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Identification of a critical regulatory site in the human interleukin-3
promoter by in vivo footprinting
S Cameron, DS Taylor, EC TePas, NA Speck and B Mathey-Prevot
Division of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA
02115.
Interleukin-3 (IL-3) is involved in proliferation and differentiation of
hematopoietic progenitor cells. Its expression is subject to precise,
tissue-specific regulation, and has been studied extensively in the gibbon
T-cell line MLA 144 by a combination of functional assays and DNA binding
experiments. To extend these studies, the gibbon IL-3 promoter was cloned
and in vivo footprinting of the gibbon and human IL- 3 proximal promoters
was performed. The gibbon IL-3 promoter was found to be highly homologous
to its human counterpart and both promoters yielded identical in vivo
footprints after induction of IL-3 synthesis. In particular, we observed
specific protection of three guanines over a core sequence TGTGGTTT
(IF-1IL3) that had not been recognized in previous studies. IF-1IL3 is not
found in other cytokine promoters, but it is conserved in the IL-3 promoter
of several species and is similar to a recurring motif in viral and
T-cell-specific cellular enhancers. IF-1IL3 binds a specific complex in MLA
144 and Jurkat nuclear extracts in vitro, which shares the same specificity
as the complex bound by the polyoma virus and T-cell receptor delta
enhancers. Mutation of the three guanines in IF-1IL3 core sequence disrupts
binding in vitro and abrogates the ability of the IL-3 promoter to mediate
inducible expression in T cells. Although IF-1IL3 is necessary for IL-3
expression, it is not sufficient: a truncated IL-3 promoter with an intact
IF-1IL3 site but no other activator sites is transcriptionally silent.
These studies describe a new regulatory element within the IL-3 promoter
that is essential for expression and conserved between species.
Volume 83,
Issue 10,
pp. 2851-2859,
05/15/1994
Copyright © 1994 by The American Society of Hematology
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