The expression of Clcn7 and Ostm1 in osteoclasts is coregulated by microphthalmia transcription factor

J Biol Chem. 2007 Jan 19;282(3):1891-904. doi: 10.1074/jbc.M608572200. Epub 2006 Nov 14.

Abstract

Microphthalmia transcription factor (MITF) regulates osteoclast function by controling the expression of genes, including tartrate-resistant acid phosphatase (TRAP) and cathepsin K in response to receptor activator of nuclear factor-kappaB ligand (RANKL)-induced signaling. To identify novel MITF target genes, we have overexpressed MITF in the murine macrophage cell line RAW264.7 subclone 4 (RAW/C4) and examined the gene expression profile after sRANKL-stimulated osteoclastogenesis. Microarray analysis identified a set of genes superinduced by MITF overexpression, including Clcn7 (chloride channel 7) and Ostm1 (osteopetrosis-associated transmembrane protein 1). Using electrophoretic mobility shift assays, we identified two MITF-binding sites (M-boxes) in the Clcn7 promoter and a single M-box in the Ostm1 promoter. An anti-MITF antibody supershifted DNA-protein complexes for promoter sites in both genes, whereas MITF binding was abolished by mutation of these sites. The Clcn7 promoter was transactivated by coexpression of MITF in reporter gene assays. Mutation of one Clcn7 M-box prevented MITF transactivation, but mutation of the second MITF-binding site only reduced basal activity. Chromatin immunoprecipitation assays confirmed that the two Clcn7 MITF binding and responsive regions in vitro bind MITF in genomic DNA. The expression of Clcn7 is repressed in the dominant negative mutant Mitf mouse, mi/mi, indicating that the dysregulated bone resorption seen in these mice can be attributed in part to transcriptional repression of Clcn7. MITF regulation of the TRAP, cathepsin K, Clcn7, and Ostm1 genes, which are critical for osteoclast resorption, suggests that the role of MITF is more significant than previously perceived and that MITF may be a master regulator of osteoclast function and bone resorption.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Animals
  • Base Sequence
  • Chloride Channels / biosynthesis*
  • Computational Biology
  • Gene Expression Regulation*
  • Humans
  • Kinetics
  • Membrane Proteins / biosynthesis*
  • Mice
  • Microphthalmia-Associated Transcription Factor / metabolism
  • Microphthalmia-Associated Transcription Factor / physiology*
  • Molecular Sequence Data
  • Osteoclasts / metabolism*
  • Promoter Regions, Genetic
  • Protein Binding

Substances

  • Chloride Channels
  • Clcn7 protein, mouse
  • Membrane Proteins
  • Microphthalmia-Associated Transcription Factor
  • OSTM1 protein, mouse