Figure 6.
Runx2 regulates AChE transcription. A, cultured osteoblasts were transfected with pcDNA3, or cDNA-encoding Runx2, and cell lysates were collected 3 days after transfection. Left panel, cell lysates were subjected to ALP and AChE assays. Right panel, total RNAs were extracted from the cultures to perform real-time PCR analysis. B, deletion of the Runx2-binding site on the ACHE promoter was shown. A schematic diagram of human ACHE promoter with key transcription elements was shown. The Runx2-binding site sequence (ACC GCC) was mutated into AGG GCC on the human promoter. C, left panel, cultured osteoblasts were transiently transfected with pAChE-Luc and its mutant pAChEΔRunx2-Luc for 24 h before the application of Wnt3a (200 ng/ml) for 3 days. Right panel, cultured osteoblasts were transiently co-transfected with pAChE-Luc, or pAChEΔRunx2-Luc, with or without cDNA encoding Runx2. The cultures were collected 3 days later for luciferase assay and ALP assay. Values are expressed as fold-increase to basal reading, and are mean ± S.E., n = 5, each with triplicate samples, *, p < 0.05; **, p < 0.01; ***, p < 0.001.