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Environmental cues regulate epigenetic reprogramming of airway-resident memory CD8+ T cells

Nature Immunology volume 21, pages 309–320 (2020)Cite this article

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Abstract

Tissue-resident memory T cells (TRM cells) are critical for cellular immunity to respiratory pathogens and reside in both the airways and the interstitium. In the present study, we found that the airway environment drove transcriptional and epigenetic changes that specifically regulated the cytolytic functions of airway TRM cells and promoted apoptosis due to amino acid starvation and activation of the integrated stress response. Comparison of airway TRM cells and splenic effector-memory T cells transferred into the airways indicated that the environment was necessary to activate these pathways, but did not induce TRM cell lineage reprogramming. Importantly, activation of the integrated stress response was reversed in airway TRM cells placed in a nutrient-rich environment. Our data defined the genetic programs of distinct lung TRM cell populations and show that local environmental cues altered airway TRM cells to limit cytolytic function and promote cell death, which ultimately leads to fewer TRM cells in the lung.

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Fig. 1: Decline of A-TRM and I-TRM cells over time.
Fig. 2: In situ apoptosis drives lung TRM cell decline.
Fig. 3: Influenza-specific lung A-TRM and I-TRM cells have distinct transcriptional profiles.
Fig. 4: Chromatin accessibility reveals a distinct epigenetic programming of lung A-TRM and I-TRM populations.
Fig. 5: Exposure to the airway environment drives activation of the ISR.
Fig. 6: The airway environment is sufficient to alter the transcriptional program of S-TEM cells but does not induce core TRM programming.
Fig. 7: Restoration of a nutrient-rich environment resolves environmentally driven cellular stress in A-TRM cells.

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Data availability

All sequencing data are available from the National Center for Biotechnology Information Gene Expression Omnibus under accession no. GSE118112. All code, data processing scripts and additional data that support the findings of this study are available from the corresponding author upon request.

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Acknowledgements

We thank the New York University Genome Technology Center and University of Albama at Birmingham Helfin Genomics Core for Illumina sequencing, the Emory Integrated Genomics Core for sequencing library Bioanalyzer expertise, Children’s Healthcare of Atlanta and Emory University Pediatric Flow Cytometry Core for cell sorting and the NIH Tetramer Core Facility (contract no. HHSN272201300006C). This project was supported by NIH grants (nos. R01HL122559 and R01HL138508) and Centers of Excellence in Influenza Research and Surveillance contracts (no. HHSN272201400004C (to J.E.K.), and nos. 1R01AI113021 and P01AI125180-01 (to J.M.B.)). S.L.H. was supported by an NIH grant (no. F31 HL136101).

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Author notes

  1. These authors contributed equally: Sarah L. Hayward, Christopher D. Scharer.

Authors and Affiliations

  1. Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA, USA

    Sarah L. Hayward, Christopher D. Scharer, Emily K. Cartwright, Zheng-Rong Tiger Li, Jeremy M. Boss & Jacob E. Kohlmeier

  2. Department of Immunology, Kindai University Faculty of Medicine, Osaka-Sayama, Japan

    Shiki Takamura

  3. Emory-UGA Center of Excellence for Influenza Research and Surveillance, Atlanta, GA, USA

    Jacob E. Kohlmeier

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Contributions

S.L.H., C.D.S., J.M.B. and J.E.K. designed the study. S.L.H., C.D.S., E.K.C., Z.-R.T.L. and S.T. performed the experiments. S.L.H., C.D.S., E.K.C., Z.-R.T.L., S.T. and J.E.K. analyzed the data. S.L.H., C.D.S., J.M.B. and J.E.K. wrote the manuscript.

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Correspondence to Jacob E. Kohlmeier.

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Peer review information I. Visan 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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Integrated supplementary information

Supplementary Fig. 1 Sendai-specific CD8 TRM cells in the airways and interstitium gradually decline over time.

(a) Example staining reflecting Sendai NP (SenNP) tetramer staining of BAL, interstitium, and spleen on days 30 and 90 post-Sendai virus infection. (b) Number of SenNP+ CD8+ T cells in the airway (A-TRM), lung interstitium (I-TRM), and spleen (S-TEM) following infection with Sendai virus. n=10 for all timepoints. Data represented as mean ± SEM.

Supplementary Fig. 2 Lung TRM cells decline more rapidly than circulating TEM cells.

(a) Staining of CXCR3 and CD62L on FluNP+ S-TEM over time. (b) Number of FluNP+ S-TEM in wild-type mice on days 35, 60, 90, and 180 post-x31 infection. (c) Fold change in S-TEM and subsets of I-TRM FluNP+ cells defined by CD69 and CD103 expression between day 35 and day 60 (left graph) and between day 35 and day 180 (right graph). n=10 for all time points and data are derived from mice in Fig. 1. Data represented as mean ± SEM.

Supplementary Fig. 3 Validation of A-TRM-specific epigenomic and transcriptome profiles.

(a) Principal component analysis of 9,970 detected genes in S-TEM (n=3), lung vascular TEM (N=3), I-TRM (N=3) and A-TRM (n=3) FluNP+ CD8+ T cells following RNA-Seq. Points denote samples and circles show 99% confidence intervals for each cell type. (b) Bar plots of FPKM normalized gene expression for the indicated genes. Data represent mean ± SD. (c) Principal component analysis of 31,049 accessible peaks in S-TEM (n=3), lung vascular TEM (N=3), I-TRM (N=3) and A-TRM (n=3) FluNP+ CD8+ T cells following ATAC-Seq. Points denote samples and circles show 99% confidence intervals for each cell type. (d) Genome plot showing the Slc7a5, Asns, and Gzma loci. Accessibility for the indicated sample is shown along with gene structure and transcription direction. Locations of DAR are boxed. Data represent the mean of three replicates for each group. (e) Percent AnnexinV+ among I-TRM and lung vascular TEM FluNP+ CD8+ T cells. N=13 and the I-TRM data are from Fig. 2h. P value: *p<0.05. Data represented as mean ± SEM.

Supplementary Fig. 4 BCL2 is up-regulated in A-TRM compared to I-TRM.

(a) Frequency of BCL2 on FluNP+ CD8+ I-TRM cells and A-TRM cells (n=5). (b) gMFI of BCL2 on FluNP+ CD8+ I-TRM cells and A-TRM cells (n=5). Data represented as mean ± SEM. P value: ** = p<0.01. (c) Example histogram of BCL2 gated on FluNP+ CD8+ I-TRM cells and A-TRM cells.

Supplementary Fig. 5 A-TRM cells from WT and Ddit3-/- mice provide similar protection following influenza challenge.

(a) Experimental design for intratracheal (IT) transfer of WT or Ddit3-/- A-TRM cells into naïve recipient mice. (b) Viral titers measured on day 4 post-challenge in mice receiving WT (n=8) or Ddit3-/- (n=10) A-TRM cells. Data represented as mean ± SD.

Supplementary Fig. 6 Alveolar macrophages do not up-regulate stress response pathways.

Gene Set Enrichment Analysis comparing transcriptome profiles of alveolar versus interstitial macrophages for the indicated gene sets. The FDR q-value for each comparison is indicated.

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Hayward, S.L., Scharer, C.D., Cartwright, E.K. et al. Environmental cues regulate epigenetic reprogramming of airway-resident memory CD8+ T cells. Nat Immunol 21, 309–320 (2020). https://doi.org/10.1038/s41590-019-0584-x

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