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Blood, Vol. 94 No. 7 (October 1), 1999:
pp. 2477-2486
Cloning and Characterization of EphA3 (Hek) Gene Promoter: DNA
Methylation Regulates Expression in Hematopoietic Tumor Cells
Mirella Dottori,
Michelle Down,
Andreas Hüttmann,
David R. Fitzpatrick, and
Andrew W. Boyd
From Queensland Institute of Medical Research and the Department of
Medicine, University of Queensland, PO Royal Brisbane Hospital,
Herston, Queensland, Australia.
The Eph family of receptor tyrosine kinases (RTK) has restricted
temporal and spatial expression patterns during development, and
several members are also found to be upregulated in tumors. Very little
is known of the promoter elements or regulatory factors required for
expression of Eph RTK genes. In this report we describe the
identification and characterization of the EphA3 gene promoter region.
A region of 86 bp located at  348 bp to 262 bp upstream from the
transcription start site was identified as the basal promoter. This
region was shown to be active in both EphA3-expressing and
-nonexpressing cell lines, contrasting with the widely different levels
of EphA3 expression. We noted a region rich in CpG dinucleotides downstream of the basal promoter. Using Southern blot analyses with
methylation-sensitive restriction enzymes and bisulfite sequencing of
genomic DNA, sites of DNA methylation were identified in hematopoietic cell lines which correlated with their levels of EphA3 gene expression. We showed that EphA3 was not methylated in normal tissues but that a
subset of clinical samples from leukemia patients showed extensive
methylation, similar to that observed in cell lines. These results
suggest that DNA methylation may be an important mechanism regulating
EphA3 transcription in hematopoietic tumors.
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