The activation of biological T cell responses requires prolonged contact with APCs and sustained signaling. We investigated whether signaling must be uninterrupted to commit T cells to cytokine production or whether T cell activation may also result from summation of interrupted signals. Upon periodic addition and removal of a src kinase inhibitor, human CD4+ T cells destroyed and re-formed immunological synapses while aborting and restarting signal transduction. Remarkably, under these conditions, T cells were eventually activated to IFN-γ production and the amount of IFN-γ produced was directly related to the total signaling time despite the repeated interruptions. Our results illustrate that T cell activation does not require a stable immunological synapse and can be achieved by interrupted signaling. It is implied that T cells can add activation signals, possibly collected on multiple APCs.
Signaling events induced by T‐cell receptor (TCR) engagement involve a cascade of tyrosine phosphorylation events leading to activation of several downstream pathways and resulting in cytokine production. TCR‐dependent interferon γ (IFN‐γ) production by Th1 cells has been shown to require tyrosine phosphorylation of numerous proteins, intracellular Ca2+ mobilization, and mitogen‐activated protein kinase activation. In contrast, the signaling pathways responsible for TCR‐dependent interleukin (IL) 4 production remain poorly understood. By using a T‐cell hybridoma that displays a hierarchized production of IL‐4 and IFN‐γ following TCR engagement (IL‐4 being produced at a lower threshold of activation than IFN‐γ), we showed that IL‐4 can be produced in spite of the absence of tyrosine phosphorylation of phospholipase Cγ1. However, protein kinase C (PKC) was found to be translocated to the cell membrane, and an increase in intracellular Ca2+ concentration was observed. The PKC‐dependent Ca2+ response and IL‐4 expression were accounted for by a dihydropyridine‐sensitive Ca2+ entry, which could occur through L‐type calcium channels. This pathway was also functional in the D10G4.1 Th2 clone. The fact that this pathway, allowing IL‐4 production, did not require optimal activation might explain why low doses of peptides or altered peptide ligands favor Th2 responses.
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