Environmental contaminants affect a wide variety of biological events in many species. Dioxins are typical environmental contaminants that exert adverse oestrogen-related effects. Although their anti-oestrogenic actions are well described, dioxins can also induce endometriosis and oestrogen-dependent tumours, implying possible oestrogenic effects. However, the molecular mechanism underlying oestrogen-related actions of dioxins remains largely unknown. A heterodimer of the dioxin receptor (AhR) and Arnt, which are basic helix-loop-helix/PAS-family transcription factors, mediates most of the toxic effects of dioxins. Here we show that the agonist-activated AhR/Arnt heterodimer directly associates with oestrogen receptors ER-alpha and ER-beta. This association results in the recruitment of unliganded ER and the co-activator p300 to oestrogen-responsive gene promoters, leading to activation of transcription and oestrogenic effects. The function of liganded ER is attenuated. Oestrogenic actions of AhR agonists were detected in wild-type ovariectomized mouse uteri, but were absent in AhR-/- or ER-alpha-/- ovariectomized mice. Our findings suggest a novel mechanism by which ER-mediated oestrogen signalling is modulated by a co-regulatory-like function of activated AhR/Arnt, giving rise to adverse oestrogen-related actions of dioxin-type environmental contaminants.

Transcriptional activators, several different coactivators, and general transcription factors are necessary to access specific loci in the dense chromatin structure to allow precise initiation of RNA polymerase II (Pol II) transcription. Histone acetyltransferase (HAT) complexes were implicated in loosening the chromatin around promoters and thus in gene activation. Here we demonstrate that the 2 MDa GCN5 HAT-containing metazoan TFTC/STAGA complexes contain a histone H2A and H2B deubiquitinase activity. We have identified three additional subunits of TFTC/STAGA (ATXN7L3, USP22, and ENY2) that form the deubiquitination module. Importantly, we found that this module is an enhancer of position effect variegation in Drosophila. Furthermore, we demonstrate that ATXN7L3, USP22, and ENY2 are required as cofactors for the full transcriptional activity by nuclear receptors. Thus, the deubiquitinase activity of the TFTC/STAGA HAT complex is necessary to counteract heterochromatin silencing and acts as a positive cofactor for activation by nuclear receptors in vivo.

Renal 25-hydroxyvitamin D3 1alpha-hydroxylase [1alpha(OH)ase] catalyzes metabolic activation of 25-hydroxyvitamin D3 into 1alpha, 25-dihydroxyvitamin D3 [1alpha,25(OH)2D3], an active form of vitamin D, and is inhibited by 1alpha,25(OH)2D3. 1alpha(OH)ase, which was cloned from the kidney of mice lacking the vitamin D receptor (VDR-/- mice), is a member of the P450 family of enzymes (P450VD1alpha). Expression of 1alpha(OH)ase was suppressed by 1alpha, 25(OH)2D3 in VDR+/+ and VDR+/- mice but not in VDR-/- mice. These results indicate that the negative feedback regulation of active vitamin D synthesis is mediated by 1alpha(OH)ase through liganded VDR.

One class of the nuclear receptor AF-2 coactivator complexes contains the SRC-1/TIF2 family, CBP/p300 and an RNA coactivator, SRA. We identi®ed a subfamily of RNA-binding DEAD-box proteins (p72/p68) as a human estrogen receptor a (hERa) coactivator in the complex containing these factors. p72/p68 interacted with both the AD2 of any SRC-1/TIF2 family protein and the hERa A/B domain, but not with any other nuclear receptor tested. p72/p68, TIF2 (SRC-1) and SRA were co-immunoprecipitated with estrogenbound hERa in MCF7 cells and in partially puri®ed complexes associated with hERa from HeLa nuclear extracts. Estrogen induced co-localization of p72 with hERa and TIF2 in the nucleus. The presence of p72/ p68 potentiated the estrogen-induced expression of the endogenous pS2 gene in MCF7 cells. In a transient expression assay, a combination of p72/p68 with SRA and one TIF2 brought an ultimate synergism to the estrogen-induced transactivation of hERa. These ®nd-ings indicate that p72/p68 acts as an ER subtypeselective coactivator through ERa AF-1 by associating with the coactivator complex to bind its AF-2 through direct binding with SRA and the SRC-1/TIF2 family proteins.

