Metabolic defects in phytanic acid catabolism have been shown to be connected with a number of human diseases which can lead to lethal defects of the nervous system and other organs. These effects are probably a result of the very high accumulation of phytanic acid in tissues throughout the body, due to defects in phytanic acid oxidation, the peroxisome being a major site for this process. The nuclear hormone receptors peroxisome proliferator-activated receptor and retinoid X receptor (RXR) have been shown to function as transcription factors in the control of the peroxisomal enzyme expression. Known activators of peroxisome proliferator-activated receptor include polyunsaturated fatty acids and, for RXR, the 9 4 s isomer of retinoic acid. Here we report that phytanic acid is also a natural ligand for RXRa, being able to activate a RXR-responsive promoter. We present evidence that phytanic acid binds to RXRa, promotes formation of an RXRa/RXR response element complex (as detected by gel retardation), and induces a RXRa conformational change similar to that induced by 9-cis-retinoic acid (as detected by protease sensitivity). These results suggest an involvement of RXRa in the control of fatty acid metabolism and could imply that RXRs have a role in the disease effects resulting from defective phytanic acid catabolism.

The pleiotropic effects of retinoic acid on cell differentiation and proliferation are mediated by two subfamilies of nuclear receptors, the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs).Recently the synthetic retinoid Ro 41-5253 was identified as a selective RARa antagonist. As demonstrated by gel retardation assays, Ro 41-5253 and two related new RARa antagonists do not influence RARcWRXRa heterodimerization and DNA binding. In a limited trypsin digestion assay, complexation of RARca with retinoic acid or several other agonistic retinoids altered the degradation of the receptor such that a 30-kDa proteolytic fragment became resistant to proteolysis. This suggests a ligand-induced conformational change, which may be necessary for the interaction of the DNA-bound RARorRXRc heterodimer with other transcription factors. Our results demonstrate that antagonists compete with agonists for binding to RARaL and may induce a different structural alteration, suggested by the tryptic resistance of a shorter 25-kDa protein fragment in the digestion assay. This RARe conformation seems to allow RARW/RXRcr binding to DNA but not the subsequent transactivation of target genes. Protease mapping with C-terminally truncated receptors revealed that the proposed conformational changes mainly occur in the DE regions of RARa. Complexation of RARI3, RARy, and RXRe, as well as the vitamin D3 receptor, with their natural ligands resulted in a similar resistance of fragments to proteolytic digestion. This could mean that ligand-induced conformational changes are a general feature in the hormonal activation of vitamin D3 and retinoid receptors.

Cellular responsiveness to retinoic acid and its metabolites is conferred through two distinct families of receptors: the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs). Herein, we report on the identification and characterization of several conformationally restricted retinoids, which selectively bind and activate RX receptors. Under the influence of retinoids, HL-60 myelocytic leukemia cells differentiate into granulocytes. This effect is mediated by RAR␣, as has been demonstrated through the use of a selective RAR␣ antagonist (Apfel, C., Bauer, F., Crettaz, M., Forni, L., Kamber, M., Kaufmann, F., LeMotte, P., Pirson, W., and Klaus, M. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 7129 -7133). Here, we show that conformationally restricted RXR-specific retinoids, at doses that are per se inactive, are able to potentiate by up to one order of magnitude the pro-differentiating effects of all-trans retinoic acid and an RAR␣-selective synthetic retinoid. We also present evidence that these RXR-selective ligands are able to bind to a DNA RXR⅐RAR heterodimer complex. This finding demonstrates that agonists for RARs and RXRs can synergistically promote HL-60 differentiation, which could be mediated through a heterodimer of these receptors.A subclass of nuclear hormone receptors have been described, termed retinoid X receptors (RXR) 1 (2-4), which are ligand-inducible transcription factors responsive to the 9-cis isomer of retinoic acid (5, 6). A related subclass of receptors, the retinoic acid receptors (RAR) (7-12), also responds to 9-cis retinoic acid (9-cis RA), as well as to the all-trans isomer of retinoic acid (t-RA). A striking feature of RXRs is their ability to heterodimerize with several members of the steroid receptor superfamily, including the RA receptors, thyroid receptors, peroxisome proliferator-activated receptors, vitamin D receptor, and chicken ovalbumin upstream promoter-transcription factor (13-21). Growing evidence suggests that heterodimers are the active species for binding to promoter response elements.In view of the involvement of RXRs in multiple signaling pathways, RXR-selective ligands can possibly be expected to show medical usefulness through their modulation of the above mentioned and perhaps also other hormone-regulated processes. The pharmacological effects of RXR ligands, which are attributable solely to activity on RXRs, are difficult to assess using 9-cis RA as an activating ligand since 9-cis RA also functions as an RAR ligand and consequently elicits the full range of classical retinoid effects. Therefore, to gain deeper insight into RXR function as well as to evaluate their pharmacological usefulness, we have designed and characterized a number of RXR-selective compounds and report on some intriguing pharmacological properties. These compounds are sterically restricted analogues of 9-cis RA, which cannot isomerize. They exhibit high potency and high selectivity for RXRs versus RARs and belong to different structural classes than other RXR-selective compounds r...

The human retinoic acid receptor u was expressed in Escherichia coli. The recombinant protein was found to be very unstable in several E. coli strains, probably due to proteolysis. Conditions were established to obtain reasonable amounts of active protein for ligand and DNA binding studies. The recombinant receptor showed the expected DNA binding activities in gel-retardation assays. Ligand binding properties were measured in a charcoal absorption assay. The dissociation constant for highly specific bound retinoic acid was found to be 0.67 nM. The affinity of several synthetic retinoids to thc recombinant protein was determined and compared to their biological activity. Some of the values presented here differ significantly from those published earlier for the receptor or its isolated hormonebinding domain.The nuclear receptors for retinoids play a major role in the regulation of genes involved in cell differentiation and growth. Their natural ligand, the vitamin A derivative alltrans-retinoic acid, has been reported to be a possible morphogen in the embryonic tissues of frogs and chicken.