Expression of the mammalian system A neutral amino acid transporter in Xenopus oocytes.
In this report, we demonstrate the expression of the mammalian System A neutral amino acid transporter in Xenopus laevis oocytes following microinjection of mRNA from rat liver, Chinese hamster ovary (CHO) cells, and human placenta. Stage 6 oocytes were injected with poly(A+) mRNA from one of these three sources and incubated for 24 h prior to assaying Na(+)-dependent 2-aminoisobutyric acid transport to monitor the increase in System A activity. The endogenous 2-aminoisobutyric acid uptake rates in oocytes were sufficiently slow so as to provide a low background value that was subtracted to obtain transport rates for the mammalian carrier alone. The degree of expression of the mammalian System A activity in Xenopus oocytes corresponded to the known transport rates in the tissue from which the mRNA was prepared. For example, hepatic mRNA from glucagon-treated rats produced greater System A activity than mRNA from control animals, and the mRNA from the CHO transport mutant cell line alar4-H3.9, which overproduces System A, resulted in higher transport rates than mRNA from the parental cell line (CHO-K1). Fractionation of total mRNA poly(A+) by nondenaturing agarose gel electrophoresis revealed transport activity associated with a 2.0-2.5-kilobase mRNA fraction common to each of the three tissues tested.
Tarnuzzer
R W
RW
Department of Biochemistry and Molecular Biology, J. Hillis Miller Health Center, University of Florida School of Medicine, Gainesville 32610.
Campa
M J
MJ
Qian
N X
NX
Englesberg
E
E
Kilberg
M S
MS
D000621:Aminoisobutyric Acids / Q000378:metabolism; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics*; D002460:Cell Line; D005260:Female; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008845:Microinjections; D009865:Oocytes / Q000378:metabolism*; D010920:Placenta / Q000378:metabolism; D011247:Pregnancy; D012333:RNA, Messenger / Q000008:administration & dosage / Q000235:genetics* / Q000302:isolation & purification; D051381:Rats; D014982:Xenopus laevis
D000621:Aminoisobutyric Acids; D002352:Carrier Proteins; D012333:RNA, Messenger; C100049:2-aminoisobutyric acid