Cytotoxic T lymphocytes are effector CD8+ T cells that eradicate infected and malignant cells. Here we show that the transcription factor NFATc1 controls the cytotoxicity of mouse cytotoxic T lymphocytes. Activation of Nfatc1 −/− cytotoxic T lymphocytes showed a defective cytoskeleton organization and recruitment of cytosolic organelles to immunological synapses. These cells have reduced cytotoxicity against tumor cells, and mice with NFATc1-deficient T cells are defective in controlling Listeria infection. Transcriptome analysis shows diminished RNA levels of numerous genes in Nfatc1 −/− CD8+ T cells, including Tbx21, Gzmb and genes encoding cytokines and chemokines, and genes controlling glycolysis. Nfatc1 −/−, but not Nfatc2 −/− CD8+ T cells have an impaired metabolic switch to glycolysis, which can be restored by IL-2. Genome-wide ChIP-seq shows that NFATc1 binds many genes that control cytotoxic T lymphocyte activity. Together these data indicate that NFATc1 is an important regulator of cytotoxic T lymphocyte effector functions.

Immune therapy of cancer is among the most promising recent advances in medicine. Whether the immune system can keep cancer in check depends on, among other factors, the efficiency of immune cells to recognize and eliminate cancer cells. We describe a time-resolved single-cell assay that reports the quality, quantity, and kinetics of target cell death induced by single primary human natural killer (NK) cells. The assay reveals that single NK cells induce cancer cell death by apoptosis and necrosis but also by mixed forms. Inhibition of either one of the two major cytotoxic pathways, perforin/granzyme release or FasL/FasR interaction, unmasked the parallel activity of the other one. Ca influx through Orai channels is important for tuning killer cell function. We found that the apoptosis/necrosis ratio of cancer cell death by NK cells is controlled by the magnitude of Ca entry and furthermore by the relative concentrations of perforin and granzyme B. The possibility to change the apoptosis/necrosis ratio employed by NK cells offers an intriguing possibility to modulate the immunogenicity of the tumor microenvironment.

Cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are required to protect the human body against cancer. Ca is a key metabolic factor for lymphocyte function and cancer homeostasis. We analysed the Ca dependence of CTL and NK cell cytotoxicity against cancer cells and found that CTLs have a bell-shaped Ca dependence with an optimum for cancer cell elimination at rather low [Ca ] (23-625 μm) and [Ca ] (122-334 nm). This finding predicts that a partial inhibition of Orai1 should increase (rather than decrease) cytotoxicity of CTLs at [Ca ] higher than 625 μm. We tested this hypothesis in CTLs and indeed found that partial down-regulation of Orai1 by siRNA increases the efficiency of cancer cell killing. We found two mechanisms that may account for the Ca optimum of cancer cell killing: (1) migration velocity and persistence have a moderate optimum between 500 and 1000 μm [Ca ] in CTLs, and (2) lytic granule release at the immune synapse between CTLs and cancer cells is increased at 146 μm compared to 3 or 800 μm, compatible with the Ca optimum for cancer cell killing. It has been demonstrated in many cancer cell types that Orai1-dependent Ca signals enhance proliferation. We propose that a decrease of [Ca ] or partial inhibition of Orai1 activity by selective blockers in the tumour microenvironment could efficiently reduce cancer growth by simultaneously increasing CTL and NK cell cytotoxicity and decreasing cancer cell proliferation.

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