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Nucleic Acids Research, 1992, Vol. 20, No. 8 1925-1932
© 1992

MOLECULAR BIOLOGY

The involvement of demethylation in the myeloid-specific function of the mouse M lysozyme gene downstream enhancer

Sabine Klages{dagger}, Bettina Möllers§ and Rainer Renkawitz*

Genzentrum, Max-Planck-lnstitut für Biochemie W-8033 Martinsried, Germany

*To whom correspondence should be addressed at present address: Genetisches Institut der Justus-Liebig-Universität, Heinrich-Buff-Ring 58–62, W-6300 Giessen, Germany

Received January 22, 1992. Revised March 19, 1992. Accepted March 19, 1992.

Lysozyme gene expression is a specific marker for the macrophage/granulocyte lineage of hematopoietic differentiation in mammals, its expression being gradually Increased during maturation. Analysis of the mechanisms regulating mouse M lysozyme gene expression during myeloid differentiation revealed a complicated pattern of DNase I hypersensitive sites (HS sites) within the flanking regions of the gene. The HS-3 site, located in the 3'-flanking region of the gene, overlapped with an enhancer element, which is the only strong enhancer identified in the vicinity of the gene. We demonstrate a positive correlation between undermethylatlon of the entire 3'-flanking region, the appearance of the HS-3 site, and M lysozyme gene expression during in vitro differentiation of hematopoietic stem cells. We furthermore show that methylation of a single CpG site within the enhancer core element, only observed in immature macrophage cells in vivo, is sufficient to inhibit nuclear factor binding to this element in vitro and to inhibit its trans-activation potential in DNA transfectlon experiments.

+Present addresses: Bristol-Myers Squibb Pharmaceutical Research Institute, Department of Molecular Biology, Biochemical Oncology Laboratory, Princeton, NJ 08543–4000, USA

§Present addresses: Boehringer Mannheim GmbH, Sandhofer Strafie 116, W-6800 Mannheim


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