Modulation effects of zinc on the formation of vitamin D receptor and retinoid X receptor α-DNA transcription complexes: Analysis by microelectrospray mass spectrometry

The vitamin D receptor (VDR) binds zinc, and the activity of vitamin D dependent genes in cells is influenced by intracellular zinc concentrations. To determine whether zinc influences vitamin D action in cells by modulating the formation of VDR and retinoid x receptor α (RXRα) heterodimer-DNA complexes, we used microelectrospray ionization mass spectrometry (μESI-MS) to assess receptor-DNA interactions in the presence of varying amounts of zinc. In the absence of DNA, VDR and RXRα proteins were primarily monomeric with small amounts of protein homodimers also observed. Zn²⁺ (up to 300 μM) did not change VDR or RXRα monomer/homodimer ratios. Mass spectra of VDR combined with RXRα were a sum of individual protein spectral data. Zn²⁺ had no effect on the interactions of receptors. With increasing amounts of Zn²⁺, additional Zn²⁺ ions were detected bound to VDR and RXRα. μESI-MS analyses of RXRα in the presence of an osteopontin vitamin D DNA response element (OP-VDRE) showed RXRα homodimer/OP-VDRE complexes. DNA-protein complex formation increased on addition of Zn²⁺ up to 200 μM; at 300 μM, Zn²⁺ dissociation of the RXRα homodimer/OP-VDRE complexes occurred, coincident with the appearance of RXRα monomeric protein. When μESI-MS analyses were carried out with VDR and OP-VDRE, VDR homodimer/OP-VDRE complexes were not detected. Addition of Zn²⁺ did not result in VDR/OP-VDRE complex formation. Heterodimeric VDR/RXRα complexes with OP-VDRE were detected by μESI-MS. Addition of 300 μM Zn²⁺ resulted in dissociation of the heterodimeric VDR/RXRα/OP-VDRE complex. Addition of Mg²⁺ in place of Zn²⁺ did not alter protein/OP-VDRE complexes. Our results show that zinc modulates steroid hormone receptor-DNA interactions.