Antiviral CD8+ T cell responses are characterized by an initial activation/priming of T lymphocytes followed by a massive proliferation, subset differentiation, population contraction and the development of a stable memory pool. The transcription factor BATF3 has been shown to play a central role in the development of conventional dendritic cells, which in turn are critical for optimal priming of CD8+ T cells. Here we show that BATF3 was expressed transiently within the first days after T cell priming and had long-lasting T cell–intrinsic effects. T cells that lacked Batf3 showed normal expansion and differentiation, yet succumbed to an aggravated contraction and had a diminished memory response. Vice versa, BATF3 overexpression in CD8+ T cells promoted their survival and transition to memory. Mechanistically, BATF3 regulated T cell apoptosis and longevity via the proapoptotic factor BIM. By programing CD8+ T cell survival and memory, BATF3 is a promising molecule to optimize adoptive T cell therapy in patients.
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We thank the Core Unit for FACS and the Core Unit SysMed of the IZKF Würzburg for supporting this study, H.C. Probst (Institute for Immunology, Mainz University) and T. Kaisho (Immunology Frontier Research Center, Osaka University) for providing the P14 and XCR1-DTR-Venus mice31, respectively. The B8R tetramer was obtained through the National Institutes of Health Tetramer Core Facility. This work was supported by grants through the German Research Foundation, SFB TR 221 (project A03 to M. Hudecek and H.E.), Germany’s Excellence Strategy – EXC2151 – (390873048 to M. Hölzel) and German Research Foundation Emmy Noether programme (GA2129/2-1 to G.G.) and by grants of the European Research Council to W.K. (819329- STEP2) and G.G. (759176-TissueLymphoContexts). W.K., G.G. and M.V. are supported by the by the Max Planck Society (Max Planck Research Groups).
M.A.A., M. Hudecek and W.K. are currently applying for patents relating to the content of this manuscript.
Peer review information Peer reviewer reports are available. L. A. Dempsey was the primary editor on this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.
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Relative abundance of leukocytes after prime (d8; VV) and recall response (d6; VV) of bone marrow chimeric mice. Data are representative of two (n = 9 at prime and n = 7 at recall) independent experiments. Error bars indicate the mean ± SEM. Comparison between groups was calculated using two-tailed unpaired Student’s t-test.
Kinetic analysis of co-transferred WT (GFP) and Batf3−/− (CD45.1) OT-I cells in the blood of WT recipients after L.m.-OVA i.p. Data are representative of three independent experiments (n = 4). Error bars indicate the mean ± SEM. Comparison between groups was calculated using the One sample t-test. ***, p = 0.0003.
Extended Data Fig. 3 Measurement of mitochondrial potential during the contraction phase of CD8+ T cell immune response.
Representative dotplots, histograms and statistical analysis of mitochondrial membrane potential of the co-transferred WT (GFP) and Batf3−/− (CD45.1) OT-I cells. Cells were analyzed at d8 and d12 post- VV-OVA infection. Data are representative of two independent experiments (n = 4). Error bars indicate the mean ± SEM. Comparison between groups was calculated using the paired Student’s t-test. * = p ≤ 0.05.
(a) Kinetic analysis of co-transferred WT (CD45.1 / CD45.2) and Batf3−/− (CD45.1) OT-I cells in the blood of IL15 deficient (IL15-/-) recipients after or VV-OVA i.p. infection. (b) qRT-PCR of Batf in ex vivo isolated WT and Batf3−/− CD8 OT-I cells after VV-OVA infection, or 48 h hours post anti-CD3/CD28 in vitro stimulation. (c) Heatmap showing the 166 differentially expressed genes by comparing WT versus Batf3−/− CD8 T cells in vitro stimulated with anti-CD3/CD28 for 48 h or 72 h. Columns represent technical replicates. (d) Sequence and schematic of the retroviral plasmid carrying the Bcl2l11 shRNA. pLMPd-shRNA-Ametrine were provided by Transomic®. The shRNA position is marked in red and reported by Ametrine (yellow). The cassette carrying the shRNA is flanked by LTRs (dark grey) and MESV Psi (light grey). Ametrine (yellow) is active by a PGK promoter (light grey). LTR: long terminal repeat; MESV: murine embryonic stem cell virus; Psi: RNA target site for packaging; PGK: PGK promoter. Data are representative of one experiment (n = 4). Error bars indicate the mean ± SEM. Comparison between groups (n = 4) was calculated using the One sample t-test. **, p = 0.007.
For both retroviral over-expression plasmids pMIG was used as a backbone. In both cases the inserts, Batf3-IRES-GFP (dark blue and green, respectively) and Batf-IRES-GFP (light blue and green, respectively), were inserted between the LTRs (dark grey) and downstream of MESV Psi (light grey) and gag (grey-blue). As mock control pMIGR1-Ametrine was used. LTR: long terminal repeat; gag: structural precursor protein for packaging; MESV: murine embryonic stem cell virus; Psi: RNA target site for packaging.
(a, b) Naïve CD8 T cells were isolated, activated and retrovirally transduced with pMIG- Ametrine (empty vector) or carrying the murine Batf3, pMIG-BATF3-GFP (BATF3) or the murine Batf, pMIG-BATF-GFP (BATF). Cells were resting for 48 h in culture after transduction and for additional 48 h after sorting before in vitro activation with anti-CD3/CD28. (a) Representative dotplots and statistical analysis of the in vitro kinetics of WT CD8 T cells coculture (50:50 – empty vector: BATF3) and (b) 50:50 – empty vector: BATF) in two different concentrations of IL-15. (c) Representative dotplots and statistical analysis of the in vitro kinetics of WT versus Batf3-/- CD8 T cells coculture (50:50 – empty vector: empty vector or empty vector: BATF3). Data are representative of one (a; n = 4) or two (b, c; n = 4) independent experiments. Error bars indicate the mean ± SEM. Comparison between groups was calculated using the One sample t-test. **, p = 0.007.
(a-d) Representative dotplots and gating strategy for (a) IL-7R, (b) ICOS, (c) intracellular CTLA-4 and (d) Tim-3 expression analysis in resting and after anti-CD3/CD28 stimulation of 50:50 (empty vector: BATF3) CD8+ T cell coculture. (e) Statistical analysis of Glycolitic Proton Eflux Rate (GlycoPER) readouts and (f) Oxygen Consuption Rate (OCR) parameters measured in resting state and 24 h after anti-CD3/CD28 stimulation. (a-f) Data are representative of two independent experiments (n = 3). Error bars indicate the mean ± SEM. Comparison between groups was calculated using two-tailed unpaired Student’s t-test.
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Ataide, M.A., Komander, K., Knöpper, K. et al. BATF3 programs CD8+ T cell memory. Nat Immunol 21, 1397–1407 (2020). https://doi.org/10.1038/s41590-020-0786-2
Nature Immunology (2020)