SUBCELLULAR REDISTRIBUTION IS INVOLVED IN ACUTE REGULATION OF THE BRUSH-BORDER NA+ H+ EXCHANGER ISOFORM 3 IN HUMAN COLON ADENOCARCINOMA CELL-LINE CACO-2 - PROTEIN-KINASE C-MEDIATED INHIBITION OF THE EXCHANGER/

Na+/H+ exchanger isoform 3 (NHE3), an epithelial brush border isoform of the Na+/H+ exchanger gene family, plays an important role in reabso rption of Na+ in the small intestine, the colon, and the kidney. In se veral cell types, phorbol 12-myristate 13-acetate (PMA) acutely inhibi ts NHE3 activity by changes in V-max, but the mechanism of this inhibi tion is unknown. We investigated the role of subcellular redistributio n of NHE3 in the PMA-induced inhibition of endogenous brush border NHE 3 in a model human colon adenocarcinoma cell line, Caco-2. Subcellular localization of NHE3 was examined by confocal morphometric analysis c omplemented with cell surface biotinylation and compared with NHE3 act ivity evaluated by fluorometric measurement of intracellular pH. PMA i nhibited NHE3 activity by 28% (p < 0.01), which was associated with a decrease of the ratio of the brush border/subapical cytoplasmic compar tment of NHE3 from similar to 4.3 to similar to 2.4. This translocatio n resulted in 10-15% of the total cell NHE3 being shifted from the bru sh border pool to the cytoplasmic pool. These effects were mediated by protein kinase C, since they were blocked by the protein kinase C inh ibitor H7. We conclude that inhibition of NHE3 by protein kinase C in Caco-2 cells involves redistribution of the exchanger from brush borde r into a subapical cytoplasmic compartment, and that this mechanism co ntributes similar to 50% to the overall protein kinase C-induced inhib ition of the exchanger.