Intramolecular determinants of Retinoid X receptor involved in heterodimeric interactions with its partner PXR: Implications in receptor dynamics and mitotic chromatin interactions.

AIMS: Retinoid X Receptor (RXR) is a central nuclear receptor (NR) that functions as an obligate heterodimeric partner for multiple NRs, including Pregnane X Receptor (PXR), to regulate diverse target gene expression. While subcellular localization of NRs is critical for cellular function, select NRs have been reported to associate with mitotic chromatin, a phenomenon linked to transcriptional memory. Although RXR heterodimers are structurally characterized, the intramolecular determinants within RXR that couple heterodimerization to nuclear import and receptor-chromatin interactions remain unclear. MATERIAL & METHODS: We employed fluorescent protein-tagged receptor constructs, live-cell imaging, site-directed mutagenesis, co-immunoprecipitation assays, promoter-reporter assay and mitotic chromatin fractionation to examine RXR-PXR heterodimerization, subcellular localisation and receptor association with mitotic chromatin. KEY FINDINGS: We identified a stretch of critical 40 amino acid residues (389-428), previously reported as 'Interaction Box' in the LBD region of RXR, that is essential for RXR-PXR heterodimerization-dependent nuclear import and partner-mediated association of RXR with mitotic chromatin. A single amino acid mutation in RXR (L420D) disrupted heterodimerization-mediated nuclear co-import, whereas a combination of triple mutations (Y397D, L420D and R421E) abolished the PXR-dependent mitotic chromatin interaction exhibited by RXR. SIGNIFICANCE: Our study reveals critical molecular determinants within the RXR I-box that are crucial for inter- and intra- molecular interactions of RXR-PXR heterodimers. The insights obtained from this study advance our understanding of how structural determinants within RXR govern partner-mediated receptor-receptor and receptor-chromatin interactions. The identified heterodimerization interface may represent a potential target for therapeutic intervation in diverse disease conditions.