The NF-κB component RelB is essential for dendritic cell (DC) differentiation and maturation. The vitamin D receptor (VDR) is a nuclear receptor that mediates inhibition of DC maturation and transcriptional repression of relB after engagement of its ligand, 1α,25-dihydroxyvitamin D₃, or related analogs (D₃ analogs). Ligand-dependent relB suppression was abolished by a histone deacetylase (HDAC) inhibitor. Constitutive association of VDR with the relB promoter was demonstrated in DCs by chromatin immunoprecipitation. Promoter binding by VDR was enhanced by ligand and reduced by LPS. Association of HDAC3 and HDAC1 with the relB VDR-binding site was observed, but only HDAC3 was reciprocally modulated by D₃ analog and LPS. Overexpression of HDAC3 caused relB promoter suppression, increased sensitivity to D₃ analog, and resistance to LPS. Depletion of HDAC3 attenuated relB suppression by D₃ analog. In vivo, D₃ analog resulted in reduced RelB in DCs from VDR WT mice but not VDR knockout mice. Other NF-lation of RelB and c-Rel in control animals. We conclude that vitamin D-regulated relB transcription in DCs is controlled by chromatin remodeling by means of recruitment of complexes including HDAC3.