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Hematol J. 2001;2(3):150-60.

A minimal serpin promoter with high activity in haematopoietic progenitors and activated T cells.

Author information

1
CRC Department of Hematopoietic Cell and Gene Therapeutics, Paterson Institute for Cancer Research, Wilmslow Road, Manchester M20 4BX, UK. lynne.hampson@man.ac.uk

Abstract

INTRODUCTION:

The serine protease inhibitor Serpin 2A is highly expressed in ex vivo bipotent granulocyte/macrophage progenitor cells and in cultured myeloid stem cells. The gene undergoes rapid down-regulation as these cells are induced to differentiate, and constitutive expression in cultured myeloid stem cells retards maturation. Serpin 2A is also expressed in T cells as a consequence of activation. We now report analysis of the upstream regulatory elements that control Serpin 2A transcription.

MATERIALS AND METHODS:

Using primer extension and rapid amplification of cDNA ends the transcription start site of the Serpin 2A gene was mapped, and a 1.2 Kb genomic upstream fragment cloned and sequenced. Promoter activity and protein binding of deletion and site-directed mutant constructs were analysed by transient transfection and by electrophoretic mobility shift assays.

RESULTS:

A minimal promoter fragment was identified with high activity dependent on NF-kappa and Moloney murine leukaemia enhancer factor LVa binding sites in both myeloid stem cells and activated T cells. NF-kappa was shown to be the main DNA binding protein in T cells, whereas that in haematopoietic stem cells appears to be novel.

CONCLUSION:

Serpin 2A promoter activity in T cells is due predominantly to NF-kappa binding to its consensus site. Activity in haematopoietic stem cells appears to be mediated by a novel protein, which recognises the NF-kappa consensus only in the context of flanking sequences. This concise regulatory element may be of potential value in gene therapeutic applications.

PMID:
11920240
DOI:
10.1038/sj.thj.6200102
[Indexed for MEDLINE]

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