β-Apo-13-carotenone Regulates Retinoid X Receptor Transcriptional Activity through Tetramerization of the Receptor*

Background: β-Apo-13-carotenone, a naturally occurring apocarotenoid, functions as an antagonist of the retinoid X receptor (RXR). Results: β-Apo-13-carotenone inhibits transactivation of RXRα but does not interfere with coactivator binding to the receptor like the known antagonist UVI3003. Conclusion: β-Apo-13-carotenone induces the formation of a transcriptionally silent RXR tetramer. Significance: β-Apo-13-carotenone is a naturally occurring rexinoid with a novel mechanism of antagonism. Retinoid X receptor (RXRα) is activated by 9-cis-retinoic acid (9cRA) and regulates transcription as a homodimer or as a heterodimer with other nuclear receptors. We have previously demonstrated that β-apo-13-carotenone, an eccentric cleavage product of β-carotene, antagonizes the activation of RXRα by 9cRA in mammalian cells overexpressing this receptor. However, the molecular mechanism of β-apo-13-carotenone's modulation on the transcriptional activity of RXRα is not understood and is the subject of this report. We performed transactivation assays using full-length RXRα and reporter gene constructs (RXRE-Luc) transfected into COS-7 cells, and luciferase activity was examined. β-Apo-13-carotenone was compared with the RXRα antagonist UVI3003. The results showed that both β-apo-13-carotenone and UVI3003 shifted the dose-dependent RXRα activation by 9cRA. In contrast, the results of assays using a hybrid Gal4-DBD:RXRαLBD receptor reporter cell assay that detects 9cRA-induced coactivator binding to the ligand binding domain demonstrated that UVI3003 significantly inhibited 9cRA-induced coactivator binding to RXRαLBD, but β-apo-13-carotenone did not. However, both β-apo-13-carotenone and UVI3003 inhibited 9-cRA induction of caspase 9 gene expression in the mammary carcinoma cell line MCF-7. To resolve this apparent contradiction, we investigated the effect of β-apo-13-carotenone on the oligomeric state of purified recombinant RXRαLBD. β-Apo-13-carotenone induces tetramerization of the RXRαLBD, although UVI3003 had no effect on the oligomeric state. These observations suggest that β-apo-13-carotenone regulates RXRα transcriptional activity by inducing the formation of the “transcriptionally silent” RXRα tetramer.