Margatoxin Binds to a Homomultimer of KV1.3 Channels in Jurkat Cells. Comparison with KV1.3 Expressed in CHO Cells

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Abstract
Voltage-gated potassium (KV) 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 KV 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, KV1.3, has been used to investigate the properties of lymphocyte KV channels. These data were compared with [125I]MgTX binding to heterotetrameric KV 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/KV1.3 membranes, and the receptor density is 20−120 fmol/mg in Jurkat membranes or 1000 fmol/mg in CHO/KV1.3 membranes. In contrast to rat brain, [125I]MgTX binding to Jurkat and CHO/KV1.3 membranes exhibits an absolute requirement for K+, with no potentiation of binding by Na+. KV1.3 was the only KV1 series channel present in either CHO/KV1.3 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 KV1 family. The relative potencies of a series of peptidyl KV 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/KV1.3 cells, except that α-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 KV inhibitors all indicate that the [125I]MgTX receptor in Jurkat lymphocytes is comprised of a homomultimer of KV1.3, unlike the heteromultimeric arrangement of the receptor in rat brain.