9-cis retinoic acid stereoisomer binds and activates the nuclear receptor RXR alpha.

Vitamin A (retinol) and its natural derivatives are required for many physiological processes. The activity of retinoids is thought to be mediated by interactions with two subfamilies of nuclear retinoic acid receptors, RAR and RXR. The RARs bind all-trans retinoic acid (t-RA) with high affinity and alter gene expression as a consequence of this direct ligand interaction. RXR alpha is activated by t-RA, yet has little binding affinity for this ligand. t-RA may be converted to a more proximate ligand that directly binds and activates RXR alpha, and we have developed a method of nuclear receptor-dependent ligand trapping to test this hypothesis. Here we report the identification of a stereoisomer of retinoic acid, 9-cis retinoic acid, which directly binds and activates RXR alpha. These results suggest a new role for isomerization in the physiology of natural retinoids.     

Homodimer formation of retinoid X receptor induced by 9-cis retinoic acid.

Retinoid response pathways are mediated by two classes of receptors, the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs). A central question is whether distinct response pathways are regulated by these two classes of receptors. The observation that the stereoisomer 9-cis-retinoic acid binds with high affinity to RXRs suggested that this retinoid has a distinct role in controlling RXR activity, but it was almost simultaneously discovered that RXRs function as auxiliary receptors for RARs and related receptors, and are essential for DNA binding and function of those receptors. Hence, although RARs seem to operate effectively only as heterodimeric RAR/RXR complexes, RXRs themselves apparently function predominantly, if not exclusively, as auxiliary receptors. Here we report that 9-cis-retinoic acid induces RXR homodimer formation. Our results demonstrate a new mechanism for retinoid action by which a ligand-induced homodimer mediates a distinct retinoid response pathway.     

Identification of a second human retinoic acid receptor

We have previously described a human complementary DNA that encodes a novel protein which is homologous to members of the steroid/thyroid nuclear receptor multigene family. This novel protein (hap for hepatoma) exhibits strong homology with the human retinoic acid receptor (RAR) which has been recently characterized. To test the possibility that the hap protein might also be a retinoid receptor, a chimaeric receptor was created by replacing the putative DNA binding domain of hap with that of the human oestrogen receptor (ER). The resulting hap-ER chimaera was then tested for its ability to trans-activate an oestrogen-responsive reporter gene (vit-tk-CAT) in the presence of possible receptor ligands. Here we show that retinoic acid (RA) at physiological concentrations is effective in inducing the expression of this reporter gene by the hap-ER chimaeric receptor. This demonstrates the existence of two human retinoic acid receptors designated RAR-alpha and RAR-beta.     

Identification of a novel retinoic acid receptor in regenerative tissues of the newt

In urodele amphibians, the progenitor cells that regenerate amputated limbs (known as the blastema) normally replace only the missing structures. After systemic delivery of retinoic acid (RA), more proximal structures are also formed, indicating that RA can control position specification in the proximal-distal axis of the regenerating limb. According to dose and experimental context, retinoids can also re-specify the anteroposterior axis of the limb, induce deletions of skeletal elements, or block re-growth completely. To study the molecular basis of these morphogenetic effects, we screened complementary DNA libraries of newt regenerative tissues (limbs and tails) for hormone nuclear receptors activated by RA. Two functional retinoic acid receptors (RARs) were identified, one of which is the newt homologue of the human alpha-receptor (RAR alpha). The second receptor, called RAR delta, is novel. Sequence analysis suggests that the composite newt RAR previously reported is chimaeric, consisting of 5'RAR-beta-like and 3' RAR delta clones. We conclude that multiple RARs are expressed during limb regeneration in amphibians and suggest that receptor heterogeneity may underlie the different effects of retinoids on limb morphogenesis.     

Spatial and temporal expression of the retinoic acid receptor in the regenerating amphibian limb

Retinoic acid is known to have dramatic effects on vertebrate limb pattern in development and regeneration, supporting a model in which a gradient of retinoic acid serves as a morphogen to differentially supply positional information to a developing limb. The discovery of a retinoic acid receptor (RAR) and its homology to the steroid and thyroid hormone receptors provided a potential molecular mechanism for limb morphogenesis. One prediction of this model is that the receptor must be expressed in the developing and regenerating limb anlage. We investigated the expression of the RAR in the adult newt, Notophthalmus viridescens, whose amputated limbs are capable of regenerating and upon which retinoic acid can act to alter pattern. We report the cloning of cDNAs encoding a functional newt RAR and the localization of high and uniform levels of RAR mRNA specifically in the regenerating cells that control limb pattern. These results indicate that the morphogenic field is established through differential activation of pre-existing retinoic acid receptors rather than differential expression of the RAR gene.     

