Lateral mobility of Na,K-ATPase and membrane lipids in renal cells. Importance of cytoskeletal integrity.
Because membrane fluidity is an important determinant of membrane function, the lateral diffusion rate (DL) of the membrane protein Na,K-ATPase was determined in intact renal proximal tubule epithelial cells by the technique of fluorescence redistribution after photobleaching (FRAP). In normal cells the DL of Na,K-ATPase in the basal membrane was 3.31 x 10(-10) cm2/sec. Treatment with cytochalasin D to promote actin filament depolymerization caused a sevenfold increase in DL. Exposure of cells to a Ca(2+)-free medium or to hypoxia and reoxygenation, which have similar disruptive effects on the cytoskeleton, also caused increases in DL. Disruption of actin microfilament structure also increased the mobile fraction of Na,K-ATPase. Using a confocal laser microscopic technique only 14.9% of total Na,K-ATPase was observed to reside in the apical membrane domain of normal cells. Microfilament depolymerization caused this fraction to increase to 47.7%. Thus, the translocation of Na,K-ATPase from the basolateral to the apical domain induced by cytoskeletal protein dysfunction was enabled by an increased rate of lateral diffusion of Na,K-ATPase. The behavior of a variety of membrane lipids following actin depolymerization was more heterogeneous. Some lipids showed a similar increase in DL, whereas others showed very little dependence upon the cytoskeleton for lateral restraint.
Paller
M S
MS
Department of Medicine, University of Minnesota, Minneapolis 55455.
D008841:Actin Cytoskeleton / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D000199:Actins / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D015687:Cell Hypoxia; D016764:Cell Polarity; D002478:Cells, Cultured; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D004058:Diffusion; D004848:Epithelium / Q000378:metabolism / Q000648:ultrastructure; D005456:Fluorescent Dyes; D007687:Kidney Tubules, Proximal / Q000378:metabolism* / Q000648:ultrastructure; D008560:Membrane Fluidity; D008563:Membrane Lipids / Q000378:metabolism*; D018613:Microscopy, Confocal; D011108:Polymers; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D000199:Actins; D005456:Fluorescent Dyes; D008563:Membrane Lipids; D011108:Polymers; D015638:Cytochalasin D; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium