Tumor Necrosis Factor Receptor Superfamily Member 19 (TNFRSF19) Regulates Differentiation Fate of Human Mesenchymal (Stromal) Stem Cells through Canonical Wnt Signaling and C/EBP

Abstract

Mechanisms controlling human multipotent mesenchymal (stromal) stem cell (hMSC) differentiation into osteoblasts or adipocytes are poorly understood. We have previously demonstrated that Wnt signaling in hMSC enhanced osteoblast differentiation and inhibited adipogenesis by comparing two hMSC cell lines overexpressing mutated forms of the Wnt co-receptor LRP5: T253I (hMSC-LRP5T253) and T244M (hMSC-LRP5T244) conducting high and low level of Wnt signaling, respectively. To explore the underlying molecular mechanisms, we compared gene expression profiles of hMSC-LRP5T253 and hMSC-LRP5T244 treated with Wnt3a using whole genome expression microarrays and found that TNFRSF19 is differentially up-regulated between the two cells lines. Bioinformatic analysis and dual luciferase assay of its promoter revealed that TNFRSF19 transcript 2 (TNFRSF19.2) is a target of canonical Wnt signaling. Knocking down TNFRSF19 in hMSC-LRP5T253 cells decreased Wnt3a-induced osteoblast differentiation marker alkaline phosphate activity and its overexpression in hMSC-LRP5T244 cells increased alkaline phosphate activity. In addition, TNFRSF19 was negatively regulated by adipogenic transcription factor CCAAT/enhancer-binding proteins (C/EBP). Knocking down TNFRSF19 in hMSC-LRP5T253 cells or its overexpression in hMSC-LRP5T244 cells significantly increased or decreased adipogenesis, respectively. In conclusion, we revealed a novel function of TNFRSF19 as a factor mediating differentiation signals that determine the hMSC differentiating fate into osteoblasts or adipocytes. Mechanisms controlling human multipotent mesenchymal (stromal) stem cell (hMSC) differentiation into osteoblasts or adipocytes are poorly understood. We have previously demonstrated that Wnt signaling in hMSC enhanced osteoblast differentiation and inhibited adipogenesis by comparing two hMSC cell lines overexpressing mutated forms of the Wnt co-receptor LRP5: T253I (hMSC-LRP5T253) and T244M (hMSC-LRP5T244) conducting high and low level of Wnt signaling, respectively. To explore the underlying molecular mechanisms, we compared gene expression profiles of hMSC-LRP5T253 and hMSC-LRP5T244 treated with Wnt3a using whole genome expression microarrays and found that TNFRSF19 is differentially up-regulated between the two cells lines. Bioinformatic analysis and dual luciferase assay of its promoter revealed that TNFRSF19 transcript 2 (TNFRSF19.2) is a target of canonical Wnt signaling. Knocking down TNFRSF19 in hMSC-LRP5T253 cells decreased Wnt3a-induced osteoblast differentiation marker alkaline phosphate activity and its overexpression in hMSC-LRP5T244 cells increased alkaline phosphate activity. In addition, TNFRSF19 was negatively regulated by adipogenic transcription factor CCAAT/enhancer-binding proteins (C/EBP). Knocking down TNFRSF19 in hMSC-LRP5T253 cells or its overexpression in hMSC-LRP5T244 cells significantly increased or decreased adipogenesis, respectively. In conclusion, we revealed a novel function of TNFRSF19 as a factor mediating differentiation signals that determine the hMSC differentiating fate into osteoblasts or adipocytes.