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Functional properties of the beta-globin locus control region in K562
erythroleukemia cells
AM Moon and TJ Ley
Department of Medicine, Jewish Hospital, Washington University Medical
Center, St Louis, MO 63110.
In this report, we compare the function of the human beta-globin locus
control region (LCR) in three K562 erythroleukemia cell assays, including
(1) a transient transfection assay for "classical" enhancer activity, (2) a
colony assay that detects "productive integration events," and (3) an assay
that detects the ability of LCR fragments to confer hemin inducibility on
linked, stably integrated gamma-globin promoters. Various LCR fragments
were inserted into an expression vector consisting of an A gamma-globin
promoter driving the neomycin phosphotransferase gene (gamma-neo). Using
these vectors, we determined that a 2.5-kb DNA fragment containing LCR
sites I through IV (previously named mu locus activation region [mu LAR])
had activity in all three assays; of the individual LCR sites, only site II
was highly active in all three assays. One region within site II,
consisting of tandem AP-1/NF-E2 consensus elements, had approximately 10%
as much colony assay activity as the entire mu LAR. However, this region
did not have detectable activity in a transient enhancer assay in uninduced
K562 cells, nor was it capable of conferring hemin inducibility on linked
gamma-globin promoters in stably transfected cells. Finally, we tested the
ability of the mu LAR to activate promoters (beta-globin and cathepsin G)
that are not normally expressed in K562 cells. beta-neo was minimally
activated by the mu LAR in transient transfection experiments. The mu LAR
increased the number of stably transfected colonies produced by beta-neo,
but the absolute number of beta-neo colonies, with or without the mu LAR,
was approximately 10% to 20% that of gamma-neo. In contrast, a minimal
cathepsin G promoter was activated by the mu LAR in K562 cells. Our results
suggest that LCR functions are dependent in part on the environments and
the promoters with which the LCR is tested.
Volume 77,
Issue 10,
pp. 2272-2284,
05/15/1991
Copyright © 1991 by The American Society of Hematology
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