EPDR1 Governs Metabolic and Immunologic Re-programming of Human Mesenchymal Stem Cells During Osteoblast Differentiation

Abstract

EPDR1 is a novel human osteoblast regulatory gene previously identified by intersecting BMD GWAS datasets with promoter-focused Capture-C and ATAC-seq generated in differentiating human osteoblasts. Recently, we showed that the open chromatin region harboring the BMD variants rs1524068, rs6975644 and rs940347 functions as an osteoblast specific EPDR1 enhancer in hFOB1.19 cells. However, the precise molecular processes controlled by EPDR1 is unknown. Herein, we knocked-down EPDR1 expression in three unique bone-marrow derived human mesenchymal stem cell donor lines (hMSC) using small interfering RNA and evaluated the global transcriptomic changes with or without BMP2 stimulation using osteopermissive media. Globally, 3,861 genes were differentially regulated (1,888 upregulated and 1,973 downregulated) by BMP in non-targeted cells, whereas 2,744 genes were differentially regulated (1,528 upregulated and 1,216 downregulated) in EPDR1 silenced cells. As expected, genes associated with osteoblast differentiation, BMP signaling pathway, Notch signaling pathway and tissue morphogenesis were expressed in non-targeted cells; however, none of these biological processes achieved significance in EPDR1 silenced cells, suggesting major pathway perturbations. To understand these changes, we performed a donor-specific comparison with EPDR1-silenced cells. Of the ~2,300 genes differentially regulated, we observed an upregulation of genes related to inflammatory response, cytokine-mediated signaling pathway, and regulation of T cell activity, while genes related to cell division were decreased. Additionally, we observed differentially expressed genes enriched for pathways involved in regulation of fatty acid metabolic process, regulation of bone resorption, arachidonic acid metabolic process, negative regulation of collagen biosynthetic process and positive regulation of tumor necrosis factor, interferon-gamma, and interleukin-8 production upon BMP2 stimulation. The crystal-structure of EPDR1 protein suggests human EPDR1 folds into a dimer using a monomeric subunit consisting of a deep hydrophobic pocket to bind to hydrophobic fatty acids and function as a lipoprotein carrier. Since EPDR1 is a secreted protein, an immune-reactive profile driven by loss of EPDR1 could be metabolically mediated via disturbances in cellular lipoprotein and fatty acid trafficking. We conclude that EPDR1 could play an important role in pathophysiological bone turnover later in life and therefore warrants further investigation.http://deepblue.lib.umich.edu/bitstream/2027.42/191456/2/EPDR1 siR RNA SEQ V2 kdh.pd