Runx2 (runt related transcription factor 2) is an essential transcription factor for osteoblast proliferation and differentiation. Uridine diphosphate (UDP)-N-acetylgalactosamine (GalNAc): polypeptide GalNAc-transferase 3 (Galnt3) prevents proteolytic processing of fibroblast growth factor 23 (Fgf23), which is a hormone that regulates the serum level of phosphorus. and were expressed in osteoblasts and osteocytes, and expression was restricted to osteocytes in bone. Overexpression and knock-down of upregulated and downregulated, respectively, the expressions of and , and Runx2 directly regulated the transcriptional activity of in reporter assays. The expressions of and in osteoblast-specific knockout () mice were about half those in mice. However, the serum levels of phosphorus and intact Fgf23 in mice were similar to those in mice. The trabecular bone volume was increased during aging in both male and female mice, but the osteoid was reduced. The markers for bone formation and resorption in mice were similar to the control in both sexes. mice exhibited hyperphosphatemia and hypercalcemia, and the intact Fgf23 was about 40% that of wild-type mice. These findings indicated that Runx2 regulates the expressions of and and that Galnt3 decelerates the mineralization of osteoid by stabilizing Fgf23.