Identification of a new class of steroid hormone receptors

The gonads and adrenal glands produce steroids classified into five major groups which include the oestrogens, progestins, androgens, glucocorticoids and mineralocorticoids. Gonadal steroids control the differentiation and growth of the reproductive system, induce and maintain sexual characteristics and modulate reproductive behaviour. Adrenal steroids also influence differentiation as well as being metabolic regulators. The effects of each steroid depend primarily on its specific receptors, the nature of which could therefore provide a basis for classification of steroid hormone action. The successful cloning, sequencing and expression of the human glucocorticoid (hGR) (ref. 1), oestrogen (hER), progesterone (hPR), and mineralocorticoid (hMR) receptors, complementary DNA, plus homologues from various species, provides the first opportunity to study receptor structure and its influence on gene expression. Sequence comparison and mutational analysis show structural features common to all groups of steroid hormone receptors. The receptors share a highly conserved cysteine-rich region which functions as the DNA-binding domain. This common segment allows the genome to be scanned for related gene products: hMR cDNA for example, was isolated using an hGR hybridization probe. In this study, using the DNA-binding domain of the human oestrogen receptor cDNA as a hybridization probe, we have isolated two cDNA clones encoding polypeptides with structural features suggestive of cryptic steroid hormone receptors which could participate in a new hormone response system.     

A novel steroid thyroid hormone receptor-related gene inappropriately expressed in human hepatocellular carcinoma

We have previously isolated from a human hepatocellular carcinoma a hepatitis B virus integration in a 147-base-pair cellular DNA fragment, similar to steroid- and c-erb-A/thyroid-hormone receptor genes. We have now cloned the corresponding complementary DNA from a human-liver cDNA library. Nucleotide sequence analysis revealed that the overall structure of the cellular gene, which we have named hap, is similar to that of the DNA-binding hormone receptors. That is, it displays two highly conserved regions identified as the putative DNA-binding and hormone-binding domains of the c-erb A/steroid receptors. Six out of seven hepatoma and hepatoma-derived cell-lines express a 2.5-kilobase (kb) hap messenger RNA species which is undetectable in normal adult and fetal livers but present in all non-hepatic tissues analysed. The data suggest that the hap gene product may be a novel ligand-responsive regulatory protein whose inappropriate expression in liver may relate to the hepatocellular carcinogenesis.     

Requirement of hormone for thermal conversion of the glucocorticoid receptor to a DNA-binding state

A central question arising from the model of eukaryotic gene regulation by steroid hormone receptors is whether or not proteins represent pre-existing gene regulatory proteins that are activated on exposure to the extracellular signal. It has been generally believed that the ligand-binding of steroid hormone receptors triggers an allosteric change in receptor structure, manifested by an increased affinity of the receptor for DNA in vitro and nuclear target elements in vivo, as monitored by nuclear translocation. But this model has been challenged by recent reports indicating that glucocorticoid and progesterone receptors bind specifically in vitro to target DNA sequences even in the absence of hormone. On the other hand, it appears that the hormone induces protection in vivo of the glucocorticoid response element of the tyrosine amino transferase gene. Here we show that under conditions permitting minimal in vitro manipulation, the steroid-free glucocorticoid receptor in crude cytosol associates with the hsp90 heat shock protein (relative molecular mass Mr approximately equal to 90,000) to form a large 300K complex, rather than the 94K liganded receptor monomer. More importantly, we have developed an assay to demonstrate the requirement of hormone to dissociate the 300K complex by heat treatment. Specific DNA-binding activity of the receptor becomes apparent in this process, showing that DNA binding occurs but is inhibited in the large heteromeric complex. We propose a model in which receptor function is repressed by association of the receptor with hsp90. Dissociation of this complex is induced by the binding of steroid and is apparently an irreversible process.     

Structural basis of steroid hormone perception by the receptor kinase BRI1

Polyhydroxylated steroids are regulators of body shape and size in higher organisms. In metazoans, intracellular receptors recognize these molecules. Plants, however, perceive steroids at membranes, using the membrane-integral receptor kinase BRASSINOSTEROID INSENSITIVE 1 (BRI1). Here we report the structure of the Arabidopsis thaliana BRI1 ligand-binding domain, determined by X-ray diffraction at 2.5?? resolution. We find a superhelix of 25 twisted leucine-rich repeats (LRRs), an architecture that is strikingly different from the assembly of LRRs in animal Toll-like receptors. A 70-amino-acid island domain between LRRs 21 and 22 folds back into the interior of the superhelix to create a surface pocket for binding the plant hormone brassinolide. Known loss- and gain-of-function mutations map closely to the hormone-binding site. We propose that steroid binding to BRI1 generates a docking platform for a co-receptor that is required for receptor activation. Our findings provide insight into the activation mechanism of this highly expanded family of plant receptors that have essential roles in hormone, developmental and innate immunity signalling.     

Primary structure and expression of a functional human glucocorticoid receptor cDNA

Identification of complementary DNAs encoding the human glucocorticoid receptor predicts two protein forms, of 777 (alpha) and 742 (beta) amino acids, which differ at their carboxy termini. The proteins contain a cysteine/lysine/arginine-rich region which may define the DNA-binding domain. Pure radiolabelled glucocorticoid receptor, synthesized in vitro, is immunoreactive and possesses intrinsic steroid-binding activity characteristic of the native glucocorticoid receptor.     

Characterization of DNA sequences through which cadmium and glucocorticoid hormones induce human metallothionein-IIA gene

Deletion experiments have defined two stretches of DNA (genetic elements), lying close to the promoter for a human gene for metallothionein, that separately mediate the induction of the gene by heavy metal ions, particularly cadmium, and by glucocorticoid hormones. The element responsible for induction by cadmium is duplicated, yet a single copy is fully functional; the element responsible for induction by glucocorticoid hormones is coincident with the DNA-binding site for the glucocorticoid hormone receptor.     

Hepatitis B virus DNA integration in a sequence homologous to v-erb-A and steroid receptor genes in a hepatocellular carcinoma

Hepatitis B virus (HBV) is clearly involved in the aetiology of human hepatocellular carcinoma (HCC) and the finding of HBV DNA integration into human liver DNA in almost all HCCs studied suggested that these integrated viral sequences may be involved in liver oncogenesis. Several HBV integrations in different HCCs and HCC-derived cell lines have been analysed after molecular cloning without revealing any obvious role for HBV. From a comparison of a HBV integration site present in a particular HCC with the corresponding unoccupied site in the non-tumorous tissue of the same liver, we now report that HBV integration places the viral sequence next to a liver cell sequence which bears a striking resemblance to both an oncogene (v-erb-A) and the supposed DNA-binding domain of the human glucocorticoid receptor and human oestrogen receptor genes. We suggest that this gene, usually silent or transcribed at a very low level in normal hepatocytes, becomes inappropriately expressed as a consequence of HBV integration, thus contributing to the cell transformation.