Hypertonic activation of AE2 anion exchanger in Xenopus oocytes via NHE-mediated intracellular alkalinization.
Xenopus oocytes express endogenous Na+/H+ exchange activity but lack significant endogenous Cl-/HCO3- exchange activity. Coupled operation of Na+/H+ exchange and Cl-/HCO3- exchange contributes in many cell types to the cellular response to hypertonic stress. We therefore examined in Xenopus oocytes the osmotic regulation of chloride transport mediated by recombinant anion exchanger proteins AE2 and AE1. Hypotonicity was without effect on either anion transporter. Hypertonicity activated AE2-associated 36Cl- influx and efflux in a time- and osmolarity-dependent manner, whether incremental osmoles were charged or uncharged, but had no measurable effect on AE1 function. Hypertonic stimulation of AE2 was completely inhibited by Na+ removal or by addition of amiloride. In contrast, neither maneuver altered isotonic activity of AE2. Hypertonicity also induced amiloride-sensitive elevation of oocyte intracellular pH (pHi), and shifted the sigmoidal relationship of extracellular pH vs. AE2 activity > or = 0.5 units to the acid. Injection of pH 7.4 buffer into oocytes attenuated both hypertonic alkalinization and activation of AE2-associated 36Cl- influx, without inhibition of isotonic AE2 function. These data demonstrate that recombinant AE2 expressed in Xenopus oocytes is activated by increased pHi and that hypertonic activation of AE2 is secondary to hypertonic activation of Na+/H+ exchange.
Molecular Medicine Unit, Beth Israel Hospital, Boston, Massachusetts 02215.
B D
BD
Humphreys
L
L
Jiang
M N
MN
Chernova
S L
SL
Alper
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000468:Alkalies / Q000378:metabolism*; D000818:Animals; D027321:Anion Transport Proteins; D017920:Antiporters; D001692:Biological Transport / Q000187:drug effects; D005260:Female; D006863:Hydrogen-Ion Concentration; D006982:Hypertonic Solutions / Q000494:pharmacology*; D007038:Hypotonic Solutions / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism*; D064606:SLC4A Proteins; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000502:physiology*; D014982:Xenopus laevis
D000468:Alkalies; D027321:Anion Transport Proteins; D017920:Antiporters; D006982:Hypertonic Solutions; D007038:Hypotonic Solutions; D008565:Membrane Proteins; D064606:SLC4A Proteins; D017923:Sodium-Hydrogen Exchangers; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid