Constitutively active RAS signaling reduces 1,25 dihydroxyvitamin D-mediated gene transcription in intestinal epithelial cells by reducing vitamin D receptor expression

Abstract

High vitamin D status is associated with reduced colon cancer risk but these studies ignore the diversity in the molecular etiology of colon cancer. RAS activating mutations are common in colon cancer and they activate pro-proliferative signaling pathways. We examined the impact of RAS activating mutations on 1,25 dihydroxyvitamin D (1,25(OH)2D)-mediated gene expression in cultured colon and intestinal cell lines. Transient transfection of Caco-2 cells with a constitutively active mutant K-RAS (G12V) significantly reduced 1,25(OH)2D-induced activity of both a human 25-hydroxyvitamin D, 24 hydroxyase (CYP24A1) promoter-luciferase and an artificial 3X vitamin D response element (VDRE) promoter-luciferase reporter gene. Young Adult Mouse Colon (YAMC) and Rat Intestinal Epithelial (RIE) cell lines with stable expression of mutant H-RAS had suppressed 1,25(OH)2D-mediated induction of CYP24A1 mRNA. The RAS effects were associated with lower Vitamin D receptor (VDR) mRNA and protein levels in YAMC and RIE cells and they could be partially reversed by VDR overexpression. RAS-mediated suppression of VDR levels was not due to either reduced VDR mRNA stability or increased VDR gene methylation. However, chromatin accessibility to the VDR gene at the proximal promoter (−300 bp), an enhancer region at −6 kb, and an enhancer region located in exon 3 was significantly reduced in RAS transformed YAMC cells (YAMC-RAS). These data show that constitutively active RAS signaling suppresses 1,25(OH)2D-mediated gene transcription in colon epithelial cells by reducing VDR gene transcription but the mechanism for this suppression is not yet known. These data suggest that cancers with RAS-activating mutations may be less responsive to vitamin D mediated treatment or chemoprevention.

Inhibition of HAS2 and hyaluronic acid production by 1,25-Dihydroxyvitamin D3 in breast cancer

“…Upon H-RAS V12 transformation, HAS2 was significantly increased while both HAS1 and HAS3 were strongly reduced. Consistent with previous reports that RAS transformation abrogates VDR signaling [34][35][36], 1,25D3 failed to suppress HAS2 mRNA in HMLE-RAS cells and did not alter the expression of HAS1 or HAS3. Of the hyaluronidase genes, HYAL1 was increased while HYAL2 was decreased in HMLE-RAS cells relative to HMLE parental cells, but no consistent effects of 1,25D3 on these genes were noted (data not shown).…”

“…These data suggest that both murine and human VDR are capable of repressing Has2 when bound by 1,25D3. It is worth noting that 1,25D3 did not suppress HAS2 in mammary epithelial cells expressing H-RAS V12 , likely because this oncogene has consistently been shown to corrupt VDR signaling [ 34 – 36 ]. Collectively these data establish that 1,25D3 regulation of Has2 is mediated through VDR rather than through alternative receptors such as the retinoic acid receptor-related orphan receptors (RORs) which can be activated by other dihydroxyvitamin D3 metabolites including 20,23(OH) 2 D 3 [ 44 ].…”

Exploring vitamin D metabolism and function in cancer

“…Third, constitutively active mutations of the Ras oncogene found in many cancers suppress VDR expression in tumors. The expression of K-Ras mutants in human colon cancer cells and H-Ras mutants in mouse colon and rat intestinal epithelial cells inhibit calcitriol-dependent VDR activation by suppressing VDR transcription 120 . In addition to reducing VDR expression, H-Ras and K-Ras mutations expressed in keratinocytes and PEC lines, respectively, suppressed VDR transcriptional activity by inducing phosphorylation of RXRα, which impairs the recruitment of co-activator SRC-1 to RXRα 121 , 122 .…”

Ancient Nuclear Receptor VDR With New Functions: Microbiome and Inflammation

“…30 Constitutive RAS signaling, which is commonly observed in colon cancer, downregulates VDR expression and thus inhibits vitamin D signaling. 31 We need also consider the heterogeneity of VDR and its variable expression in physiological and pathological conditions in the intestine when assessing the efficacy of vitamin D / vitamin D analogue treatments in IBD.…”