Regulatory phosphorylation of the secretory Na-K-Cl cotransporter: modulation by cytoplasmic Cl
Lytle, C.; Forbush, B.
American Journal of Physiology 270(2 Pt 1): C437-C448
1996
ISSN/ISBN: 0002-9513 PMID: 8779905 Document Number: 455024
The effect of cytoplasmic Cl concentration ((Cl)-i) on the activation state ((3H)benzmetanide binding rate) and phosphorylation state (32P incorporation) of the Na-K-Cl cotransporter was evaluated in secretory tubules isolated from the dogfish shark rectal gland. Reduction of (CI)-i at relatively constant cell volume (by removal of extracellular Cl or Na or by addition of bumetanide) increased cotransporter activation and phosphorylation. Raising extracellular K concentration ((K)-o) from 4 to 80 mM, a maneuver that elevated (CI)-i above normal, reduced basal cotransport activity and rendered it entirely refractory to forskolin. High (K)-o also blocked activation and phosphorylation in response to cell shrinkage, despite the fact that (CI)-i was already greatly elevated as a consequence of osmotic water loss. The phosphatase inhibitor calyculin A also promoted activation, but not in cells preexposed briefly to high (K)-o. In summary, maneuvers that lower (CI)-i activate the cotransporter, whereas those that elevate (CI)-i (or prevent it from decreasing) block activation in response to secretory stimuli. Cell Cl appears to govern its own rate of entry via Na-K-Cl cotransport by impeding regulatory phosphorylation of the Na-K-Cl cotransport protein.