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Inhibition of endocytosis causes phosphorylation (S256)-independent plasma membrane accumulation of AQP2.
- Source :
-
American journal of physiology. Renal physiology [Am J Physiol Renal Physiol] 2004 Feb; Vol. 286 (2), pp. F233-43. Date of Electronic Publication: 2003 Sep 30. - Publication Year :
- 2004
-
Abstract
- Inhibition of clathrin-mediated endocytosis by expression of a GTPase-deficient dynamin mutant (dynamin-2/K44A) for 16 h results in an accumulation of plasma membrane aquaporin-2 (AQP2) in epithelial cells stably transfected with wild-type AQP2. We now show a similar effect of K44A dynamin in LLC-PK1 cells transfected with an S256 phosphorylation-deficient AQP2 mutant, AQP2(S256A), and in AQP2-transfected inner medullary collecting duct (IMCD) cells. More acute blockade of endocytosis in these cells with the cholesterol-depleting agent methyl-beta-cyclodextrin (mbetaCD; 10 mM) resulted in a rapid and extensive cell-surface accumulation of both wild-type AQP2 and AQP2 (S256A) within 15 min after treatment. This effect was similar to that induced by treatment of the cells with vasopressin. Blockade of endocytosis by mbetaCD was confirmed using quantitative analysis of FITC-dextran uptake and AQP2 membrane insertion was verified by cell-surface biotinylation. These data indicate that AQP2 recycles constitutively and rapidly between intracellular stores and the cell surface in LLC-PK1 and IMCD cells. The constitutive trafficking process is not dependent on phosphorylation of the serine-256 residue of AQP2, which is, however, an essential step for regulated vasopressin/cAMP-mediated translocation of AQP2. Our data show that rapid and extensive plasma membrane accumulation of AQP2 can occur in a vasopressin receptor (V2R)- and phosphorylation-independent manner, pointing to a potential means of bypassing the mutated V2R in X-linked nephrogenic diabetes insipidus to achieve cell surface expression of AQP2.
- Subjects :
- Adenoviridae genetics
Animals
Aquaporin 2
Aquaporin 6
Aquaporins genetics
Biotinylation
Cell Membrane drug effects
Cell Membrane metabolism
Cholesterol metabolism
Cyclodextrins pharmacology
Dynamin II genetics
Dynamin II metabolism
Endocytosis drug effects
Hybridomas
Kidney Tubules, Collecting metabolism
LLC-PK1 Cells
Phosphorylation
Renal Agents pharmacology
Serine metabolism
Swine
Vasopressins pharmacology
Aquaporins metabolism
Endocytosis physiology
beta-Cyclodextrins
Subjects
Details
- Language :
- English
- ISSN :
- 1931-857X
- Volume :
- 286
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- American journal of physiology. Renal physiology
- Publication Type :
- Academic Journal
- Accession number :
- 14519593
- Full Text :
- https://doi.org/10.1152/ajprenal.00179.2003