Cloning, functional expression, and regulation of two K+ channels in human T lymphocytes.
Low stringency screening of a Jurkat cDNA library with a rat brain K+ channel (RCK1) probe has resulted in the isolation of HLK3, a voltage-gated K+ channel. In Xenopus oocytes, the HLK3 clone directs the expression of a rapidly activating transient outward K+ current similar to the type n K+ current recorded in Jurkat T cells. The HLK3 gene is located on the short arm of human chromosome 1 (p13.3). Polymerase chain reaction was used to clone HIsK from Jurkat cDNA. The HIsK clone shares the same sequence with a previously described genomic clone (Murai, T., Kazikuka, A., Takumi, T., Ohkubo, H., and Nakanishi, S. (1989) Biochem. Biophys. Res. Commun. 161, 176-181). In Xenopus oocytes, it encodes a slowly activating, noninactivating K+ channel which cannot be recorded in Jurkat cells by conventional patch-clamp techniques. Transcripts of both clones are present at a similar level before and after activation of purified human T lymphocytes and Jurkat cells, reflecting a constitutive expression of K+ channel messages. This finding is in good agreement with the electrophysiological results for type n K+ current density on the same cells. HLK3 current is very sensitive to the scorpion toxin charybdotoxin (IC50 = 0.8 nM). HIsK current is totally insensitive to this toxin but is blocked by the antiarrhythmic clofilium (IC50 = 80 microM). While charybdotoxin has no effect on interleukin 2 mRNA induction, clofilium potently inhibits interleukin 2 mRNA expression upon mitogen-induced T cell activation. It is concluded that the HLK3 channel is not an important component of the T cell mitogenic response. Other targets for K+ channel blockers, such as the HIsK protein, could be involved in the activation process.
Attali
B
B
Institut de Pharmacologie Moléculaire et Cellulaire, Valbonne, France.
Romey
G
G
Honoré
E
E
Schmid-Alliana
A
A
Mattéi
M G
MG
Lesage
F
F
Ricard
P
P
Barhanin
J
J
Lazdunski
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002455:Cell Division / Q000187:drug effects; D018999:Charybdotoxin; D002874:Chromosome Mapping; D002878:Chromosomes, Human, Pair 1; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D015221:Potassium Channels / Q000187:drug effects / Q000235:genetics*; D014176:Protein Biosynthesis; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics; D012604:Scorpion Venoms / Q000494:pharmacology; D016415:Sequence Alignment; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism*; D014158:Transcription, Genetic; D014981:Xenopus
D007376:Interleukin-2; D015221:Potassium Channels; D000644:Quaternary Ammonium Compounds; D012333:RNA, Messenger; D012604:Scorpion Venoms; D018999:Charybdotoxin; C022799:clofilium; D004247:DNA