Individual Subunits of Heterodimers Comprised of Retinoic Acid and Retinoid X Receptors Interact with Their Ligands Independently

Abstract

The retinoid X receptor (RXR) is a member of a family of transcription factors, known as hormone nuclear receptors, that mediate the effects of hydrophobic hormones on gene transcription. RXR, which is activated by 9-cis-retinoic acid (9cRA), can modulate several signaling pathways by virtue of its ability to form heterodimers with other members of the receptor family, as, for example, the retinoic acid receptor (RAR). The roles of the individual receptors within heterodimers are not clear as yet. It was recently reported that heterodimerization inhibits transcriptional activation by RXR, an effect that was attributed to an inability of RXR within heterodimers to bind its ligand. This inhibition was reported to depend on the association of heterodimers with cognate DNA and on the level of saturation of the RAR subunit within the heterodimers. In the present work, the ligand-binding characteristics of RXR and RAR individually and within heterodimers were examined by fluorescence-based methods. The results indicate that heterodimerization with RAR does ot alter the ligand-binding capacity of RXR nor the rate of dissociation of the ligand from this receptor. The ligand-binding capacity of RXR also was not affected by association of heterodimers with cognate DNA nor by the level of saturation of the RAR subunit. The data indicate further that the affinity of RAR for 9cRA is considerably higher as compared to RXR and that this differential affinity is retained within RAR-RXR heterodimers. Thus, binding of ligands by subunits within RAR-RXR heterodimers proceeds independently.

Distinct Roles for Cellular Retinoic Acid-binding Proteins I and II in Regulating Signaling by Retinoic Acid

“…RA transfer from CRABP to vesicles could thus be followed by monitoring the time-dependent decrease in the fluorescent of the ligand. In this assay, because the amount of vesicles used was sufficient to draw Ͼ95% of the ligand from the protein, the rate constant of the observed reaction directly reflects the rate constant for dissociation of RA from the protein (k off ) (16,20). Representative traces showing transfer of RA from CRABP-I or CRABP-II to vesicles are shown in Fig.…”

Suppression of Mammary Carcinoma Cell Growth by Retinoic Acid: the Cell Cycle Control Gene Btg2 Is a Direct Target for Retinoic Acid Receptor Signaling

“…Recombinant histidine-tagged proteins were expressed in Escherichia coli and purified as previously described (28). Purified proteins were dialyzed against HEK buffer [10 mmol/L HEPES (pH 8), 0.1 mmol/L EDTA, 100 mmol/L KCl] and stored at À20jC in 50% glycerol.…”

Retinoid X Receptor Signalling in the Specification of Skeletal Muscle Lineage

“…The exact response is therefore highly dependent on the identity of the ligand and cannot simply be classified as agonistic versus antagonistic. Although RXR can engage in ligand binding when RAR is ligand occupied and/or if a suitable synthetic ligand is present (Chen et al, 1996;Kersten et al, 1996;Lala et al, 1996;Minucci et al, 1997;Roy et al, 1995), bexarotene is unable to transactivate the RXR/RAR heterodimer . In fact, bexarotene has been reported to reduce interactions between RXRs and RARs whereas ligand such as 9-cis increases the binding of RXRs to RARs (Dong and Noy, 1998).…”