EXPRESSION OF RAT HOMEOBOX GENE, RHOX, IN DEVELOPING AND ADULT TISSUES IN MICE AND REGULATION OF ITS MESSENGER-RNA EXPRESSION IN OSTEOBLASTS BY BONE MORPHOGENETIC PROTEIN-2 AND PARATHYROID HORMONE-RELATED PROTEIN

The rat homeobox gene, rHox, was cloned from a rat osteosarcoma cDNA l ibrary. Southwestern and gel mobility shift analyses showed that rHox binds to the promoter regions of collagen (alpha 1)I and osteocalcin g enes while transient transfection with rHox resulted in repression of their respective promoter activities. In situ hybridization studies sh owed that rHox mRNA was widely expressed in osteoblasts, chondrocytes, skeletal muscle, skin epidermis, and bronchial and intestinal epithel ial cells, as well as cardiac muscle in embryonic and newborn mice. Ho wever in 3-month-old mice, rHox mRNA expression was restricted to oste oblasts, megakaryocytes, and myocardium. Bone morphogenetic protein 2, a growth factor that commits mesenchymal progenitor cells to differen tiate into osteoblasts, down-regulated rHox mRNA expression by 40-50% in UMR 201, a rat preosteoblast cell line, in a time- and dose-depende nt manner. In contrast, PTH-related protein (PTHrP), recently shown to be a negative regulator of chondrocyte differentiation, significantly enhanced rHox mRNA expression in UMR 106-06 osteoblastic cells by 3-f old at 24 h while at the same time down-regulating expression of pro-a lpha 1(I) collagen mRNA by 60%. Expression of rHox mRNA in calvarial o steoblasts derived from PTHrP -/- mice was approximately 15% of that o bserved in similar cells obtained from normal mice. In conclusion, cur rent evidence suggests that rHox acts as a negative regulator of osteo blast differentiation. Furthermore, down-regulation of rHox mRNA by bo ne morphogenetic protein 2 and its up-regulation by PTHrP support a ro le of the homeodomain protein, rHox, in osteoblast differentiation.