Osteoblast-Specific Expression of the α2(I) Collagen Promoter in Transgenic Mice: Correlation with the Distribution of TGF-β1

Abstract

To begin to assess the transcriptional mechanisms that regulate type I collagen gene expression in differentiating osteoblasts, we have sought to determine the minimal promoter sequences that confer osteoblast-specific expression to the α₂(I) collagen gene during murine development. Transgenic mice were generated harboring DNA constructs in which the −2000, −500, and −350 to +54 regions located upstream of the start of transcription were linked to the Escherichia coli β-galactosidase reporter gene (LacZ). Histochemical staining using X-gal indicated that the −2000 lacZ transgene was strongly expressed in newly differentiated and fully functional osteoblasts at intramembranous and endochondral sites of ossification. The promoter was also active in osteocytes in regions of bone remodeling within alveolar bone. The temporal and spatial activity of this region of the promoter closely resembled the developmental patterns of expression of the endogenous α₂(I) collagen gene as determined by in situ hybridization. The cis-acting elements within the 500 and 350 bp segments of the α₂(I) collagen promoter also drove reporter gene expression in forming osteoblasts, although levels of transgene expression were not as marked as that seen with the 2000 bp promoter. Furthermore, the synthesis and secretion of TGF-β₁ in osteogenic zones coincided with areas where the α₂(I) collagen promoter constructs were transcriptionally active. Since a nuclear factor 1 binding site present at −300 has been shown to mediate the effects of TGF-β₁ on the α₂(I) collagen promoter, these data support a role for TGF-β, in the control of this gene during development.