Phorbol 12-myristate 13-acetate down-regulates Na,K-ATPase independent of its protein kinase C site: Decrease in basolateral cell surface area

被引:37
作者
Beron, J [1 ]
Forster, I [1 ]
Beguin, P [1 ]
Geering, K [1 ]
Verrey, F [1 ]
机构
[1] UNIV LAUSANNE,INST PHARMACOL & TOXICOL,CH-1005 LAUSANNE,SWITZERLAND
关键词
D O I
10.1091/mbc.8.3.387
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
The effect of protein kinase C (PKC) stimulation on the pump current (I-p) generated by the Na,K-ATPase was measured in A6 epithelia apically permeabilized with amphotericin B. Phorbol 12-myristate 13-acetate (PMA) produced a decrease in I-p carried by sodium pumps containing the endogenous Xenopus laevis or transfected Bufo marinus alpha 1 subunits (similar to 30% reduction within 25 min, maximum after 40 min) independent of the PKC phosphorylation site (T15A/S16A). In addition to this major effect of PMA, which was independent of the intracellular sodium concentration and was prevented by the PKC inhibitor bisindolylmaleimide GF 109203X (BIM), another BIM-resistant, PKC site-independent decrease was observed when the I-p was measured at low sodium concentrations (total reduction similar to 50% at 5 mM sodium). Using ouabain binding and cell surface biotinylation, stimulation of PKC was shown to reduce surface Na,K-ATPase by 14 to 20% within 25 min. The same treatment stimulated fluid phase endocytosis sevenfold and decreased by 16.5% the basolateral cell surface area measured by transepithelial capacitance measurements. In conclusion, PKC stimulation produces a decrease in sodium pump function which can be attributed, to a large extent, to a withdrawal of sodium pumps from the basolateral cell surface independent of their PKC site. This reduction of the number of sodium pumps is parallel to a decrease in basolateral membrane area.
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页码:387 / 398
页数:12
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