title: The GPA-dependent, spherostomatocytosis mutant AE1 E758K induces GPA-independent, endogenous cation transport in amphibian oocytes.
abstract: The previously undescribed heterozygous missense mutation E758K was discovered in the human AE1/SLC4A1/band 3 gene in two unrelated patients with well-compensated hereditary spherostomatocytic anemia (HSt). Oocyte surface expression of AE1 E758K, in contrast to that of wild-type AE1, required coexpressed glycophorin A (GPA). The mutant polypeptide exhibited, in parallel, strong GPA dependence of DIDS-sensitive (36)Cl(-) influx, trans-anion-dependent (36)Cl(-) efflux, and Cl(-)/HCO(3)(-) exchange activities at near wild-type levels. AE1 E758K expression was also associated with GPA-dependent increases of DIDS-sensitive pH-independent SO(4)(2-) uptake and oxalate uptake with altered pH dependence. In marked contrast, the bumetanide- and ouabain-insensitive (86)Rb(+) influx associated with AE1 E758K expression was largely GPA-independent in Xenopus oocytes and completely GPA-independent in Ambystoma oocytes. AE1 E758K-associated currents in Xenopus oocytes also exhibited little or no GPA dependence. (86)Rb(+) influx was higher but inward cation current was lower in oocytes expressing AE1 E758K than previously reported in oocytes expressing the AE1 HSt mutants S731P and H734R. The pharmacological inhibition profile of AE1 E758K-associated (36)Cl(-) influx differed from that of AE1 E758K-associated (86)Rb(+) influx, as well as from that of wild-type AE1-mediated Cl(-) transport. Thus AE1 E758K-expressing oocytes displayed GPA-dependent surface polypeptide expression and anion transport, accompanied by substantially GPA-independent, pharmacologically distinct Rb(+) flux and by small, GPA-independent currents. The data strongly suggest that most of the increased cation transport associated with the novel HSt mutant AE1 E758K reflects activation of endogenous oocyte cation permeability pathways, rather than cation translocation through the mutant polypeptide.
authors.affiliation: Division of Nephrology, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02215, USA.
authors.forename: Andrew K
authors.initials: AK
authors.lastname: Stewart
authors.forename: David H
authors.initials: DH
authors.lastname: Vandorpe
authors.forename: John F
authors.initials: JF
authors.lastname: Heneghan
authors.forename: Fouad
authors.initials: F
authors.lastname: Chebib
authors.forename: Kathleen
authors.initials: K
authors.lastname: Stolpe
authors.forename: Arash
authors.initials: A
authors.lastname: Akhavein
authors.forename: E Jennifer
authors.initials: EJ
authors.lastname: Edelman
authors.forename: Yelena
authors.initials: Y
authors.lastname: Maksimova
authors.forename: Patrick G
authors.initials: PG
authors.lastname: Gallagher
authors.forename: Seth L
authors.initials: SL
authors.lastname: Alper
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000558:Ambystoma mexicanum / Q000378:metabolism; D000595:Amino Acid Sequence; D000663:Amphibians / Q000235:genetics / Q000378:metabolism*; D000745:Anemia, Hemolytic, Congenital / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D001639:Bicarbonates / Q000378:metabolism*; D002034:Bumetanide / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability; D002712:Chlorides / Q000378:metabolism*; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D005260:Female; D006021:Glycophorins / Q000235:genetics / Q000378:metabolism*; D006579:Heterozygote; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials; D008875:Middle Aged; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D009865:Oocytes / Q000378:metabolism*; D010042:Ouabain / Q000494:pharmacology; D019815:Oxalic Acid / Q000378:metabolism; D012414:Rubidium Radioisotopes / Q000378:metabolism; D012720:Severity of Illness Index; D049994:Sodium Potassium Chloride Symporter Inhibitors / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism; D013431:Sulfates / Q000378:metabolism; D014982:Xenopus laevis / Q000378:metabolism
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D001639:Bicarbonates; D002712:Chlorides; C409972:GYPA protein, human; D006021:Glycophorins; D012414:Rubidium Radioisotopes; C454201:SLC4A1 protein, human; D049994:Sodium Potassium Chloride Symporter Inhibitors; D013431:Sulfates; D002034:Bumetanide; D010042:Ouabain; D019815:Oxalic Acid; D000254:Sodium-Potassium-Exchanging ATPase; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid