Cyclic AMP regulates aquaporin 5 expression at both transcriptional and post-transcriptional levels through a protein kinase A pathway.
The membrane water channel aquaporin 5 (AQP5) plays an important role in transporting water across the apical surface of the alveolar epithelium and the epithelia of submucosal glands in the upper airway and nasopharynx. It is thus a potentially important target for modulating the fluid content of upper airway and nasopharyngeal secretions in disorders such as cystic fibrosis. Here, we have used an in vitro system to identify a molecular mechanism through which transcriptional and post-transcriptional regulation of AQP5 is accomplished. In a murine lung epithelial cell line (MLE-12), the addition of chlorophenylthio-cAMP (cpt-cAMP) causes a 4-fold increase in AQP5 mRNA and protein levels and induces translocation of AQP5 to the apical plasma membrane. Treatment with forskolin and isoproternol also caused similar increases in AQP5 expression both in vitro and in mouse lung tissue slices. The addition of actinomycin D abolished the cAMP-mediated increase in AQP5 mRNA and showed that there was no increase in the half-life of AQP5 mRNA, and inhibition of protein kinase A by H-89 blocked the cpt-cAMP-mediated increase of AQP5. Pretreatment of cells with cycloheximide blocked the cpt-cAMP-mediated increase of AQP5 mRNA, indicating that de novo protein synthesis is essential for increased AQP5 transcription. Immunofluorescent micrographs of cells treated with cpt-cAMP showed a significantly stronger AQP5 signal at the plasma membrane as compared with untreated cells. These results show that cAMP regulates AQP5 at multiple levels, by increasing synthesis of AQP5 mRNA and by triggering translocation of AQP5 to the plasma membrane.
Yang
Fan
F
University of Cincinnati College of Medicine, Department of Molecular Genetics, Biochemistry and Microbiology, Cincinnati, Ohio 45267, USA.
Kawedia
Jitesh D
JD
Menon
Anil G
AG
D051402:Aquaporin 5; D020346:Aquaporins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D007545:Isoproterenol / Q000494:pharmacology; D008565:Membrane Proteins; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D051402:Aquaporin 5; D020346:Aquaporins; D008565:Membrane Proteins; D012333:RNA, Messenger; D005576:Colforsin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D007545:Isoproterenol