Transcriptional control of the tissue-specific, developmentally regulated osteocalcin gene requires a binding motif for the Msx family of homeodomain proteins
Biochemistry & Molecular Pharmacology
Graduate School of Biomedical Sciences; Department of Cell Biology and Cancer Center
Medical Subject Headings
Animals; Base Sequence; Binding Sites; Cell Differentiation; Cells, Cultured; DNA-Binding Proteins; *Gene Expression Regulation, Developmental; Homeodomain Proteins; MSX1 Transcription Factor; Molecular Sequence Data; Mutagenesis, Site-Directed; Oligodeoxyribonucleotides; Osteoblasts; Osteocalcin; Promoter Regions (Genetics); RNA, Messenger; Rats; Structure-Activity Relationship; *Transcription Factors; Transcription, Genetic
Life Sciences | Medicine and Health Sciences
The OC box of the rat osteocalcin promoter (nt -99 to -76) is the principal proximal regulatory element contributing to both tissue-specific and developmental control of osteocalcin gene expression. The central motif of the OC box includes a perfect consensus DNA binding site for certain homeodomain proteins. Homeodomain proteins are transcription factors that direct proper development by regulating specific temporal and spatial patterns of gene expression. We therefore addressed the role of the homeodomain binding motif in the activity of the OC promoter. In this study, by the combined application of mutagenesis and site-specific protein recognition analysis, we examined interactions of ROS 17/2.8 osteosarcoma cell nuclear proteins and purified Msx-1 homeodomain protein with the OC box. We detected a series of related specific protein-DNA interactions, a subset of which were inhibited by antibodies directed against the Msx-1 homeodomain but which also recognize the Msx-2 homeodomain. Our results show that the sequence requirements for binding the Msx-1 or Msx-2 homeodomain closely parallel those necessary for osteocalcin gene promoter activity in vivo. This functional relationship was demonstrated by transient expression in ROS 17/2.8 osteosarcoma cells of a series of osteocalcin promoter (nt -1097 to +24)-reporter gene constructs containing mutations within and flanking the homeodomain binding site of the OC box. Northern blot analysis of several bone-related cell types showed that all of the cells expressed msx-1, whereas msx-2 expression was restricted to cells transcribing osteocalcin. Taken together, our results suggest a role for Msx-1 and -2 or related homeodomain proteins in transcription of the osteocalcin gene.
Rights and Permissions
Citation: Proc Natl Acad Sci U S A. 1994 Dec 20;91(26):12887-91.
Proceedings of the National Academy of Sciences of the United States of America
Hoffmann, Heidi M.; Catron, Katrina M.; Van Wijnen, Andre J.; McCabe, Laura R.; Lian, Jane B.; Stein, Gary S.; and Stein, Janet L., "Transcriptional control of the tissue-specific, developmentally regulated osteocalcin gene requires a binding motif for the Msx family of homeodomain proteins" (1994). GSBS Student Publications. 530.