The biological effects of estrogens are mediated by the estrogen receptors ER␣ and ER␤. These receptors regulate gene expression through binding to DNA enhancer elements and subsequently recruiting factors such as coactivators that modulate their transcriptional activity. Here we show that ARNT (aryl hydrocarbon receptor nuclear translocator), the obligatory heterodimerization partner for the aryl hydrocarbon receptor and hypoxia inducible factor 1␣, functions as a potent coactivator of ER␣-and ER␤-dependent transcription. The coactivating effect of ARNT depends on physical interaction with the ERs and involves the C-terminal domain of ARNT and not the structurally conserved basic helixloop-helix and PAS (Per-ARNT-Sim) motifs. Moreover, we show that ARNT͞ER interaction requires the E2-activated ligand binding domain of ER␣ or ER␤. These observations, together with the previous role of ARNT as an obligatory partner protein for conditionally regulated basic helix-loop-helix-PAS proteins like the aryl hydrocarbon receptor or hypoxia inducible factor 1␣, expand the cellular functions of ARNT to include regulation of ER␣ and ER␤ transcriptional activity. ARNT was furthermore recruited to a natural ER target gene promoter in a estrogen-dependent manner, supporting a physiological role for ARNT as an ER coactivator.cross-talk ͉ chromatin immunoprecipitation E strogens regulate important physiological processes, such as development and function of the male and female reproductive system and maintenance of bone mass in women, and represent a protective factor against cardiovascular disease (1). The physiological effects of estrogens are mediated by estrogen receptors ␣ (ER␣) and ␤ (ER␤), which belong to the nuclear receptor superfamily (1, 2). Nuclear receptors are liganddependent transcription factors characterized by a conserved structural arrangement composed of a centrally located DNA binding domain (DBD) containing two highly conserved Zn finger motifs. This domain is flanked in the N terminus by a variable A͞B region, which contains an activation function (AF-1). The ligand binding domain (LBD), located C-terminally of the DBD, contains a second AF (AF-2) and is also responsible for ligand binding, receptor dimerization, and cofactor interaction (3).The ERs are, in the absence of ligand, present in the nucleus in a nonactivated form. The latent ERs interact with corepressors such as N-Cor, SHP, or SMRT, which inhibit constitutive transcriptional activity (1). Binding of agonists induces release of repressor proteins and allows interaction with coactivators of the p160 class like SRC-1 or transcription intermediary factor 2 (TIF-2). These proteins have been shown to increase access to chromatin through acetylation of histones, mediate contact with general transcription factors, and enhance receptor AF-1͞AF-2 synergy (4, 5).Interestingly, p160 coactivators share considerable sequence homology to basic helix-loop-helix (bHLH)-PAS (Per-ARNTSim) transcription factors. This family also includes factors such as the aryl h...

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Bone tissue homeostasis is governed by hormones, growth factors, and cytokines and can be distorted by environmental pollutants, such as ligands to the aryl hydrocarbon receptor (AhR). A transgenic mouse expressing a constitutively active aryl hydrocarbon receptor (CA-AhR), mimicking continuous low-dose exposure to AhR ligands, was used to explore potential long-term effects of these ligands on bone. The density, content, and dimensions of cortical and trabecular bone, as well as physical properties, were significantly altered in female transgenic mice, while almost no alterations were detected in males. Osteoclast volume density and serum level of C-telopeptide of type I collagen (CTX), reflecting osteoclast activity, were both increased by approximately 60% in female CA-AhR mice, while serum tartrate-resistant acid phosphatase (TRAP) 5b, reflecting osteoclast numbers, was unchanged. Subsequently, the resorption index (CTX/TRAP 5b) was increased by 90%, indicating increased osteoclast activity in female CA-AhR. Moreover, the protein level of the osteoclast collagenase cathepsin K was increased by 40% in bone extracts of female CA-AhR mice. The messenger RNA expression of several osteoclast- and osteoblast-associated genes was altered in female transgenic mice but not in males. Notably, early markers for osteoclast and osteoblast differentiation were normal, while the expression of functional markers of osteoclasts and osteoblasts were reduced. In conclusion, a low continuous activation of the AhR leads to a skeletal phenotype with increased bone resorption associated with more ductile bones in females but not in males. The results indicate the presence of an interaction between the AhR and a female-specific mechanism implicated in inhibition of osteoclast development and function. Female bone tissue appears more susceptible to dioxins and other AhR ligands than male bone tissue.

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