Dexamethasone inhibits the binding of nuclear factors to the IL-5 promoter in human CD4 T cells.
BACKGROUND
IL-5 is produced by the T(H)2 subset of CD4(+) T lymphocytes and is necessary for the eosinophilia typical of allergic conditions. Glucocorticoids such as dexamethasone are highly effective inhibitors of eosinophilic inflammation, and one of their effects is inhibition of IL-5 gene expression.


OBJECTIVE
We wished to examine the effect of dexamethasone on the binding of nuclear factors from primary human CD4(+) T lymphocytes to the RE-I and RE-II positively acting regulatory elements of the IL-5 promoter.


METHODS
CD4(+) T cells, purified from PBMCs by magnetic bead separation, were activated with anti-CD3 antibody and phorbol myristate acetate. Nuclear extracts were tested in electrophoretic mobility shift assays with probes based on RE-I and RE-II.


RESULTS
In extracts from activated cells, the RE-II region of the promoter formed a complex that was shown by supershift assay to contain NFATc. This complex was abolished by treatment of the cells with dexamethasone before activation and was weak or absent in unactivated cells. By contrast, binding to the RE-I region and to the GATA-3 site within RE-I was observed in resting cells and was not affected by activation or treatment with dexamethasone.


CONCLUSIONS
Dexamethasone inhibits the inducible binding of factors to the RE-II region but does not affect the constitutive binding to the RE-I region. Characterization of such molecular effects of glucocorticoids could enable the development of specific inhibitors of IL-5 expression that lack the side effects of glucocorticoids.
Quan
A
A
Centre for Immunology, St Vincent's Hospital and University of New South Wales, Australia.
McCall
M N
MN
Sewell
W A
WA
D001667:Binding, Competitive; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D004268:DNA-Binding Proteins / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology*; D050990:GATA3 Transcription Factor; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D015848:Interleukin-5 / Q000235:genetics*; D016853:Muromonab-CD3 / Q000494:pharmacology; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding / Q000187:drug effects; D020218:Response Elements; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015534:Trans-Activators / Q000378:metabolism
D004268:DNA-Binding Proteins; D050990:GATA3 Transcription Factor; C494749:GATA3 protein, human; D005938:Glucocorticoids; D015848:Interleukin-5; D016853:Muromonab-CD3; D009687:Nuclear Proteins; D015534:Trans-Activators; D003907:Dexamethasone; D013755:Tetradecanoylphorbol Acetate