Establishment and characterization of a Chinese hamster ovary cell line, CHO-4J, stably expressing a number of Na+/I- symporters.
The cDNA of the Na+/I- symporter playing a key role in thyroid iodide transport was cloned very recently. To characterize its function, we transfected the Na+/I- symporter gene into Chinese hamster ovary (CHO) cells and established a cell line stably expressing a number of rat Na+/I- symporters, named CHO-4J. Iodide uptake was dependent on [Na+] and reached a plateau within 30 min. Kinetic studies revealed that the K(m) for iodide was 35 microM, similar to that of FRTL-5 thyroid cells. The maximal velocity (Vmax) at cell protein level was 6-10-fold higher than in FRTL-5 cells and that at single cell level was approximately 1000-fold higher. CIO4- and SCN- dose-dependently inhibited iodide uptake in a competitive manner. The Ki was 1.5 and 16 microM, respectively. Iodide efflux from CHO-4J cells was apparently slower (t1/2 = 15 min) than FRTL-5 cells (t/2 = 2 min). Electrophysiological characteristics were examined using the whole cell patch clamp technique. Rapid inward current was observed when CHO-4J cells were perfused with 50-1000 microM Nal, suggesting a transport stoichiometry of at least 2 Na+ per I-. The current-voltage relation revealed that this current was membrane potential-dependent. The reversal potential was very close to that of Na+ in agreement with dependency on the Na+ electrochemical gradient. CHO-4J cells with a slow iodide efflux, expressing a number of Na+/I-symporters whose characteristics are identical to those of FRTL-5 cells will function as a new tool for sensitive of iodide uptake.
Kosugi
S
S
Department of Laboratory Medicine, Kyoto University School of Medicine, Japan. kosugi@kuhp.kyto.u.ac.jp
Sasaki
N
N
Hai
N
N
Sugawa
H
H
Aoki
N
N
Shigemasa
C
C
Mori
T
T
Yoshida
A
A
D000818:Animals; D016466:CHO Cells; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D017136:Ion Transport; D007700:Kinetics; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D027981:Symporters; D000099209:Sodium-Iodide Symporters
D002352:Carrier Proteins; D008565:Membrane Proteins; D027981:Symporters; D000099209:Sodium-Iodide Symporters