Molecular regulation of granulocyte macrophage colony-stimulating factor in human lung epithelial cells by interleukin (IL)-1beta, IL-4, and IL-13 involves both transcriptional and post-transcriptional mechanisms.
Interleukin (IL)-1beta stimulates the release of granulocyte macrophage colony-stimulating factor (GM-CSF) from lung epithelial cells. To investigate the molecular mechanisms underlying GM-CSF regulation, we studied GM-CSF production, messenger RNA (mRNA) expression levels, and GM-CSF promoter activity in A549 human alveolar carcinoma cells stimulated with IL-1beta. Coincubation with IL-4 or IL-13 dose-dependently inhibited IL-1beta-induced GM-CSF release. Time-course studies of intracellular and extracellular protein release and mRNA expression indicated tight coupling of protein and mRNA synthesis within 6 h after stimulation. IL-4 and IL-13 both inhibited expression of GM-CSF mRNA and protein by 2 h after stimulation. Stable transfection of A549 cells, with GM-CSF promoter/ enhancer constructs containing up to 3.3 kb upstream of the transcription start site, revealed maximal activation by IL-1beta and phorbol 12-myristate 13-acetate (PMA) with a reporter containing the proximal promoter (-627 to +35). This excludes sequences further upstream from a major regulatory role in GM-CSF promoter activation by IL-1beta or PMA in these cells. IL-4 and IL-13 downregulated promoter activation but had no effect on GM-CSF mRNA half-life. However, IL-1beta activation of all constructs was far less pronounced than in Jurkat T cells, suggesting a requirement for additional mechanisms, possibly post-transcriptional, to potentiate the observed transcriptional induction.
Bergmann
M
M
Franz-Volhard Clinic at Max-Delbrück Center, Charité, Humboldt University, Berlin, Germany.
Barnes
P J
PJ
Newton
R
R
D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D018793:Interleukin-13 / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology; D007378:Interleukins / Q000494:pharmacology*; D019169:Jurkat Cells; D008168:Lung / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D014407:Tumor Cells, Cultured
D007375:Interleukin-1; D018793:Interleukin-13; D007378:Interleukins; D012333:RNA, Messenger; D003609:Dactinomycin; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D003513:Cycloheximide; D013755:Tetradecanoylphorbol Acetate