Molecular cloning of NHE1 from winter flounder RBCs: activation by osmotic shrinkage, cAMP, and calyculin A.
In this report, we describe the cloning, cellular localization, and functional characteristics of Na(+)/H(+) exchanger 1 (NHE1) from red blood cells of the winter flounder Pseudopleuronectes americanus (paNHE1). The paNHE1 protein localizes primarily to the marginal band and exhibits a 74% similarity to the trout beta-NHE, and 65% to the human NHE1 (hNHE1). Functionally, paNHE1 shares characteristics of both beta-NHE and hNHE1 in that it is activated both by manipulations that increase cAMP and by cell shrinkage, respectively. In accordance, the paNHE1 protein exhibits both protein kinase A consensus sites as in beta-NHE and a region of high homology to that required for shrinkage-dependent activation of hNHE1. After shrinkage-dependent activation of paNHE1 and resulting activation of a Cl(-)/HCO(3)(-) exchanger, their parallel operation results in net uptake of NaCl and osmotically obliged water. Activation of paNHE1 by cAMP is at least additive to that elicited by osmotic shrinkage, suggesting that these stimuli regulate paNHE1 by distinct mechanisms. Finally, exposure to the serine/threonine phosphatase inhibitor calyculin A potently activates paNHE1, and this activation is also additive to that induced by shrinkage or cAMP.
Department of Human Physiology, School of Medicine, University of California-Davis, Davis, California 95616, USA. sfpederson@aki.ku.dk
Stine F
SF
Pedersen
Scott A
SA
King
Robert R
RR
Rigor
Zhenpeng
Z
Zhuang
Jaimie M
JM
Warren
Peter M
PM
Cala
D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism; D048429:Cell Size; D002712:Chlorides / Q000378:metabolism; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000378:metabolism*; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D005432:Flounder / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D008387:Marine Toxins; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D010080:Oxazoles / Q000378:metabolism*; D010802:Phylogeny; D011188:Potassium / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000378:metabolism*
D000318:Adrenergic beta-Agonists; D002712:Chlorides; D004791:Enzyme Inhibitors; D008387:Marine Toxins; D010080:Oxazoles; D017923:Sodium-Hydrogen Exchangers; C069139:growth factor-activatable Na-H exchanger NHE-1; C059041:calyculin A; D012964:Sodium; D000242:Cyclic AMP; D007545:Isoproterenol; D011188:Potassium