Molecular cloning of a chloride channel that is regulated by dehydration and expressed predominantly in kidney medulla.
Complementary DNA encoding a rat kidney chloride channel (CIC-K1) was isolated by a polymerase chain reaction (PCR) cloning strategy. We designed degenerate primers, based on the regions where previously cloned chloride channels (CIC-0, -1, and -2) possess significant amino acid identity, and performed reverse transcription PCR with whole kidney mRNA. The 686-amino acid protein encoded by CIC-K1 is about 40% identical to the previously cloned chloride channels and has a similar hydropathy profile. Expression of CIC-K1 in Xenopus oocytes induced Cl- currents that activate instantaneously upon hyperpolarization and depolarization, and displayed a slightly outwardly rectifying current-voltage relationship. The message for CIC-K1 was 2.4 kilobases and was found predominantly in kidney, especially in the inner medulla. Reverse transcription PCR technique using micro-dissected nephron segments revealed that the main site of expression in kidney was the thin ascending limb of Henle's loop, which has the highest Cl- permeability among the nephron segments and is thought to be involved in a counter-current system for urine concentration in the inner medulla. The abundance of CIC-K1 mRNA in kidney increased about 4-fold as rats became dehydrated by deprivation of water for 5 days. The site of expression and the regulation by dehydration suggest that CIC-K1 function may be important in urinary concentrating mechanisms.
Uchida
S
S
Second Department of Internal Medicine, Tokyo Medical and Dental University, Japan.
Sasaki
S
S
Furukawa
T
T
Hiraoka
M
M
Imai
T
T
Hirata
Y
Y
Marumo
F
F
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018118:Chloride Channels; D002712:Chlorides; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded; D005786:Gene Expression Regulation; D007473:Ion Channels / Q000235:genetics*; D007679:Kidney Medulla / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014867:Water / Q000502:physiology; D014981:Xenopus
D018118:Chloride Channels; D002712:Chlorides; D004277:DNA, Single-Stranded; D007473:Ion Channels; D008565:Membrane Proteins; D012333:RNA, Messenger; D014867:Water