Transcriptional regulation of the tartrate-resistant acid phosphatase (TRAP) gene by iron

O. Alcantara, S. V. Reddy, G. David Roodman, D. H. Boldt

Research output: Contribution to journalArticle

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Abstract

Tartrate-resistant acid phosphatase (TRAP) was first identified in cells from patients with hairy cell leukaemia. Subsequently, it has been found in other leukaemias, B-lymphoblastoid cell lines, osteoclasts and subsets of normal lymphocytes, macrophages, and granulocytes. Recent data indicate that TRAP and porcine uteroferrin, a placental iron-transport protein, represent a single gene product. However, the intracellular role of TRAP is unknown. We used a full-length human placental TRAP cDNA probe to examine TRAP expression in human peripheral mononuclear cells (PMCs). TRAP mRNA increased 50-75-fold after 24 h in unstimulated PMC cultures. Cell-fractionation experiments indicated that monocytes were the main cell population accounting for increased TRAP mRNA transcripts, and this was confirmed by histochemical staining for TRAP enzyme activity. Because expression of other iron-binding and -transport proteins is controlled by iron availability, we examined the role of iron in regulating TRAP expression. Increase of TRAP mRNA transcripts in PMCs was inhibited by 50 μM desferrioxamine, a potent iron chelator. The 5' flanking region of the TRAP gene was cloned from a mouse genomic library. In preliminary transient transfection experiments, it was determined that the 5'-flanking region of the TRAP gene contained iron-responsive elements. Therefore, a series of stably transfected HRE H9 cell lines was developed bearing genetic constructs containing various segments of the murine TRAP 5' promoter region driving a luciferase reporter gene. Treatment of transfectants with 100 μg/ml iron-saturated human transferrin (FeTF) was performed to assess iron responsiveness of the constructs. Constructs containing a full-length TRAP promoter (comprising base pairs -1846 to +2) responded to FeTF with a 4-5-fold increase of luciferase activity whereas constructs containing only base pairs -363 to +2 of the TRAP promoter did not respond. Constructs containing 1240 or 881 bp of the TRAP promoter gave only a 1.5- to 2-fold increase of luciferase activity with FeTF. In all cases, increase of luciferase activity was blocked by desferrioxamine. Cells transfected with another luciferase construct driven by a simian virus 40 promoter did not show any increase of luciferase activity with FeTF. These data indicate that expression of TRAP is regulated by iron and that this regulation is exerted at the level of gene transcription. The transfection experiments also suggest that the region of the TRAP 5'-flanking sequence between base pairs -1846 and -1240 contains an iron regulatory element.

Original languageEnglish (US)
Pages (from-to)421-425
Number of pages5
JournalBiochemical Journal
Volume298
Issue number2
StatePublished - 1994
Externally publishedYes

Fingerprint

Acid Phosphatase
Iron
Genes
Luciferases
5' Flanking Region
Base Pairing
tartaric acid
Tartrate-Resistant Acid Phosphatase
Deferoxamine
Cells
Messenger RNA
Transfection
Carrier Proteins
Bearings (structural)
Iron-Binding Proteins
Cell Fractionation
Hairy Cell Leukemia
Cell Line
Lymphocytes
Genomic Library

ASJC Scopus subject areas

  • Biochemistry

Cite this

Transcriptional regulation of the tartrate-resistant acid phosphatase (TRAP) gene by iron. / Alcantara, O.; Reddy, S. V.; Roodman, G. David; Boldt, D. H.

In: Biochemical Journal, Vol. 298, No. 2, 1994, p. 421-425.

Research output: Contribution to journalArticle

Alcantara, O. ; Reddy, S. V. ; Roodman, G. David ; Boldt, D. H. / Transcriptional regulation of the tartrate-resistant acid phosphatase (TRAP) gene by iron. In: Biochemical Journal. 1994 ; Vol. 298, No. 2. pp. 421-425.
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