Genome-wide analyses of gene expression profile identify key genes and pathways involved in skeletal response to phosphate and 1,25-dihydroxyvitamin D3 in vivo

Abstract

Phosphate (P) is an essential element involved in various biological actions, such as bone integrity, energy production, cell signaling and molecular component. P homeostasis is modulated by 4 main tissues; intestine, kidney, bone, and parathyroid gland, where 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃), parathyroid hormone and fibroblast growth factor 23 (FGF23) are produced and/or have an influence. In bone, serum P level modulates the production of FGF23 which then controls not only P excretion but also vitamin D metabolism in kidney in an endocrine manner. The hormonally active form of vitamin D, 1,25(OH)₂D₃, also has a significant effect on skeletal cells via its receptor, the vitamin D receptor, to control gene expression which mediates bone metabolism as well as mineral homeostasis. In this study, we adopted RNA-seq analysis to understand genome-wide skeletal gene expression regulation in response to P and 1,25(OH)₂D₃. We examined lumbar 5 vertebrae from the mice that were fed P deficient diet for a week followed by an acute high P diet for 3, 6, and 24 h as well as mice treated with 1,25(OH)₂D₃ intraperitoneally for 6 h. Further identification and exploration of the genes regulated by P and 1,25(OH)₂D₃ showed that P dynamically modulates the expression of skeletal genes involved in various biological processes while 1,25(OH)₂D₃ regulates genes highly related to bone metabolism. Our in vivo data were then compared with in vitro data that we previously obtained, which suggests that the gene expression profiles presented in this report mainly represent those of osteocytes. Interestingly, it was found that even though the skeletal response to P is distinguished from that to 1,25(OH)₂D₃, both factors have an effect on Wnt signaling pathway to modulate bone homeostasis. Taken together, this report presents genome-wide data that provide a foundation to understand molecular mechanisms by which skeletal cells respond to P and 1,25(OH)₂D₃.

Keywords: 1,25-dihydroxyvitamin D(3); Bone; Phosphate; RNA-seq; Transcription; Transcriptome; Wnt signaling pathway.