Margatoxin binds to a homomultimer of K(V)1.3 channels in Jurkat cells. Comparison with K(V)1.3 expressed in CHO cells

• Published in: Biochemistry (Easton) 1997-03, Vol.36 (12), p.3737
• Main Authors: Helms, L M, Felix, J P, Bugianesi, R M, Garcia, M L, Stevens, S, Leonard, R J, Knaus, H G, Koch, R, Wanner, S G, Kaczorowski, G J, Slaughter, R S
• Format: Article
• Language: English
• Subjects: Animals; Blotting, Western; CHO Cells; Chromatography, High Pressure Liquid; Cricetinae; Humans; Iodine - metabolism; Jurkat Cells; Kinetics; Kv1.3 Potassium Channel; Neurotoxins - metabolism; Potassium Channels - metabolism; Potassium Channels, Voltage-Gated; Rats; Rubidium - metabolism; Scorpion Venoms - metabolism
• ISSN: 0006-2960
• DOI: 10.1021/bi962351p

Voltage-gated potassium (K(V)) channels play key roles in setting the resting potential and in the activation cascade of human peripheral T lymphocytes. Margatoxin (MgTX), a 39-amino acid peptide from Centruroides margaritatus, is a potent inhibitor of lymphocyte K(V) channels. The binding of monoiodotyrosinyl margatoxin ([125I]MgTX) to plasma membranes prepared from either Jurkat cells, a human leukemic T cell line, or CHO cells stably transfected with the Shaker-type voltage-gated K+ channel, K(V)1.3, has been used to investigate the properties of lymphocyte K(V) channels. These data were compared with [125I]MgTX binding to heterotetrameric K(V) channels in rat brain synaptic plasma membranes [Knaus, H. G., et al. (1995) Biochemistry 34, 13627-13634]. The affinity for [125I]MgTX is 100-200 fM in either Jurkat or CHO/K(V)1.3 membranes, and the receptor density is 20-120 fmol/mg in Jurkat membranes or 1000 fmol/mg in CHO/K(V)1.3 membranes. In contrast to rat brain, [125I]MgTX binding to Jurkat and CHO/K(V)1.3 membranes exhibits an absolute requirement for K+, with no potentiation of binding by Na+. K(V)1.3 was the only K(V)1 series channel present in either CHO/K(V)1.3 or Jurkat plasma membranes as determined by immunoprecipitation of [125I]MgTX binding or by Western blot analyses using sequence-specific antibodies prepared against members of the K(V)1 family. The relative potencies of a series of peptidyl K(V) channel inhibitors was essentially the same for inhibition of [125I]MgTX binding to Jurkat, CHO, or rat brain membranes and for blocking 86Rb+ efflux from the CHO/K(V)1.3 cells, except that alpha-dendrotoxin was more potent at blocking binding to rat brain membranes than in the other assays. The characteristics of [125I]MgTX binding, the antibody profiles, and the effects of the peptidyl K(V) inhibitors all indicate that the [125I]MgTX receptor in Jurkat lymphocytes is comprised of a homomultimer of K(V)1.3, unlike the heteromultimeric arrangement of the receptor in rat brain.