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Transcriptional regulation of interleukin-3 expression in megakaryocytes
S Nimer, J Zhang, H Avraham and Y Miyazaki
Laboratory of Molecular Aspects of Hematopoiesis, Sloan-Kettering
Institute, New York, NY, USA.
Interleukin-3 (IL-3) is a potent stimulator of megakaryocyte proliferation,
and autocrine production of IL-3 by megakaryocytic leukemia cell lines and
bone marrow-derived megakaryocytes has recently been demonstrated. To
characterize the transcriptional regulation of IL- 3 in megakaryocytes, we
transiently transfected IL-3 promoter CAT constructs that contain variable
amounts of 5' flanking sequences into the human CMK and CMK-6
megakaryocytic cell lines and identified two positive acting
transcriptional regulatory regions, one located between bp -315 and -284,
which contains consensus AP-1 and ets binding sites, and a second located
between bp -173 and -61. DNase I footprinting assays using CMK or CMK-6
nuclear extracts demonstrate DNA-protein interactions in the identical
region protected by T cell or natural killer cell nuclear extracts (between
bp -165 to -128), and electrophoretic mobility shift assays demonstrate the
binding of proteins to three distinct portions of this region. To
characterize the transcription factors in megakaryocytic cells that could
bind to these two regulatory regions, we performed Northern blot analyses,
which showed the presence of ets-1, elf-1 (which is thought to be
restricted to T cells), NF-IL3A and AML1 mRNAs, as well as c-fos, jun B,
and jun D, but not c-jun mRNA. These studies show that the transcriptional
regulation of IL-3 expression in megakaryocytic leukemia cell lines is
similar, but not identical to normal human T cells.
Volume 88,
Issue 1,
pp. 66-74,
07/01/1996
Copyright © 1996 by The American Society of Hematology
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