Retinoid X receptor activities of source waters in China and their removal efficiencies during drinking water treatment processes

There is increasing evidence of estrogenic activities of source waters and drinking waters in China based on estrogen receptors(ERs) testing.However,relating such activities to retinoid X receptors(RXRs) in both drinking and source waters are lacking.To rectify this situation,we assessed 23 source water samples from six major river systems in China.We also collected samples at various stages of water processing from three drinking water treatment plants(DWTPs) using a two-hybrid RXR yeast assay with and without metabolism.No RXR agonistic activity was observed,but significant antagonistic activity was detected in all sample extracts.The RXR antagonistic activities of source water sample extracts ranged from 15.2% to 57.8% without metabolism and 11.5% to 68.3% with metabolism,respectively.In the drinking water treatment processes,RXR antagonistic activities without metabolism and with metabolism of up to 31.4% and 37.5% were removed,respectively.Nevertheless,the remaining RXR antagonists in treated drinking water from these source waters could still be harmful to human health.To the best of our knowledge,the occurrence of in vitro RXR disruption activities in source and drinking water has not been previously reported in China.Therefore,an attempt was made to conduct detailed studies investigating RXR disrupting activities and their possible risks in source and drinking water.     

视黄素受体介导的视黄素（retinoids）抗癌作用

对近年来视黄素受体介导视黄素抗癌作用机理的研究进展作一综述,主要有：1）视黄素受体;2）视黄素受体与细胞生长;3）视黄素受体与孤生受体的的相关性;4）视黄素受体对AP-1活性的抑制作用;5）视黄素受体介导的信号转导途径。通过研究,对于探讨视黄素受体的功能、阐明视黄素的抗癌机理、合成更多的受体选择性视黄素具有重要的现实意义。     

A human retinoic acid receptor which belongs to the family of nuclear receptors

A cDNA encoding a protein that binds retinoic acid with high affinity has been cloned. The protein is homologous to the receptors for steroid hormones, thyroid hormones and vitamin D3, and appears to be a retinoic acid-inducible trans-acting enhancer factor, suggesting that the molecular mechanisms of the effect of retinoids (vitamin A) on embryonic development, differentiation and tumour cell growth are similar to those described for other members of this nuclear receptor family.     

Epidermal growth factor receptors increase during the differentiation of embryonal carcinoma cells.

Mouse teratocarcinoma stem cells (embryonal carcinoma, or EC cells) bind very small amounts of mouse epidermal growth factor (EGF) and the latter hormone seems to have no stimulatory effect on the growth of two cloned lines of EC cells. However, when EC cells are induced to differentiate into large flat endodern-like cells (END cells), EGF receptors increase in number reaching a plateau in 6 to 8 days. At 8 to 10 days after induction, END cells multiply very slowly, but when EGF is added (3 x 10(-10) M) to the medium, cell division is stimulated and a further change in morphology occurs. This letter describes the binding characteristics and numbers of the EGF receptors on EC and END cells and shows that exogenous retinoic acid increases the numbers of EGF receptors on END cells. We were unable to find endogenous competing factors produced by EC cells. Such factors could account for the lack of detectable binding of EGF on these cells. As EC cells differentiate to END cells, so the ability of the cells to form tumours is reduced. Since this change is accompanied by an increase in the number of EGF receptors there may be a relationship between these two events.     

Sequential activation of HOX2 homeobox genes by retinoic acid in human embryonal carcinoma cells

RETINOIC acid had been implicated as a natural morphogen in chicken and frog embryogenesis, and is presumed to act through the gene regulatory activity of a family of nuclear receptors. Homeobox genes, which specify positional information in Drosophila and possibly in vertebrate embryogenesis, are among the candidate responsive genes. We previously reported that retinoic acid specifically induces human homeobox gene (HOX) expression in the embryonal carcinoma cell line NT2/D1. We now show that the nine genes of the HOX2 cluster are differentially activated in NT2/D1 cells exposed to retinoic acid concentrations ranging from 10(-8) to 10(-5) M. Genes located in the 3' half of the cluster are induced at peak levels by 10(-8) M retinoic acid, whereas a concentration of 10(-6) to 10(-5) M is required to fully activate 5' genes. At both high and low retinoic acid concentrations, HOX2 genes are sequentially activated in embryonal carcinoma cells in the 3' to 5' direction.