Changes in the kidney proteome of vitamin D receptor knock-out mice.

Vitamin D is a steroid hormone whose main function in organism is regulation of Ca2+ metabolism. The hormone acts in various tissues, but mostly in kidney, bone and intestine, either directly, influencing genomic and nongenomic responses, or indirectly via regulation of Ca2+ homeostasis. To exert its genomic actions vitamin D binds to the vitamin D receptor (VDR), whereupon liganded receptor binds to vitamin D response elements (VDRE) in target gene promoter regions and activates transcription. The mechanism of nongenomic responses, which take only seconds/minutes and modulate different signalling pathways, is still unclear. VDR belongs to the steroid hormone receptor superfamily and contains several domains common to all members of the family, including an N-terminal DNA binding domain, a ligand binding domain, a transcription factor domain and a C-terminal domain responsible for interactions with cofactors. In this study, VDR knock-out mice were examined, in which the DNA binding domain of VDR was partially deleted, thus disabling DNA binding, but the receptor was expressed and had an intact ligand binding domain. This mutation caused ablation of both, genomic and nongenomic responses in mice. Therefore, changes in the expression level of a large number of different proteins, which are regulated by vitamin D, or involved in vitamin D mediated response, were expected. Hence 2-dimensional electrophoresis was used to investigate changes in the kidney proteome of VDR knock-out mice. This approach enables an unbiased exploratory survey of a huge number of proteins at the same time. Using this technique a number of differentially expressed proteins were identified: VDR, cytosolic malate dehydrogenase, lactate dehydrogenase, hydroxyacid oxidase and adenosine kinase. Except VDR, all of them were significantly downregulated in VDR knock-out mice and they were generally involved in central metabolic processes in the cell. Taken together, downregulated expression of these enzymes implied lower energy supplies in kidneys of VDR deficient animals. However, no major functional abnormalities in VDR knock-out mice could be observed. Identification of VDR itself provided additional evidence that the receptor was expressed, even with partially deleted DNA binding domain, and therefore points to the importance of the N-terminal part of the protein in vitamin D response