MARGATOXIN BINDS TO A HOMOMULTIMER OF K(V)1.3 CHANNELS IN JURKAT CELLS - COMPARISON WITH K(V)1.3 EXPRESSED IN CHO CELLS

Voltage-gated potassium (K-V) channels play key roles in setting the r esting potential and in the activation cascade of human peripheral T l ymphocytes. Margatoxin (MgTX), a 39-amino acid peptide from Centruroid es margaritatus, is a potent inhibitor of lymphocyte Ky channels. The binding of monoiodotyrosinyl margatoxin ([I-125]MgTX) to plasma membra nes prepared from either Jurkat cells, a human leukemic T cell line, o r CHO cells stably transfected with the Shaker-type voltage-gated K+ c hannel, K(V)1.3, has been used to investigate the properties of lympho cyte Ky channels. These data were compared with [I-125]MgTX binding to heterotetrameric Ky channels in rat brain synaptic plasma membranes [ Knaus, H. G., et al. (1995) Biochemistry 34, 13627-13634]. The affinit y for [I-125]MgTX is 100-200 fM in either Jurkat or CHO/K(V)1.3 membra nes, 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, [I-1 25]MgTX binding to Jurkat and CHO/K(V)1.3 membranes exhibits an absolu te requirement for K+, with no potentiation of binding by Nai K(V)1.3 was the only K(V)1 series channel present in either CHO/K(V)1.3 or Jur kat plasma membranes as determined by immunoprecipitation of [I-125]Mg TX binding or by Western blot analyses using sequence-specific antibod ies prepared against members of the K(V)1 family. The relative potenci es of a series of peptidyl Ky channel inhibitors was essentially the s ame for inhibition of [I-125]MgTX binding to Jurkat, CHO, or rat brain membranes and for blocking Rb-86(+) efflux from the CHO/K(V)1.3 cells , except that cl-dendrotoxin was more potent at blocking binding to ra t brain membranes than in the other assays. The characteristics of [I- 125]MgTX binding, the antibody profiles, and the effects of the peptid yl Ky inhibitors all indicate that the [I-125]MgTX receptor in Jurkat lymphocytes is comprised of a homomultimer of K(V)1.3, unlike the hete romultimeric arrangement of the receptor in rat brain.