Regulation of Na+/H+ exchange activity by recruitment of new Na+/H+ antiporters: effect of calyculin A.
The Na+/H+ antiporter of trout red blood cells, beta-NHE, is activated by agonists of the adenosine 3',5'-cyclic monophosphate-dependent protein kinase A (PKA) and by those of protein kinase C (PKC). beta-NHE, once activated, shifts into a refractory state, accounting for its desensitization. It had previously been shown that desensitization is blocked and reversed by the protein phosphatase inhibitor okadaic acid (OA). In this study we examined the effect of another protein phosphatase inhibitor, calyculin A (CIA). CIA was at least 10 times more potent than OA in blocking beta-NHE desensitization, suggesting that desensitization is controlled by phosphatase-1. Furthermore, CIA alone induced a large Na+/H+ exchange in unstimulated red blood cells, a property not shared by OA. The characteristics of ClA-induced Na+/H+ exchange are very different from those of the exchange triggered by activation of beta-NHE by PKA or PKC agonists, i.e., a flat pH dependence and total insensitivity to PKA and PKC inhibitors. Simultaneous addition of maximal concentrations of ClA and catecholamine produced an additive stimulation of the Na+/H+ exchange, consistent with the interpretation that these agents act on two distinct pools of exchangers. Screening of different cDNA libraries suggested that only one isoform of antiporter exists in the trout red blood cell; it therefore seems likely that regulation of the Na+/H+ antiporter beta-NHE involves a recycling mechanism. The reasons why intracellular beta-NHE show different properties from membrane beta-NHE are discussed.
Guizouarn
H
H
Département de Biologie Cellulaire et Moléculaire, Commisseriat à l'Energic Atomique, Villefranche-sur-Mer, France.
Borgese
F
F
Pellissier
B
B
Garcia-Romeu
F
F
Motais
R
R
D000143:Acids / Q000494:pharmacology; D000818:Animals; D000242:Cyclic AMP / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D004988:Ethers, Cyclic / Q000494:pharmacology; D008387:Marine Toxins; D019319:Okadaic Acid; D010080:Oxazoles / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D011493:Protein Kinase C / Q000502:physiology; D054645:Protein Phosphatase 1; D017923:Sodium-Hydrogen Exchangers / Q000145:classification* / Q000378:metabolism*; D013696:Temperature; D014337:Trout
D000143:Acids; D004988:Ethers, Cyclic; D008387:Marine Toxins; D010080:Oxazoles; D017923:Sodium-Hydrogen Exchangers; D019319:Okadaic Acid; C059041:calyculin A; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1