Heterologous expression of the glutamine transporter SNAT3 in Xenopus oocytes is associated with four modes of uncoupled transport.
The glutamine transporter SNAT3 (SLC38A3, former SN1) plays a major role in glutamine release from brain astrocytes and in glutamine uptake into hepatocytes and kidney epithelial cells. Here we expressed rat SNAT3 in oocytes of Xenopus laevis and reinvestigated its transport modes using two-electrode voltage clamp and pH-sensitive microelectrodes. In addition to the established coupled Na+-glutamine-cotransport/H+ antiport, we found that there are three conductances associated with SNAT3, two dependent and one independent of the amino acid substrate. The glutamine-dependent conductance is carried by cations at pH 7.4, whereas at pH 8.4 the inward currents are still dependent on the presence of external Na+ but are carried by H+. Mutation of threonine 380 to alanine abolishes the cation conductance but leaves the proton conductance intact. Under Na+-free conditions, where the substrate-dependent conductance is suppressed, a substrate-independent, outwardly rectifying current becomes apparent at pH 8.4 that is carried by K+ and H+. In addition, we identified a glutamine-dependent uncoupled Na+/H+ exchange activity that becomes apparent upon removal of Na+ in the presence of glutamine. In conclusion, our results suggest that, in addition to coupled transport, SNAT3 mediates four modes of uncoupled ion movement across the membrane.
Schneider
Hans-Peter
HP
Abteilung für Allgemeine Zoologie, Fachbereich Biologie, TU Kaiserslautern, P B 3049, D-67653 Kaiserslautern, Germany.
Bröer
Stefan
S
Bröer
Angelika
A
Deitmer
Joachim W
JW
D000595:Amino Acid Sequence; D026921:Amino Acid Transport Systems, Neutral / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D005260:Female; D005973:Glutamine / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism* / Q000502:physiology; D021381:Protein Transport / Q000502:physiology; D011522:Protons; D051381:Rats; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis
D026921:Amino Acid Transport Systems, Neutral; D011522:Protons; D017923:Sodium-Hydrogen Exchangers; C403222:system N protein 1; D005973:Glutamine; D012964:Sodium