Cloning and characterization of the 5′-flanking region of the mouse tartrate-resistant acid phosphatase gene

Sakamuri V. Reddy, Thierry Scarcez, Jolene J. Windle, Robin J. Leach, Jeffrey E. Hundley, John Chirgwin, Janice Y. Chou, G. David Roodman

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

Little information is available on the molecular mechanisms controlling osteoclastic bone resorption. We used tartrate-resistant acid phosphatase (TRAP) to begin to investigate the regulation of bone resorption at the molecular level. TRAP is expressed at high levels in osteoclasts and may play an important role in the bone resorptive process. Therefore, we isolated the murine TRAP gene from a mouse spleen genomic library and characterized its promoter. A restriction map was generated for the 17 kb TRAP insert. A 2 kb Smal fragment, containing the 5′-flanking region, was subcloned and the nucleotide sequence determined. Sequence analysis of the Smal fragment revealed the presence of numerous candidate transcription factor binding sequences, including those for AP1 and H-APF-1. The H-APF-1 site matches the consensus sequence for the IL-6-regulated transcription factor. An intron was identified at -1 to -393 bp relative to the ATG. The presence of an intron was confirmed by PCR analysis of RNA isolated from murine osteoclasts. Primer extension analysis indicated the presence of a transcription initiation site at -552 bp from the ATG. The region from -1846 to 2 bp relative to the ATG initiation codon drove the transient expression of a luciferase reporter gene when transfected into HRE H9 rabbit endometrial cells. PMA treatment of HRE H9 cells enhanced luciferase transcription approximately threefold. These data suggest that the TRAP promoter is complex and contains multiple regulatory elements. The availability of the TRAP promoter may also permit production of transgenic mice, which can be used to develop previously unavailable osteoclast cell lines.

Original languageEnglish (US)
Pages (from-to)1263-1270
Number of pages8
JournalJournal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research
Volume8
Issue number10
StatePublished - Oct 1993
Externally publishedYes

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5' Flanking Region
Organism Cloning
Osteoclasts
Genes
Bone Resorption
Ubiquitin
Luciferases
Introns
Transcription Factors
Initiator Codon
Genomic Library
Transcription Initiation Site
Consensus Sequence
Reporter Genes
Transgenic Mice
Sequence Analysis
Tartrate-Resistant Acid Phosphatase
Interleukin-6
Spleen
RNA

ASJC Scopus subject areas

  • Surgery

Cite this

Cloning and characterization of the 5′-flanking region of the mouse tartrate-resistant acid phosphatase gene. / Reddy, Sakamuri V.; Scarcez, Thierry; Windle, Jolene J.; Leach, Robin J.; Hundley, Jeffrey E.; Chirgwin, John; Chou, Janice Y.; Roodman, G. David.

In: Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research, Vol. 8, No. 10, 10.1993, p. 1263-1270.

Research output: Contribution to journalArticle

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AU - Hundley, Jeffrey E.

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