Reciprocal regulation of the osteocalcin promoter by the homeodomain proteins DLX5 & MSX2

D. A. Towler, E. P. Newberry, T. Latifi

Research output: Contribution to journalArticle

Abstract

The osteocalcin (OC) gene encodes a gamma - carboxylated protein that controls matrix mineralization in bones and teeth. OC expression in calvarial osteoblasts is regulated in part by protein-protein interactions between the homeodomain represser Msx2 and TFIIF. Recently, a myoblast-specific enhancer was shown to be reciprocally regulated by Msxl and the Dix family. We wished to assess whether DlxS (expressed in bone) and Msx2 regulate the OC promoter in a similar fashion in MC3T3-E1 osteoblasts. CMV- Msx2 and CMV-Dlx5 expression vectors were co-transfected with the 0.2 kb OC basal promoter linked to a luciferase reporter. Msx2 suppresses OC promoter activity by 90%. By contrast, Dlx5 stimulates (2-fold) basal OC promoter activity, and completely reverses Msx2 - dependent OC promoter repression. Deletion of the DlxS N-terminal homeodomain extension abolishes Dlx5 "antirepressor" function. Dlx5 does not bind the OC HOXBOX; moreover, regulation of OC promoter activity by Msx2 and Dlx5 is independent of the HOXBOX cognate, suggesting that antagonism occurs via protein-protein interactions. In Far Western blot assays, radio]abeled DlxS interacts with Msx2, TFIIF-30, and weakly with TFIIF-74, but not with TBP. Msx2 binds DlxS, TFIIF-74 and TFIIF-30_ but not TBP - via suppressor domain residues 132-148. Finally, a point mutation in Msx2(P148H) that causes craniosynostosis gives rise to a variant that is not regulated by Dlx5; viz., Dlx5 cannot reverse OC promoter suppression by Msx2<Pl 48H). These data suggest that DlxS and Msx2 reciprocally regulate OC gene expression during calvarial osteoblast differentiation, and that the dysregulated activity of Msx2(P148H) leads to craniosynostosis in vivo. Dlx5 and Msx2 participate in functionally important protein-protein interactions with each other and TFIIF.

Original languageEnglish (US)
Pages (from-to)A1322
JournalFASEB Journal
Volume12
Issue number8
StatePublished - Dec 1 1998

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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