Induction of glycinebetaine uptake into Xenopus oocytes by injection of poly(A)+ RNA from renal cells exposed to high extracellular NaCl.
Madin-Darby canine kidney (MDCK) cells accumulate glycinebetaine via Na(+)-dependent transport in response to hypertonic stress. When extracellular tonicity is increased by the addition of NaCl, Vmax for glycinebetaine transport increases without an associated change in Km, consistent with an increase in the number of functioning transporters. To test whether increased transport activity results from increased gene expression, we injected poly(A)+ RNA (mRNA) from MDCK cells into Xenopus oocytes and assayed for glycinebetaine uptake in ovo. RNA-induced Na(+)-dependent uptake is observed in oocytes injected with mRNA from cells exposed to high extracellular NaCl, but not in oocytes injected with either water or mRNA from cells maintained in isotonic medium. Unfractionated mRNA induces glycinebetaine uptake in ovo at a rate which is approximately 3-fold higher than in water-injected controls. Size-fractionated mRNA (median size 2.8 kilobases) induces uptake at a rate which is approximately 7-fold higher than controls. Such RNA-induced transport activity in ovo is consistent with heterologous expression of Na(+)/glucinebetaine cotransporters encoded by renal mRNA. Increased transporter mRNA in cells exposed to hypertonicity probably underlies the pattern of expression observed in ovo. This can account for the observed rise in MDCK cell glycinebetaine transport during hypertonic stress.
Robey
R B
RB
Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
Kwon
H M
HM
Handler
J S
JS
Garcia-Perez
A
A
Burg
M B
MB
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D004285:Dogs; D004587:Electrophoresis, Agar Gel; D029968:Escherichia coli Proteins; D007668:Kidney / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008845:Microinjections; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D033902:Periplasmic Binding Proteins; D011061:Poly A / Q000494:pharmacology*; D012333:RNA, Messenger / Q000494:pharmacology*; D012965:Sodium Chloride / Q000737:chemistry*; D014981:Xenopus
D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; C053437:PROX protein, E coli; D033902:Periplasmic Binding Proteins; D012333:RNA, Messenger; D011061:Poly A; D012965:Sodium Chloride