Cutting Edge: Activation of STING in T Cells Induces Type I IFN Responses and Cell Death

Abstract

Stimulator of interferon genes (STING) was initially described as a sensor of intracellular bacterial and viral DNA and a promising adjuvant target in innate immune cells; more recently STING has also been shown to detect endogenous DNA and play a role in tumor immunity and autoimmune disease development. Thus far STING has been studied in macrophages and dendritic cells. In this study, to our knowledge we provide the first evidence of STING activation in T cells, in which STING agonists not only provoke type I IFN production and IFN-stimulated gene expression, mirroring the response of innate cells, but are also capable of activating cell stress and death pathways. Our results suggest a re-evaluation of STING agonist-based therapies may be necessary to identify the possible effects on the T cell compartment. Conversely, the effects of STING on T cells could potentially be harnessed for therapeutic applications.

Fig 1. T cells exhibit a functional IFN-I response to STING agonists

(A) STING protein levels in unstimulated T cells and macrophages; ISG mRNA expression in T cells (B) after 4 hours treatment with DMXAA or (D) 5 hours after electroporation with R'S'cGAMP; (C,E,F) DMXAA-induced IFNβ and IFNγ production, with and without anti-CD3 and -CD28, in total CD3⁺ or isolated CD4⁺ and CD8⁺ T cells; (G) TBK1-IRF3, NF_KB, and MAPK phosphorylation in DMXAA-activated T cells. Results are representative of 3-4 independent experiments.

Fig 2. DMXAA inhibits TCR-induced proliferation independent of IFN-I

(A) Anti-CD3 and -CD28 induced proliferation of B6, KO, and IFNAR^-/- CD3⁺ T cells in the presence of 10μg/ml DMXAA; (B) Effect of adding DMXAA concurrent with TCR stimulation (day 0), or after 24 (day 1) or 48 (day 2) hours. Results shown are representative of 3 independent experiments.

Fig 3. STING activation increases B6 T cell expression of apoptosis, cell stress related genes in addition to dramatic increases in ISG expression

(A) Gene pathways activated in response to TCR or TCR+DMXAA stimulation as determined using RNA sequencing data clustered based on similarity of samples using GenomePattern software (Broad Institute); (B) Changes in T cell expression of anti- and pro-apoptotic and ER stress-related genes in T cells in response to TCR or TCR+DMXAA activation; (C) STING-dependent upregulation of IFNs and selected ISGs in T cells; (D) STING-dependent activation of pro-apoptotic pathways in T cells versus macrophages. TCR: anti-CD3 and-CD28. RNA-sequencing experiment was done once.

Fig 4. DMXAA-induces a STING-dependent ER stress response and cell death

zVAD and Nec1 together rescue DMXAA (10μg/ml)-induced cell death after 24 hours (A), but not proliferation at 72 hours (B), in anti-CD3 and -CD28 activated T cells. (C) Cytotoxicity in CD3⁺ T cells of DMXAA alone after 12 and 24 hours is dose-dependent. (D) The DMXAA-induced, STING-dependent increase in IFIT2 mirrors (E) the increased ratio of XBP-1s:XBP-1u and (F) decreased expression of anti-apoptotic BCL2 following DMXAA treatment. Results are representative of 3 independent experiments. (G) In vivo DMXAA treatment has no effect on pLN CD4⁺ and CD8⁺ T cell populations but (H) increases CD3⁺ T cell ISG expression independent of non-T cell IFN sources. Symbols represent individual mice.