Regulation of the rabbit ileal brush-border Na+/H+ exchanger by an ATP-requiring Ca++/calmodulin-mediated process

R. P. Rood, E. Emmer, J. Wesolek, J. McCullen, Z. Husain, M. E. Cohen, R. S. Braithwaite, H. Murer, G. W.G. Sharp, M. Donowitz

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Abstract

Brush-border vesicles purified from rabbit ileal villus cells were used to evaluate how Ca++/calmodulin (CaM) regulates the neutral linked NaCl absorptive process, part of which is a Na+/H+ exchanger. After freezing and thawing to allow incorporation of macromolecules into the vesicles, the effect of Ca++/CaM on brush-border Na+ uptake with an acid inside pH gradient, and on Na+/H+ exchange was determined. Freezing and thawing vesicles with 0.85 μM free Ca++ plus 5 μM exogenous CaM failed to alter Na+/H+ exchange as did the addition of exogenous ATP plus an ATP regenerating system, which was sufficient to elevate intravesicular ATP to 47 μM from a basal level of 0.4 μM. However, the combination of Ca++/CaM plus ATP inhibited Na+ uptake in the presence of an acid inside pH gradient and inhibited Na+/H+ exchange, while Na+ uptake in the absence of a pH gradient was not altered. This effect required a hydrolyzable form of ATP, and did not occur when the nonhydrolyzable ATP analogue, AMP-PNP, replaced ATP. Under the identical intravesicular conditions used for the transport studies, Ca++ (0.85 μM) plus exogenous CaM (5 μM), in the presence of magnesium plus ATP, increased phosphorylation of five brush-border peptides. These data are consistent with Ca++/CaM acting via phosphorylation to regulate the ileal brush-border Na+/H+ exchanger.

Original languageEnglish
Pages (from-to)1091-1097
Number of pages7
JournalJournal of Clinical Investigation
Volume82
Issue number3
DOIs
StatePublished - Jan 1 1988
Externally publishedYes

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    Rood, R. P., Emmer, E., Wesolek, J., McCullen, J., Husain, Z., Cohen, M. E., Braithwaite, R. S., Murer, H., Sharp, G. W. G., & Donowitz, M. (1988). Regulation of the rabbit ileal brush-border Na+/H+ exchanger by an ATP-requiring Ca++/calmodulin-mediated process. Journal of Clinical Investigation, 82(3), 1091-1097. https://doi.org/10.1172/JCI113665