C/EBP transcription factors in lung cellular differentiation and development

During embryonic development the lung is lined with a primitive epithelium. As development proceeds, the epithelium matures and differentiates into several cell types that together with cells of mesenchymal origin constitute the adult lung. From studies investigating the regulation of differentiation-dependent genes in the lung, such as the Clara Cell Secretory Protein (Ccsp/Scgb1a1), a role for CCAAT/enhancer binder protein (C/EBP) transcription factors in lung cellular differentiation has been suggested.

As C/EBPs have been shown to be important regulators of development, differentiation and proliferation in other organs, their expression was investigated during mouse lung development. C/EBPalpha, C/EBPbeta and C/EBPdelta were detected from late pseudoglandular stage and expression increased prior to birth, correlating with the extensive cellular differentiation, and onset of Ccsp/Scgb1a1 expression, that occurs during this period.

When investigating combinatorial actions of C/EBPs and other transcription factors in the regulation of Ccsp/Scgb1a1, C/EBPalpha was found to synergistically transactivate the promoter together with the epithelial-specific transcription factor NKX2.1. Together with the expression-pattern of C/EBPalpha, this indicates that the synergy could be a major determinant for the high-level, epithelial-specific expression of Ccsp/Scgb1a1 in adult lung, and for the onset of Ccsp/Scgb1a1 during development.

To further examine the role of C/EBPalpha during lung development we generated transgenic mice ectopically expressing C/EBPalpha in the lung epithelium using the human Surfactant protein-C promoter, giving a premature, more widespread C/EBPalpha expression. Lungs from these mice were characterized by fewer and larger developing epithelial tubules. However no defects in overall proliferation or cellular differentiation were observed. A similar phenotype was observed in Cebpa-/- mice and we suggest that these similar phenotypes could possibly stem from dysregulation of airway branching. From our results we conclude that C/EBPalpha has a role in the later stages of lung development.

Glucocorticoids stimulate cellular differentiation during late lung development. In other organs, a role for C/EBPs in glucocorticoid signaling has been suggested. We investigated the role of C/EBPs in the action of glucocorticoids in the lung epithelium by studying the regulation of the Ccsp/Scgb1a1 and CYP2B1 genes. Both these genes are positively regulated by glucocorticoids and C/EBPs. However functional binding sites for the glucorticoid receptor have not been found in their promoters.

In transient transfection studies using a lung epithelial cell line, we found that glucocorticoids induced expression from the Ccsp/Scgb1a1 and CYP2B1 promoters. In both promoters, induction was lost when the C/EBP-binding sites were mutated. Electro-phoretic mobility shift assays revealed that glucocorticoids increased the DNA-binding activity of C/EBPbeta and C/EBPdelta within 10 minutes. The effect was mediated through the glucocorticoid receptor, independent of protein synthesis and involved phosphorylation of C/EBPbeta at residue Thr235.

We further demonstrated that C/EBPbeta is the predominant C/EBP-factor in both human and mouse lung epithelium and that glucocorticoids increase DNA-binding of C/EBPbeta in primary cells from mouse lung. These results indicate a previously unknown role for C/EBP-transcription factors in glucocorticoid signaling in the lung epithelium.

List of scientific papers

I. Cassel TN, Berg T, Suske G, Nord M (2002). Synergistic transactivation of the differentiation-dependent lung gene Clara cell secretory protein (secretoglobin 1a1) by the basic region leucine zipper factor CCAAT/enhancer-binding protein alpha and the homeodomain factor Nkx2.1/thyroid transcription factor-1. J Biol Chem. 277(40): 36970-7. Epub 2002 Aug 2
https://pubmed.ncbi.nlm.nih.gov/12161423

II. Berg T, Nord M (2005). Ectopic expression of the transcription factor C/EBPalpha in the lung epithelium disrupts late lung development. [Manuscript]

III. Berg T, Cassel TN, Schwarze PE, Nord M (2002). Glucocorticoids regulate the CCSP and CYP2B1 promoters via C/EBPbeta and delta in lung cells. Biochem Biophys Res Commun. 293(3): 907-12.
https://pubmed.ncbi.nlm.nih.gov/12051744

IV. Berg T, Didon L, Barton J, Andersson O, Nord M (2005). Glucocorticoids increase C/EBPbeta activity in the lung epithelium via phosphorylation. Biochem Biophys Res Commun. 334(2): 638-45.
https://pubmed.ncbi.nlm.nih.gov/16009338