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Charybdotoxin blocks voltage-gated K+ channels in human and murine T lymphocytes
S B Sands1, R S Lewis, M D Cahalan
1Department of Physiology and Biophysics, University of California, Irvine 92717.
Abstract:
A variety of scorpion venoms and purified toxins were tested for effects on ion channels in human T lymphocytes, a human T leukemia cell line (Jurkat), and murine thymocytes, using the whole-cell patch-clamp method. Nanomolar concentrations of charbdotoxin (CTX), a purified peptide component of Leiurus quinquestriatus venom known to block Ca2+-activated K+ channels from muscle, blocked "type n" voltage-gated K+ channels in human T lymphoid cells. The Na+ channels occasionally expressed in these cells were unaffected by the toxin. From the time course of development and removal of K+ channel block we determined the rates of CTX binding and unbinding. CTX blocks K+ channels in Jurkat cells with a Kd value between 0.5 and 1.5 nM. Of the three types of voltage-gated K+ channels present in murine thymocytes, types n and n' are blocked by CTX at nanomolar concentrations. The third variety of K+ channels, "type l," is unaffected by CTX. Noxiustoxin (NTX), a purified toxin from Centruroides noxius known to block Ca2+-activated K+ channels, also blocked type n K+ channels with a high degree of potency (Kd = 0.2 nM). In addition, several types of crude scorpion venoms from the genera Androctonus, Buthus, Centruroides, and Pandinus blocked type n channels. We conclude that CTX and NTX are not specific for Ca2+ activated K+ channels and that purified scorpion toxins will provide useful probes of voltage-gated K+ channels in T lymphocytes. The existence of high-affinity sites for scorpion toxin binding may help to classify structurally related K+ channels and provide a useful tool for their biochemical purification.
Insights
Scorpion toxins charibdotoxin (CTX) and noxiustoxin (NTX) effectively block specific voltage-gated potassium channels in human T lymphocytes and murine thymocytes. These toxins offer valuable tools for studying T lymphocyte ion channel function and purification.
Area of Science:
- Immunology
- Neuroscience
- Biochemistry
Background:
- Ion channels are crucial for T lymphocyte function.
- Scorpion venoms contain diverse toxins that modulate ion channel activity.
- Voltage-gated potassium channels play a key role in T cell activation and proliferation.
Purpose of the Study:
- To investigate the effects of scorpion toxins on ion channels in human T lymphocytes and murine thymocytes.
- To characterize the binding kinetics and specificity of charibdotoxin (CTX) and noxiustoxin (NTX) on T cell ion channels.
- To explore the potential of scorpion toxins as molecular probes for voltage-gated potassium channels.
Main Methods:
- Whole-cell patch-clamp electrophysiology was employed to record ion channel currents.
- Purified toxins (CTX, NTX) and crude scorpion venoms were applied to T lymphocyte and thymocyte cell lines.
- Dose-response relationships and kinetic analyses were performed to determine toxin affinity and binding rates.
Main Results:
- Charibdotoxin (CTX) potently blocked "type n" voltage-gated potassium channels in human T lymphocytes and Jurkat cells (Kd = 0.5–1.5 nM).
- CTX also blocked "type n" and "type n'" voltage-gated potassium channels in murine thymocytes, sparing "type l" channels.
- Noxiustoxin (NTX) demonstrated high affinity for "type n" potassium channels (Kd = 0.2 nM), and crude venoms also exhibited inhibitory effects on these channels.
- Sodium channels in T lymphocytes were unaffected by CTX.
Conclusions:
- Charibdotoxin (CTX) and noxiustoxin (NTX) are not exclusively specific for Ca2+-activated K+ channels.
- These purified scorpion toxins serve as effective pharmacological tools for investigating voltage-gated potassium channels in T lymphocytes.
- High-affinity scorpion toxin binding sites can aid in the classification and biochemical purification of structurally related potassium channels.