title: Inhibition of rat Na+(-)HCO3(-) cotransporter (NBCn1) function and expression by the alternative splice domain.
abstract: The Na(+)-HCO(3)(-) cotransporter NBCn1 (SLC4A7) has multiple variants depending upon splice domains in the cytoplasmic amino- and carboxy-termini of the protein. In this study, we examined the role of the amino-terminal splice domain containing 123 amino acids (cassette II) in the regulation of NBCn1 function and expression. Polymerase chain reaction detected NBCn1 mRNAs containing cassette II in a variety of tissues. Two variants, NBCn1-B containing cassette II and NBCn1-E lacking cassette II, were expressed in Xenopus oocytes and assessed by two-electrode voltage clamp to measure the ionic current mediated by the transporters. The two variants showed similar current-voltage (I-V) relations when measured 3-4 days after RNA injection. Replacment of Cl() with gluconate did not affect the I-V relations. When exposed to solutions containing 20-50 mm Na(+), the current produced by NBCn1-B was slightly more positive than that produced by NBCn1-E. The two currents were similar at 100 mm Na(+). The slope conductances for the two variants were progressively increased at higher Na(+) levels, and the increases were parallel and superimposed. Measured at different time points after RNA injection, NBCn1-B produced lower conductance than NBCn1-E at 24-48 h. Protein expression of NBCn1-B was also low at these time points as determined by immunoblot of oocyte membrane preparation. Expressed in opossum kidney (OK) cells, NBCn1-E caused a 1.5-fold increase in ouabain-sensitive production of p-nitrophenol from p-phenyl phosphate compared with control preparations, whereas NBCn1-B had negligible effect. We conclude that the primary function of cassette II is to reduce NBCn1 protein expression.
authors.affiliation: Department of Physiology, Emory University, Atlanta, GA 30322, USA.
authors.forename: Han Soo
authors.initials: HS
authors.lastname: Yang
authors.forename: Deborah S
authors.initials: DS
authors.lastname: Cooper
authors.forename: Ira
authors.initials: I
authors.lastname: Rajbhandari
authors.forename: Hae Jeong
authors.initials: HJ
authors.lastname: Park
authors.forename: Soojung
authors.initials: S
authors.lastname: Lee
authors.forename: Inyeong
authors.initials: I
authors.lastname: Choi
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D015153:Blotting, Western; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004594:Electrophysiology; D005260:Female; D015870:Gene Expression / Q000235:genetics / Q000502:physiology; D007473:Ion Channels / Q000502:physiology; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D027982:Sodium-Bicarbonate Symporters / Q000235:genetics* / Q000502:physiology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D014981:Xenopus
chemical_list: D018528:ATP-Binding Cassette Transporters; D018076:DNA, Complementary; D007473:Ion Channels; D012333:RNA, Messenger; C498902:Slc4a7 protein, rat; D027982:Sodium-Bicarbonate Symporters; D000254:Sodium-Potassium-Exchanging ATPase