Repolarization of Na+-K+ pumps during establishment of epithelial monolayers.
Madin-Darby canine kidney (MDCK) cells plated at confluence and incubated for 20 h in low (5 microM) Ca2+ have no tight junctions (TJs), and their Na+-K+-ATPase is randomly distributed over the surface. On transfer to normal Ca2+ levels (1.8 mM) ("Ca2+ switch"), TJs and transepithelial resistance develop quickly, trapping a considerable fraction (35%) of the surface Na+-K+-ATPase on the apical (incorrect) side. This misplaced enzyme is subsequently removed from this region or inactivated, demonstrating that polarization proceeds despite TJs. Simultaneously, the amount of Na+-K+-ATPase on the basolateral side increases in a higher proportion (125%), than could be accounted for by relocation of the misplaced apical enzyme. This incorporation is prevented by cycloheximide, ammonium chloride, primaquine, or chloroquine, suggesting that Na+-K+-ATPase originates in an intracellular pool and that its surface insertion requires synthesis of new enzyme or of a protein factor, since it is carried to the surface membrane through a mechanism of exocytosis. In summary, asymmetric distribution of ion pumps depends 1) on polarized insertion of Na+-K+-ATPase as well as 2) on removal or inactivation of misplaced enzyme.
Contreras
R G
RG
Center of Research and Advanced Studies, Mexico City D.F., Mexico.
Avila
G
G
Gutierrez
C
C
Bolívar
J J
JJ
González-Mariscal
L
L
Darzon
A
A
Beaty
G
G
Rodriguez-Boulan
E
E
Cereijido
M
M
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D003584:Cytological Techniques; D004594:Electrophysiology; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism* / Q000502:physiology; D007365:Intercellular Junctions / Q000502:physiology; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology*; D010042:Ouabain / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D014018:Tissue Distribution
D007473:Ion Channels; D010042:Ouabain; D003513:Cycloheximide; D000254:Sodium-Potassium-Exchanging ATPase