Palytoxin acts on Na(+),K (+)-ATPase but not nongastric H(+),K (+)-ATPase.
Palytoxin (PTX) opens a pathway for ions to pass through Na,K-ATPase. We investigate here whether PTX also acts on nongastric H,K-ATPases. The following combinations of cRNA were expressed in Xenopus laevis oocytes: Bufo marinus bladder H,K-ATPase alpha(2)- and Na,K-ATPase beta(2)-subunits; Bufo Na,K-ATPase alpha(1)- and Na,K-ATPase beta(2)-subunits; and Bufo Na,K-ATPase beta(2)-subunit alone. The response to PTX was measured after blocking endogenous Xenopus Na,K-ATPase with 10 microM ouabain. Functional expression was confirmed by measuring (86)Rb uptake. PTX (5 nM: ) produced a large increase of membrane conductance in oocytes expressing Bufo Na,K-ATPase, but no significant increase occurred in oocytes expressing Bufo H,K-ATPase or in those injected with Bufo beta(2)-subunit alone. Expression of the following combinations of cDNA was investigated in HeLa cells: rat colonic H,K-ATPase alpha(1)-subunit and Na,K-ATPase beta(1)-subunit; rat Na,K-ATPase alpha(2)-subunit and Na,K-ATPase beta(2)-subunit; and rat Na,K-ATPase beta(1)- or Na,K-ATPase beta(2)-subunit alone. Measurement of increases in (86)Rb uptake confirmed that both rat Na,K and H,K pumps were functional in HeLa cells expressing rat colonic HKalpha(1)/NKbeta(1) and NKalpha(2)/NKbeta(2). Whole-cell patch-clamp measurements in HeLa cells expressing rat colonic HKalpha(1)/NKbeta(1) exposed to 100 nM PTX showed no significant increase of membrane current, and there was no membrane conductance increase in HeLa cells transfected with rat NKbeta(1)- or rat NKbeta(2)-subunit alone. However, in HeLa cells expressing rat NKalpha(2)/NKbeta(2), outward current was observed after pump activation by 20 mM K(+) and a large membrane conductance increase occurred after 100 nM PTX. We conclude that nongastric H,K-ATPases are not sensitive to PTX when expressed in these cells, whereas PTX does act on Na,K-ATPase.
Guennoun-Lehmann
Saida
S
Department of Biological Sciences, Ohio University, Athens, OH 45701, USA.
Fonseca
James E
JE
Horisberger
Jean-Daniel
JD
Rakowski
Robert F
RF
D000178:Acrylamides / Q000494:pharmacology*; D000818:Animals; D002025:Bufonidae; D003001:Cloning, Molecular; D003064:Cnidarian Venoms / Q000494:pharmacology*; D003106:Colon / Q000201:enzymology; D017506:H(+)-K(+)-Exchanging ATPase / Q000187:drug effects / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D009865:Oocytes / Q000201:enzymology; D018408:Patch-Clamp Techniques; D021122:Protein Subunits / Q000378:metabolism; D051381:Rats; D012414:Rubidium Radioisotopes; D000254:Sodium-Potassium-Exchanging ATPase / Q000187:drug effects / Q000378:metabolism*; D013270:Stomach / Q000201:enzymology; D001743:Urinary Bladder / Q000201:enzymology; D014982:Xenopus laevis; D000095702:Polyether Toxins
D000178:Acrylamides; D003064:Cnidarian Venoms; D017506:H(+)-K(+)-Exchanging ATPase; D021122:Protein Subunits; D012414:Rubidium Radioisotopes; D000254:Sodium-Potassium-Exchanging ATPase; C010272:palytoxin; D000095702:Polyether Toxins