Clinical observations show that an increase in serum phosphorus (Pi) is linked to higher cardiovascular (CV) mortality, while vitamin D receptor (VDR) agonist therapy is associated with survival benefit in Stage 5 chronic kidney disease. Smooth muscle cells (SMCs) play an important role in CV pathophysiology, but the interaction between Pi and the VDR signaling pathway in SMCs is not known. Real-time RT-PCR studies revealed that elevated Pi (2.06 mM) dampened VDR-mediated regulation of a panel of genes including thrombomodulin and osteopontin in SMCs. DNA microarray results demonstrate that increasing Pi from 0.9 to 2.06 mM exerted a widespread dampening effect on VDR-mediate gene expression. A total of 325 target genes were affected by paricalcitol at 0.9 mM Pi, with 195 up- and 130 down-regulated. The number of target genes affected by paricalcitol at 2.06 mM Pi decreased to 86, with 55 up- and 31 down-regulated. VDR-mediated gene expression in As4.1 cells (a JG cell-like cell line derived from kidney tumors in SV40 T-antigen transgenic mice) and PPAR mediated gene expression in SMCs were also altered by elevated Pi, suggesting that the observation is not unique to VDR in SMCs. Mechanism analysis showed that elevated Pi had no effect on the VDR or PPAR protein level, but altered the cytosolic vs. nuclear distribution of NFkB or Ncor1. Our results demonstrated for the first time that elevated Pi affects VDR-mediated gene expression in human coronary artery SMCs and the effect is not limited to VDR in SMCs.