In Vivo Regulation of Signal Transduction Pathways by Vitamin D Stabilizes Homeostasis of Human Immune Cells and Counteracts Molecular Stress

Abstract

Vitamin D₃ is a pre-hormone that regulates hundreds of target genes and dozens of physiological functions, including calcium homeostasis and the activity of the immune system, via its metabolite 1,25-dihydroxyvitamin D₃, which is a high-affinity ligand for the transcription factor vitamin D receptor. In this study, we took advantage of data from the VitDHiD vitamin D₃ intervention trial (25 healthy individuals) indicating that 442 protein-coding genes were significantly (false discovery rate < 0.05) up- or downregulated in peripheral blood mononuclear cells one day after taking a vitamin D₃ bolus. Since more than half of the encoded proteins had "signaling" assigned as a primary biological function, we evaluated their involvement in signal transduction cascades included in the KEGG (Kyoto Encyclopedia of Genes and Genomes) database and found 88 of the vitamin D targets contributing to 16 different pathways. Eight of the pathways show an approximately even contribution of up- and downregulated genes, suggesting that the actions of vitamin D stabilize homeostasis of the physiological processes driven by the respective signaling cascades. Interestingly, vitamin D target genes involved in the signaling pathways of hypoxia-inducible factor 1 (HIF1), tumor necrosis factor (TNF), mitogen-activated protein kinases (MAPKs) and nuclear factor κB (NFκB) are primarily downregulated. This supports the observation that the physiological role of vitamin D in healthy individuals is to tone down certain processes rather than activate them. In conclusion, under in vivo conditions, vitamin D either alleviates the homeostasis of immune cells in healthy individuals or counteracts molecular responses to oxygen deprivation (HIF1), microbe infection (TNF), growth stimulation (MAPKs) and inflammation (NFκB).

Keywords: HIF1; MAPK; NFκB; TNF; immune cells; signal transduction; vitamin D; vitamin D target genes.

Figure 1

Main roles of vitamin D targets. The databases Human Protein Atlas and GeneCards were used to classify 442 vitamin D target genes identified in the VitDHiD study (Table S1) according to the type of protein that they encode (A) and their main biological function (B).

Figure 2

Vitamin D target genes in signal transduction pathways. Proteins encoded by vitamin D target genes were found in 16 signal transduction pathways listed in KEGG (left). The number of vitamin D-responsive genes/proteins listed directly in the KEGG pathways is increased by related proteins found in the pool of 442 targets (right). Please note that the pathways have some overlapping proteins.

Figure 3

Range of basal expression of vitamin D target genes involved in signal transduction pathways. Colored dots indicate the association of the genes with the four different categories of signal transduction pathways (see Figure 2). Mean basal gene expression (log scale) for the 25 individuals is indicated by black dots, and the range is indicated by horizonal lines.

Figure 4

Vitamin D-triggered homeostasis. The signal transduction pathways p53, FOXO, JAK-STAT and PI3K-STAT have approximately equal numbers of protein components, the genes of which are upregulated (green) and downregulated (red) by vitamin D supplementation in healthy individuals. In each pathway the given percentage reflects the number of genes, the direction of regulation of which will contribute to signal inhibition. The color intensity is proportional to the log fold change (FC) of gene expression between d1 and d0. The structure of the pathway follows the design of KEGG. Functionally similar proteins not indicated in KEGG are marked by an asterisk.

Figure 5

Vitamin D-triggered repression of stress signaling. The signal transduction pathways HIF1, TNF, MAPK and NFκB have more than 75% of their vitamin D target genes downregulated (red), i.e., only less than 25% of the genes are upregulated (green). In each pathway the given percentage reflects the number of genes, the direction of regulation of which will contribute to signal inhibition. The color intensity is proportional to the log fold change (FC) of gene expression between d1 and d0. The structure of the pathway follows the design of KEGG. Functionally similar proteins not indicated in KEGG are marked by an asterisk.