Inorganic phosphate regulates multiple genes during osteoblast differentiation, including Nrf2

The process of osteoblast differentiation and matrix mineralization requires a rise in alkaline phosphatase enzymatic activity resulting in the generation of free phosphate. The ability of inorganic phosphate to regulate gene transcription and cellular function represents a potentially novel extracellular signaling mechanism. Using microarray analysis we have identified a discrete set of genes that are either positively or negatively regulated by increased phosphate in MC3T3-E1 cells. The genes downregulated by phosphate encode for osteoblast-related extracellular factors such as collagens, periostin, and decorin. The genes increased by phosphate encode a novel group of transcription factors that may be important in the later stages of osteoblast development in which the environment is high in phosphate. The transcription factor Nrf2 is one such gene. Elevated phosphate levels stimulate an increase in Nrf2 RNA that is not blocked by the translation inhibitor cycloheximide, suggesting that Nrf2 is an immediate response gene. Cloning of the murine nrf2 promoter reveals that elevated phosphate produces an increase in promoter activity that is both time and dose dependent. This analysis reveals multiple genes regulated by the increase in phosphate associated with osteoblast differentiation, adding to our understanding of the intricate communication between osteoblasts and their extracellular environment.