Discovery of novel vitamin D receptor interacting proteins that modulate 1,25-dihydroxyvitamin D3 signaling

Abstract

The nuclear vitamin D receptor (VDR) modulates gene transcription in 1,25-dihydroxyvitamin D3 (1,25D) target tissues such as kidney, intestine, and bone. VDR is also expressed in heart, and 1,25D deficiency may play a role in the acceleration of cardiovascular disease. Employing a yeast two-hybrid system and a human heart library, using both a 1,25D-independent and 1,25D-dependent screen, we discovered six candidate VDR interacting proteins (VIPs). These novel VIPs include CXXC5, FASTK, NR4A1, TPM2, MYL3 and XIRP1. Mammalian two-hybrid assays as well as GST pull-downs were used to confirm VIP-VDR interaction, and the combination of these two assays reveals that CXXC5, XIRP1, FASTK and NR4A1 interactions with VDR may be modulated by 1,25D. The functional effects of these VIPs on 1,25D-mediated gene expression were explored in transcriptional assays employing three separate and distinct 1,25D-responsive element (VDRE)-driven luciferase reporter genes in transfected Caco-2 and HEK-293 cells, and in a C2C12 myoblast line. FASTK and TPM2 activated expression in all cell line and promoter contexts, while CXXC5 and XIRP1 exhibited differing effects depending on the cell line and promoter employed, suggesting promoter and cell-specific effects of these unique VIPs on VDR signaling. Further evaluation of the interaction between CXXC5 and VDR revealed that CXXC5 acts in a dose-dependent manner to stimulate VDR-mediated transcription on select VDREs. Identification of novel heart VIPs and their influence on VDR activity may increase our understanding of how vitamin D impacts cardiac physiology and may facilitate development of VDR/VIP drug analogs to combat heart disease.

Estradiol-Estrogen Receptor α Mediates the Expression of the CXXC5 Gene through the Estrogen Response Element-Dependent Signaling Pathway

“…Moreover, it was also suggested that CXXC5 acts as a transcription repressor for hypoxia-mediated Cytochrome C oxidase subunit 4/2 ( COX4/2 ) through a direct interaction with an oxygen responsive element in the proximal promoter of the gene16. Interestingly, CXXC5 was reported to act as a vitamin D (VitD) receptor interactor to enhance or repress transcription of VitD responsive genes depending on the corresponding promoter components21.…”

“…Corroborating the importance of CXXC5 in cellular events, de-regulation of CXXC5 expression appears to correlate with a number of pathologies including diminished ovarian reserve (DOR), Blepharophimosis Ptosis Epicantus inversus Syndrome (BPES), cardiovascular disease, myelodysplastic syndrome19202122. Altered CXXC5 expression was also found to be associated with locally advanced breast tumors, metastatic malignant melanomas, papillary thyroid carcinomas18 and Acute Myeloid Leukemia38.…”

“…However, it appears that CXXC5 expressed in different tissues at varying levels12 is involved in the modulation of cellular proliferation, differentiation and death as a transcription factor, transcription co-regulator and/or chromatin modifier in response to retinoic acid, bone morphogenetic protein 4 (BMP4), Wnt signaling as well as to hypoxia1213141617. The de-regulated expression of CXXC5 is also suggested to correlate with the development, and resistance to therapies, of various pathologies including cardiovascular disease, diminished ovarian reserve (DOR), Blepharophimosis Ptosis Epicantus inversus Syndrome (BPES), Acute Myeloid Leukemia (AML) and breast cancer12181920212223.…”

Vitamin D receptor ChIP-seq in primary CD4+ cells: relationship to serum 25-hydroxyvitamin D levels and autoimmune disease

“…One possibility is that in vivo VDR binding is modulated by protein-protein interactions with co-factors: SP1 and ETS1 are known to modulate VDR binding, and there is some evidence that interactions between SP1 and VDR may enable modulation of genes that lack a classical VDR recognition motif [43,44]. Several other proteins are known to bind in association with VDR, including NR4A1 and c-MYC [45,46]. CTCF is known to modulate DNA binding via protein-protein interactions with other nuclear receptors [47-49].…”

Downregulation of the Wnt inhibitor CXXC5 predicts a better prognosis in acute myeloid leukemia

“…Interestingly, Kim et al demonstrated that the ability of CXXC5 to bind Dvl was less important for Wnt inhibition than presence of the CXXC domain, further suggesting other mechanisms of inhibition. 4 It has been shown that CXXC5 interacts with the vitamin D receptor, 43 which could lead to Wnt suppression. In our GEP analysis of Wnt-stimulated K562 cells, we identified Wnt repressors HBP1 and AXIN1 significantly altered by CXXC5 expression, which could mediate CXXC5-associated effects on Wnt signaling.…”