Molecular regulation of granulocyte macrophage colony-stimulating factor in human lung epithelial cells by interleukin (IL)-1β, IL-4, and IL-13 involves both transcriptional and post-transcriptional mechanisms

Interleukin (IL)-1 beta stimulates the release of granulocyte macrophage colony-stimulating factor (GM-CSF) from lung epithelial cells. To investigate the molecular mechanisms underlying CM-CSF regulation, we studied GM-CSF production, messenger RNA (mRNA) expression levers, and CM-CSF promoter activity in A549 human alveolar carcinoma cells stimulated with IL-1 beta. Coincubation with IL-4 or IL-13 dose-dependently inhibited IL-1 beta-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 CM-CSF mRNA and protein by 2 h after stimulation. Stable transfection of A549 cells, with CM-CSF promoter/enhancer constructs containing up to 3.3 kb upstream of the transcription start site, revealed maximal activation by IL-1 beta 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-1 beta 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-1 beta 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.