K1.3 channel blockade with the Vm24 scorpion toxin attenuates the CD4 effector memory T cell response to TCR stimulation.

BACKGROUND

In T cells, the K1.3 and the K3.1 potassium channels regulate the membrane potential and calcium homeostasis. Notably, during T cell activation, the number of K1.3 channels on the cell membrane dramatically increases. K1.3 blockade results in inhibition of Ca signaling in T cells, thus eliciting an immunomodulatory effect. Among the naturally occurring peptides, the Vm24 toxin from the Mexican scorpion Vaejovis mexicanus is the most potent and selective K1.3 channel blocker known, which makes it a promissory candidate for its use in the clinic. We have shown that addition of Vm24 to TCR-activated human T cells inhibits CD25 expression, cell proliferation and reduces delayed-type hypersensitivity reactions in a chronic inflammation model. Here, we used the Vm24 toxin as a tool to investigate the molecular events that follow K1.3 blockade specifically on human CD4 T cells as they are actively involved in inflammation and are key mediators of autoimmune diseases.

METHODS

We combined cell viability, activation, and multiplex cytokine assays with a proteomic analysis to identify the biological processes affected by K1.3 blockade on healthy donors CD4 T cells, following TCR activation in the presence or absence of the Vm24 toxin.

RESULTS

The peptide completely blocked K1.3 channels currents without impairing T cell viability, and in response to TCR stimulation, it inhibited the expression of the activation markers CD25 and CD40L (but not that of CD69), as well as the secretion of the pro-inflammatory cytokines IFN-γ and TNF and the anti-inflammatory cytokines IL-4, IL-5, IL-9, IL-10, and IL-13. These results, in combination with data from the proteomic analysis, indicate that the biological processes most affected by the blockade of K1.3 channels in a T cell activation context were cytokine-cytokine receptor interaction, mRNA processing via spliceosome, response to unfolded proteins and intracellular vesicle transport, targeting the cell protein synthesis machinery.

CONCLUSIONS

The Vm24 toxin, a highly specific inhibitor of K1.3 channels allowed us to define downstream functions of the K1.3 channels in human CD4 T lymphocytes. Blocking K1.3 channels profoundly affects the mRNA synthesis machinery, the unfolded protein response and the intracellular vesicle transport, impairing the synthesis and secretion of cytokines in response to TCR engagement, underscoring the role of K1.3 channels in regulating T lymphocyte function.