotypic, and haplotypic associations of a promoter and two intronic polymorphisms of CUL1 with RA and with methotrexate response in patients with RA, were tested. A significant association (P=0.00056, adjusted) of a haplotype A-T-T with RA (odds ratio=3.68; 95% confidence interval=1.86-7.27) and in patients with RA poorly responding to methotrexate treatment (P=0.04, adjusted) was observed. Association with CUL1 haplotype indicates a possible role of CUL1 variation(s) in RA and its response to methotrexate.     

Differential gene expression in response to methoxychlor and estradiol through ERalpha, ERbeta, and AR in reproductive tissues of female mice.

The reproductive and developmental effects of 17beta-estradiol (E2) and methoxychlor (MXC) observed in treated rodents appear to be linked to some unique but also overlapping patterns of gene expression. The MXC metabolite 2,2-bis(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE) was previously shown to have selective agonist activity through estrogen receptor alpha (ERalpha) and antagonist activity through ERbeta and androgen receptor (AR). To discover gene families regulated by HPTE and E2, and to characterize similarities and differences in patterns of gene expression induced by these selective ER ligands, we analyzed tissues from mice treated for 3 days with a combined treatment of E2 and HPTE (E2 + HPTE), or the antiandrogen flutamide (FLU). RNA from uteri and ovaries was analyzed with cDNA microarrays and real-time RT-PCR. Results indicate that HPTE and E2 acted similarly to regulate most gene families in the uterus, which expresses predominantly ERalpha. However, in both the uterus and the ovary, there were a few genes that displayed differential patterns of gene regulation by E2 or HPTE treatment, presumably through ERbeta, AR, or other unidentified pathways. In the uterus, progesterone receptor, ERalpha, AR, insulin-like growth factor 1, insulin-like growth factor binding protein 5, and clusterin mRNAs were significantly reduced with both E2 or HPTE treatments, whereas cathepsin B was induced. Conversely, in the ovary, induction of cathepsin B by E2 was reversed after cotreatment with HPTE, and ERbeta expression was induced similarly by HPTE and FLU but not by E2. In addition, E2 uniquely regulated glutathione peroxidase 3, glutathione S-transferase, and cytochrome P450 17alpha-hydroxylase, with no effect of HPTE or FLU treatments. This analysis demonstrated several gene families that appear to be regulated in a ligand-specific pattern, which may explain the unique but overlapping reproductive tissue pathologies following exposure to E2 and MXC.     

Effects of tryptophan photoproducts in the circadian timing system: searching for a physiological role for aryl hydrocarbon receptor.

The aryl hydrocarbon receptor (AhR) mediates adverse effects of dioxins, but its physiological role remains ambiguous. The similarity between AhR and canonical circadian clock genes suggests potential involvement of AhR in regulation of circadian timing. Photoproducts of tryptophan (TRP), including 6-formylindolo[3,2-b]carbazole (FICZ), have high affinity for AhR and are postulated as endogenous ligands. Although TRP photoproducts activate AhR signaling in vitro, their effects in vivo have not been investigated in mammals. Because TRP photoproducts may act as transducers of light, we examined their effects on the circadian clock. Intraperitoneal injection of TRP photoproducts or FICZ to C57BL/6J mice dose dependently induced AhR downstream targets, cytochrome P4501A1 (CYP1A1) and cytochrome P4501B1 mRNA expression, in liver. c-fos mRNA, a commonly used marker for light responses, was also induced with FICZ, and all responses were AhR dependent. A rat-immortalized suprachiasmatic nucleus (SCN) cell line, SCN 2.2, was used to examine the direct effect of TRP photoproducts on the molecular clock. Both TRP photoproducts and FICZ-increased CYP1A1 expression and prolonged FICZ incubation altered the circadian expression of clock genes (Per1, Cry1, and Cry2) in SCN 2.2 cells. Furthermore, FICZ inhibited glutamate-induced phase shifting of the mouse SCN electrical activity rhythm. Circadian light entrainment is critical for adjustment of the endogenous rhythm to environmental light cycle. Our results reveal a potential for TRP photoproducts to modulate light-dependent regulation of circadian rhythm through triggering of AhR signaling. This may lead to further understanding of toxicity of dioxins and the role of AhR in circadian rhythmicity.     

Fitting a xenobiotic receptor into cell homeostasis: how the dioxin receptor interacts with TGFbeta signaling

As our knowledge on the mechanisms that control cell function increases, more complex signaling pathways and quite intricate cross-talks among regulatory proteins are discovered. Establishing accurate interactions between cellular networks is essential for a healthy cell and different alterations in signaling are known to underline human disease. Transforming growth factor beta (TGFbeta) is an extracellular cytokine that regulates such critical cellular responses as proliferation, apoptosis, differentiation, angiogenesis and migration, and it is assumed that the latency-associated protein LTBP-1 plays a relevant role in TGFbeta targeting and activation in the extracellular matrix (ECM). The dioxin receptor (AhR) is a unique intracellular protein long studied because of its critical role in xenobiotic-induced toxicity and carcinogenesis. Yet, a large set of studies performed in cellular systems and in vivo animal models have suggested important xenobiotic-independent functions for AhR in cell proliferation, differentiation and migration and in tissue homeostasis. Remarkably, AhR activity converges with TGFbeta-dependent signaling through LTBP-1 since cells lacking AhR expression have phenotypic alterations that can be explained, at least in part, by the coordinated regulation of both proteins. Here, we will discuss the existence of functional interactions between AhR and TGFbeta signaling. We will focus on regulatory and functional aspects by analyzing how AhR status determines TGFbeta activity and by proposing a mechanism through which LTBP-1, a novel AhR target gene, mediates such effects. We will integrate ECM proteases in the AhR-LTBP-1-TGFbeta axis and suggest a model that could help explain some in vivo phenotypes associated to AhR deficiency.