Expression of a rabbit renal ascorbic acid transporter in Xenopus laevis oocytes
| Autor: | Hamid M. Said, David L. Dyer, M. A. Hediger, Stanley A. Rubin, Yoshikatsu Kanai |
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| Rok vydání: | 1994 |
| Předmět: |
Male
Time Factors Physiology Sodium Xenopus chemistry.chemical_element Ascorbic Acid Kidney Injections Xenopus laevis medicine Animals RNA Messenger Microinjection Messenger RNA Dose-Response Relationship Drug biology Transporter Cell Biology biology.organism_classification Ascorbic acid Molecular biology Kinetics Glucose medicine.anatomical_structure Biochemistry chemistry Oocytes RNA Rabbits Carrier Proteins Poly A Cotransporter |
| Zdroj: | American Journal of Physiology-Cell Physiology. 267:C301-C306 |
| ISSN: | 1522-1563 0363-6143 |
| Popis: | We examined the expression of renal ascorbic acid transporter(s) in Xenopus laevis oocytes after microinjection of cells with poly(A)+ RNA extracted from rabbit kidney cortex. Concomitant expression of the Na+-glucose cotransporter served as a control in these studies. Injection of poly(A)+ RNA into oocytes produced over a fivefold increase in the uptake of [14C]ascorbic acid (570 microM) compared with water-injected cells. Size fractionation of the kidney cortex mRNA by sucrose gradient revealed that the mRNA species that induced ascorbic acid transporter expression in oocytes was present in a fraction centered around 2.0 kilobases (kb) and had a size range of 1.8-3.1 kb. Injection of the active fraction into oocytes produced a > 40-fold increase in ascorbic acid uptake compared with water-injected controls. Expression of ascorbic acid transporter(s) was noticeable as early as 2 days after injection and was maximal after 7 days; it was also dependent on the amount of mRNA injected into oocytes. The induced uptake of [14C]ascorbic acid after injection of mRNA into oocytes was 1) Na+ dependent, as indicated by the almost complete lack of transport on removal of Na+ from the incubation medium; 2) significantly inhibited by unlabeled ascorbic acid and its structural analogue isoascorbic acid but not by D-glucose; and 3) saturable as a function of increasing the substrate concentration in the incubation medium (100-1,000 microM), with an apparent Km of 258 +/- 72.5 microM and a maximum velocity of 29.6 +/- 2.8 pmol.oocyte-1.2 h-1. These data demonstrate that X. laevis oocytes are a suitable system to functionally express the mammalian renal ascorbic acid transporter.(ABSTRACT TRUNCATED AT 250 WORDS) |
| Databáze: | OpenAIRE |
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