Differential pathways for calcium influx activated by concanavalin A and CD3 stimulation in Jurkat T cells

Abstract

Sustained increase in [Ca2+](c) (Delta[Ca2+](c)) is a critical early signal from T-cell receptor (TCR/CD3). In general, Ca2+-release activated Ca2+ channels (CRAC) are responsible for the Ca2+ influx and Delta[Ca2+](c) after TCR/CD3 stimulation. However, T cells also express Ca2+-permeable nonselective cation channels such as TRPM2 and TRPC. Gd3+ is a relatively selective blocker for CRAC at micromolar concentrations. Here, Jurkat T cells were used to investigate the Gd3+-resistant Ca2+ influx (Delta[Ca2+](c,Gd)) induced by concanavalin A (ConA, 1 mu g/ml), a widely used mitogenic agent for T cells, or by anti-CD3 Ab (alpha CD3). alpha CD3-induced Delta[Ca2+](c) was partly (similar to 60%) inhibited by 1 mu M Gd3+ while thapsigargin-induced Delta[Ca2+] was almost completely abolished. ConA-induced Delta[Ca2+] was mostly inhibited by 1 mu M Gd3+ during the early phase (< 30 s of ConA application) and became resistant during the late phase (> 2 min). Induction of Delta[Ca2+](c,Gd) by alpha CD3 and ConA was inhibited by 2-aminoethoxydiphenyl borate (2-APB) and by N-(p-amylcinnamoyl) anthranilic acid, indicating that TRPM2 and TRPC are involved in this process. Treatment with Pyr-3, a TRPC3-specific inhibitor, potently suppressed Delta[Ca2+](c,Gd) by alpha CD3 (IC50, 0.16 mu M). Patch clamp experiments demonstrated that the TRPM2 channels were activated by ConA, and the TRPC-like channels were activated by alpha CD3. Our present study suggests that TRPM2 and TRPC3 are activated by ConA and TCR/CD3, respectively, in Jurkat T cells and are responsible for the induction of Delta[Ca2+](c,Gd).