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Modulation of CFTR gene expression in HT-29 cells by extracellular hyperosmolarity
M Baudouin-Legros1, F Brouillard, M Cougnon
1Institut National de la Santé et de la Recherche Médicale Unité 467, Faculté de Médecine Necker-Enfants Malades, 75015 Paris, France. legros@necker.fr
American Journal of Physiology. Cell Physiology
|January 25, 2000
Summary
Hypertonicity, or high salt conditions, decreases cystic fibrosis transmembrane conductance regulator (CFTR) mRNA levels in colon cells. This reduction in CFTR gene expression is mediated by the p38 kinase pathway and occurs at the transcriptional level.
Area of Science:
- Cell Biology
- Molecular Biology
- Physiology
Background:
- Hypertonicity impacts cell functions, including gene expression.
- Cystic fibrosis transmembrane conductance regulator (CFTR) is crucial for ion transport in secretory epithelia, often exposed to hypertonic environments.
Purpose of the Study:
- To investigate the effect of hypertonicity on cystic fibrosis transmembrane conductance regulator (CFTR) gene expression.
- To elucidate the molecular mechanisms underlying CFTR regulation by hyperosmotic stress.
Main Methods:
- HT-29 colon cells were cultured under various hypertonic conditions (NaCl, urea, mannitol).
- Kinase inhibitors (PKA, PKC, serine/threonine, tyrosine, p38 MAPK) were used to identify signaling pathways.
- Luciferase reporter assays assessed CFTR promoter activity in transfected HeLa cells.
Main Results:
- Hypertonicity ( >150 mosmol/l) significantly reduced CFTR mRNA levels in a time-dependent manner.
- The decrease in CFTR gene expression by NaCl required p38 kinase cascade activity.
- NaCl diminished CFTR promoter-driven luciferase activity, indicating transcriptional regulation.
- This transcriptional regulation necessitates protein synthesis.
Conclusions:
- Hypertonic stress downregulates CFTR gene expression at the transcriptional level.
- The p38 kinase pathway is essential for mediating this hypertonicity-induced decrease in CFTR mRNA.
- The observed decrease in CFTR mRNA represents a general cellular response to hyperosmolar stress.