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Imaging T cell activation
When an antigen-presenting cell engages the T cell receptor (TCR) of a T cell, an immune synapse forms between the cells. The lipid diacylglycerol (DAG) accumulates in the T cell plasma membrane at the immune synapse. Using a fluorescent sensor, Andrada et al. imaged DAG generation and the movement of DAG-enriched organelles toward the immune synapse in T cells stimulated with antigens of different strengths (TCR binding affinities). Although all antigens stimulated an initial generation of DAG at the immune synapse, only the highest affinity antigen triggered the movement of DAG-enriched organelles. The TCR-stimulated activity of the enzyme DGKζ, which converts DAG to phosphatidic acid, was also required for full T cell polarization. Together, these data suggest that the DAG signaling gradient interprets TCR signal strength to ensure appropriate T cell activation.
Abstract
The antigen-induced formation of an immune synapse (IS) between T cells and antigen-presenting cells results in the rapid generation of the lipid second messenger diacylglycerol (DAG) in T cells. Diacylglycerol kinase ζ (DGKζ) converts DAG into phosphatidic acid (PA). Cytotoxic T lymphocytes (CTLs) from mice deficient in DGKζ have enhanced antiviral and antitumor activities, indicating that the amount of DAG controls the effectiveness of the T cell response. We characterized the second C1 domain of protein kinase Cθ (PKCθ), a DAG-binding protein that is specifically recruited to the IS, as a biological sensor to observe the generation of a DAG gradient during IS formation. In experiments with transgenic mouse CTLs expressing the OT-I T cell receptor (TCR), we showed that both strong and weak interactions between antigen and the TCR led to the rapid generation of DAG, whereas only strong interactions induced the movement of DAG-enriched organelles toward the IS. In DGKζ-deficient CTLs, antigen stimulation led to the enhanced accumulation of DAG-containing organelles at the IS; however, impaired activation of the PA effector PKCζ resulted in lack of reorientation of the microtubule-organizing center toward the IS, a process needed for effective T cell activation. Together, these data suggest that the activation of DGKζ downstream of antigen recognition provides a mechanism that ensures the activation of PA-dependent signaling as a direct result of the strength of TCR-dependent DAG mobilization.
