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Phosphorylation of serine 256 is required for cAMP-dependent regulatory exocytosis of the aquaporin-2 water channel
- Source :
- The Journal of biological chemistry. 272(23)
- Publication Year :
- 1997
-
Abstract
- The aquaporin-2 (AQP2) vasopressin water channel is translocated to the apical membrane upon vasopressin stimulation. Phosphorylation of serine 256 of AQP2 by cAMP-dependent protein kinase has been shown, but its relation to vasopressin-regulated translocation has not been elucidated. To address this question, wild type (WT) AQP2 and a mutant with alanine in place of serine 256 of AQP2 (S256A) were expressed in LLC-PK1 cells by electroporation. Measurements by a stopped-flow light-scattering method revealed that the osmotic water permeability (Pf) of LLC-PK1 cells transfected with WT was 69.6 +/- 6.5 microm/s (24.8 +/- 2.2 microm/s for mock-transfected), and stimulation by 500 microM 8-(4-chlorophenylthio)-cAMP increased the Pf by 85 +/- 12%. When S256A AQP2 was transfected, the cAMP-dependent increase in the Pf was only 8 +/- 5%. After cAMP stimulation, the increase in surface expression of AQP2 determined by surface biotin labeling was 4 +/- 10%, significantly less than that for WT (88 +/- 5%). In addition, an in vivo [32P]orthophosphate labeling assay demonstrated significant phosphorylation of WT AQP2 and only minimal phosphorylation of S256A AQP2 in LLC-PK1 cells. Our results indicated that serine 256 of AQP2 is necessary for regulatory exocytosis and that cAMP-responsive redistribution of AQP2 may be regulated by phosphorylation of AQP2.
- Subjects :
- Light
Stimulation
Biology
urologic and male genital diseases
Aquaporins
Transfection
Biochemistry
Exocytosis
Ion Channels
Cell Line
Serine
Cyclic AMP
Animals
Point Mutation
Scattering, Radiation
Phosphorylation
Protein kinase A
Molecular Biology
Alanine
Aquaporin 2
urogenital system
Wild type
Cell Biology
Apical membrane
Thionucleotides
Molecular biology
Aquaporin 6
Recombinant Proteins
Cell biology
Rats
Mutagenesis, Site-Directed
Subjects
Details
- ISSN :
- 00219258
- Volume :
- 272
- Issue :
- 23
- Database :
- OpenAIRE
- Journal :
- The Journal of biological chemistry
- Accession number :
- edsair.doi.dedup.....5e43902be62ebeaaa97df89bd0d9d0fc