Na+-K+-ATPase is localized to the basolateral cell surface of most epithelial cells. Conflicting results regarding the intracellular trafficking of Na+-K+-ATPase in Madin-Darby canine kidney cells have been reported, with delivery to both apical and basolateral membranes or exclusively to the basolateral cell surface. We examined the delivery and steady-state distribution of Na+K+-ATPase in the amphibian epithelial cell line A6 using an antibody raised against Na+-K+-ATPase α-subunit and sulfo-N-hydroxysuccinimidobiotin to tag cell surface proteins. The steady- state distribution of the Na+-K+-ATPase was basolateral, as confirmed by immunocytochemistry. Delivery of newly synthesized Na+-K+-ATPase to the cell surface was examined using [35S]methionine and [35S]cysteine in a pulse-chase protocol. After a 20-min pulse, the α-subunit and core glycosylated β-subunit were present at both apical and basolateral cell surfaces. The α-subunit and core glycosylated β-subunit delivered to the apical cell surface were degraded within 2 h. Mature α/β-heterodimer was found almost exclusively at the basolateral surface after a 1- to 24-h chase. These data suggest that immature Na+-K+-ATPase α-subunit and core glycosylated β-subunits are not retained in the endoplasmic reticulum of A6 cells and apparently lack sorting signals. Mature Na+-K+-ATPase is targeted to the basolateral surface, suggesting that basolateral targeting of the protein is conformation dependent.
|Number of pages||9|
|Journal||American Journal of Physiology - Cell Physiology|
|Issue number||6 41-6|
|Publication status||Published - 1997|
- Intracellular trafficking
- Sodium pump
- Sodium-potassium adenosinetriphosphatase