Constitutive and regulated membrane expression of aquaporin 1 and aquaporin 2 water channels in stably transfected LLC-PK1 epithelial cells.

摘要: Abstract The aquaporins (AQPs) are a family of homologous water-channel proteins that can be inserted into epithelial cell plasma membranes either constitutively (AQP1) or by regulated exocytosis following vasopressin stimulation (AQP2). LLC-PK1 porcine renal cells were stably transfected with cDNA encoding AQP2 (tagged C-terminal c-Myc epitope) rat kidney AQP1 in an expression vector containing cytomegalovirus promoter. Immunofluorescence staining revealed was mainly localized to the membrane, whereas predominantly located on intracellular vesicles. After treatment forskolin for 10 min, relocated indicating this relocation induced cAMP. The location did not change. basal water permeability AQP1-transfected 2-fold greater than nontransfected cells, AQP2-transfected increased significantly only after treatment. Endocytotic uptake fluorescein isothiocyanate-coupled dextran stimulated 6-fold but slightly wild-type cells. This vasopressin-induced endocytosis inhibited low-K+ medium, which selectively affects clathrin-mediated endocytosis. These channel-transfected represent vitro system will allow detailed dissection mechanisms involved processing, targeting, and trafficking via constitutive versus transport pathways.