Different ionic conditions prompt NHE2 and NHE3 translocation to the plasma membrane.
We tested whether NHE3 and NHE2 Na(+)/H(+) exchanger isoforms were recruited to the plasma membrane (PM) in response to changes in ion homeostasis. NHE2-CFP or NHE3-CFP fusion proteins were functional Na(+)/H(+) exchangers when transiently expressed in NHE-deficient PS120 fibroblasts. Confocal morphometry of cells whose PM was labeled with FM4-64 measured the fractional amount of fusion protein at the cell surface. In resting cells, 10-20% of CFP fluorescence was at PM and stable over time. A protocol commonly used to activate the Na(+)/H(+) exchange function (NH(4)-prepulse acid load sustained in Na(+)-free medium), increased PM percentages of PM NHE3-CFP and NHE2-CFP. Separation of cellular acidification from Na(+) removal revealed that only NHE3-CFP translocated when medium Na(+) was removed, and only NHE2-CFP translocated when the cell was acidified. NHE2/NHE3 chimeric proteins demonstrate that the Na(+)-removal response element resides predominantly in the NHE3 cytoplasmic tail and is distinct from the acidification response sequence of NHE2.
Gens
J Scott
JS
Biocomplexity Institute, Indiana University, Bloomington, IN 47405, USA.
Du
Hongwei
H
Tackett
Lixuan
L
Kong
Shen-Shen
SS
Chu
Shaoyou
S
Montrose
Marshall H
MH
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007700:Kinetics; D009994:Osmolar Concentration; D011422:Propionates / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012964:Sodium / Q000302:isolation & purification / Q000494:pharmacology; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000737:chemistry / Q000378:metabolism*
D000906:Antibodies; C485182:Cyan Fluorescent Protein; D011422:Propionates; D000644:Quaternary Ammonium Compounds; D011993:Recombinant Fusion Proteins; C513110:Slc9a2 protein, rat; C000613387:Slc9a3 protein, rat; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D049452:Green Fluorescent Proteins; D012964:Sodium; C029658:propionic acid