Inactivating mutations in the 25-hydroxyvitamin D3 1alpha-hydroxylase gene are a cause of pseudovitamin D-deficiency rickets.

Wnt and estrogen signaling represent important regulatory pathways, each controlling a wide range of biological processes. While an increasing number of observations suggest potential convergence between these pathways, no direct evidence of their functional interaction has been reported. Using human colon and breast cancer cells, we found that estrogen receptor (ER) ␣-and ␤-catenin precipitated within the same immunocomplexes, reciprocally enhanced the transactivation of cognate reporter genes, and were reciprocally recruited to cognate response elements in the promoters of endogenous target genes. Using transgenic Drosophila that ectopically expressed human ER␣ alone or together with metabolically stable ␤-catenin/Armadillo mutants, we demonstrated genetic interaction between these signal transducers in vivo. Thus, we present here the first direct evidence of cross-talk between Wnt and estrogen signaling pathways via functional interaction between ␤-catenin and ER␣.

No abstract

For the transactivation function of VDR, only the ligand-binding domain (E region) is thought to be responsible in a ligandbinding-dependent way (27), although two transactivation domains, one at the N terminus (AF-1) and one at the C terminus (AF-2), are present in most nuclear receptors. To achieve ligand-induced transactivation, the nuclear receptors recruit several nuclear receptor coactivators. They include members of the SRC-1/TIF2 family (38, 48), CBP/p300 (9, 29), and RIP140 (8). Members of the SRC-1/TIF2 family [SRC-1 (p160, ERAP160) (18, 23), TIF2 (Grip-1) (10), and AIB-1 (ACTR) (3)] mediate the function of the AF-2 of the nuclear receptors, and the interaction site has been mapped to the minimal activation domain (AD) of AF-2 (13, 23, 50). Interestingly, it was recently shown that the interactions of estrogen receptor with SRC-1 or TIF2 are induced by estrogen (E 2 ) but not by its antagonists, tamoxifen and ICI164,384. These findings indicate that the structure of the ligand-bound E region recruiting coactivators is ligand specific (18). This idea is further supported by recent findings from crystallographic analysis that the position of the AF-2 AD (helix 12) in the estrogen receptor E region which binds the E 2 antagonists clearly differs from the one which binds E 2 (17). From the structural similarity of the ligand-binding domains of nuclear receptors, ligand type-specific alterations in their structures, at least in the helix 12 positions, are postulated (7).Several synthetic 1␣,25(OH) 2 D 3 derivatives, such as F 6 -1␣,25-(OH) 2 D 3 [26,26,26,27,27,25(OH) OCT (25,40,44). Taking these facts together, it is reasonable to speculate that the selective interactions of VDR with coactivators induced by vitamin D analogs specify the biological activities of the vitamin D analogs. Such differential combinations of transcription factors and coactivators are believed to activate only particular sets of target gene promoters (46).To test this possibility, we studied the interaction of vitamin D analog-bound VDR with nuclear receptor coactivators and interacting factors. We found that although the in vivo and in vitro interactions of VDR with SRC-1, TIF2, and AIB-1 were induced by F 6 -1␣,25(OH) 2 D 3 and ED-71 as well as by 1␣,25 (OH) 2 D 3 , OCT induced interaction only with TIF2. Such interactions were also observed in the VDR-RXR heterodimer bound to DNA. Consistent with the interactions, only TIF2 potentiated the transactivation function of VDR bound to OCT. Thus, the present findings suggest that the VDR structure is altered in a vitamin D analog-specific way, resulting in selective interaction of VDR with coactivators. Such selective coactivator interaction with VDR may specify the array of biological actions of a vitamin D analog such as OCT, possibly through activating a particular set of target gene promoters. MATERIALS AND METHODSYeast two-hybrid system and ␤-galactosidase assay. The pGBT9(GAL4-DBD)-VDR(DEF) fusion plasmid was constructed by inserting rat VDR-DEF regions (encoding amino acids ...