Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

mRNA vaccines engage unconventional pathways in CD8+ T cell priming

Nature (2026) Cite this article

Subjects

Abstract

Vaccines composed of mRNA and lipid nanoparticles (LNPs) activate B cells and T cells by inducing in vivo production of specific protein antigens. While B cells can be activated directly by antigens, T cell activation requires antigen processing and presentation by MHC molecules on specialized antigen-presenting cells (APCs). In response to viral infections, tumours, and protein- and cDNA-based vaccines, antigen presentation to CD8+ T cells is particularly dependent on type 1 conventional dendritic (cDC1) cells, which are specialized for efficient cross-presentation of exogenous antigens1,2,3,4. However, whether similar mechanisms have a role in mRNA–LNP vaccination is unclear. Here we report that mRNA–LNP vaccines do not require cDC1 cells or the WDFY4-dependent cross-presentation pathway for CD8+ T cell priming but instead engage both cDC1 and cDC2 cells redundantly. While CD8+ T cells primed exclusively by either cDC1 or cDC2 cells showed phenotypic differences, both could mediate anti-tumour responses and memory formation. Importantly, acquisition by cDCs of peptide–MHC-I complexes from non-haematopoietic cells, called cross-dressing, provides a substantial component of CD8+ T cell priming, in a manner dependent on type I interferon. mRNA–LNP induction of cross-dressing might explain their ability to activate CD8+ T cells against antigens not encoded by the vaccine.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to the full article PDF.

USD 39.95

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Both cDC subsets, cDC1 and cDC2 cells, prime CD8+ T cells after mRNA vaccination.
The alternative text for this image may have been generated using AI.
Fig. 2: MHC-I on APCs is not required for CD8+ T cell priming after mRNA vaccination.
The alternative text for this image may have been generated using AI.
Fig. 3: cDCs acquire MHC-I from non-haematopoietic cells and prime CD8+ T cells.
The alternative text for this image may have been generated using AI.
Fig. 4: CD8+ T cells primed by cDC2 cells are capable of an anti-tumour response.
The alternative text for this image may have been generated using AI.
Fig. 5: Distinct profiles of CD8+ T cells primed by cDC1 or cDC2 cells.
The alternative text for this image may have been generated using AI.

Similar content being viewed by others

Data availability

The scRNA-seq and TCR-seq data underlying Fig. 5 and Extended Data Fig. 7 is openly available in the National Center for Biotechnology Information Gene Expression Omnibus (GEO) under accession number GSE296093. All data in this study are available in the published Article and its Supplementary Information.

References

  1. den Haan, J. M. M., Lehar, S. M. & Bevan, M. J. CD8+ but not CD8− dendritic cells cross-prime cytotoxic T cells in vivo. J. Exp. Med. 192, 1685–1696 (2000).

    Article  Google Scholar 

  2. Hildner, K. et al. Batf3 deficiency reveals a critical role for CD8α+ dendritic cells in cytotoxic T cell immunity. Science 322, 1097–1100 (2008).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  3. Li, L. et al. Cross-dressed CD8α+/CD103+ dendritic cells prime CD8+ T cells following vaccination. Proc. Natl Acad. Sci. USA 109, 12716–12721 (2012).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  4. Kim, E. H. et al. Squalene emulsion-based vaccine adjuvants stimulate CD8 T cell, but not antibody responses, through a RIPK3-dependent pathway. eLife 9, e52687 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Baden, L. R. et al. Efficacy and safety of the mRNA-1273 SARS-CoV-2 vaccine. N. Engl. J. Med. 384, 403–416 (2021).

    Article  CAS  PubMed  Google Scholar 

  6. Dickerman, B. A. et al. Comparative effectiveness of BNT162b2 and mRNA-1273 vaccines in U.s. veterans. N. Engl. J. Med. 386, 105–115 (2022).

    Article  CAS  PubMed  Google Scholar 

  7. Rubin, E. J. & Longo, D. L. Covid-19 mRNA vaccines—six of one, half a dozen of the other. N. Engl. J. Med. 386, 183–185 (2022).

    Article  CAS  PubMed  Google Scholar 

  8. Dimitriadis, G. J. Translation of rabbit globin mRNA introduced by liposomes into mouse lymphocytes. Nature 274, 923–924 (1978).

    Article  CAS  PubMed  ADS  Google Scholar 

  9. Wolff, J. A. et al. Direct gene transfer into mouse muscle in vivo. Science 247, 1465–1468 (1990).

    Article  CAS  PubMed  ADS  Google Scholar 

  10. Martinon, F. et al. Induction of virus-specific cytotoxic T lymphocytes in vivo by liposome-entrapped mRNA. Eur. J. Immunol. 23, 1719–1722 (1993).

    Article  CAS  PubMed  Google Scholar 

  11. Pardi, N. et al. Nucleoside-modified mRNA vaccines induce potent T follicular helper and germinal center B cell responses. J. Exp. Med. 215, 1571–1588 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Pardi, N. et al. Nucleoside-modified mRNA immunization elicits influenza virus hemagglutinin stalk-specific antibodies. Nat. Commun. 9, 3361 (2018).

    Article  PubMed  PubMed Central  ADS  Google Scholar 

  13. Corbett, K. S. et al. SARS-CoV-2 mRNA vaccine design enabled by prototype pathogen preparedness. Nature 586, 567–571 (2020).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  14. Oberhardt, V. et al. Rapid and stable mobilization of CD8+ T cells by SARS-CoV-2 mRNA vaccine. Nature 597, 268–273 (2021).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  15. Rojas, L. A. et al. Personalized RNA neoantigen vaccines stimulate T cells in pancreatic cancer. Nature 618, 144–150 (2023).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  16. Kranz, L. M. et al. Systemic RNA delivery to dendritic cells exploits antiviral defence for cancer immunotherapy. Nature 534, 396–401 (2016).

    Article  PubMed  ADS  Google Scholar 

  17. Ohara, R. A. & Murphy, K. M. The evolving biology of cross-presentation. Semin. Immunol. 66, 101711 (2023).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Jo, S. et al. Shared pathway of WDFY4-dependent cross-presentation of immune complexes by cDC1 and cDC2. J. Exp. Med. 222, e20240955 (2025).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Liu, K. & Nussenzweig, M. C. Origin and development of dendritic cells. Immunol. Rev. 234, 45–54 (2010).

    Article  CAS  PubMed  Google Scholar 

  20. Murphy, T. L. et al. Transcriptional control of dendritic cell development. Annu. Rev. Immunol. 34, 93–119 (2016).

    Article  CAS  PubMed  Google Scholar 

  21. Toubai, T. et al. Host-derived CD8+ dendritic cells are required for induction of optimal graft-versus-tumor responses after experimental allogeneic bone marrow transplantation. Blood 121, 4231–4241 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Sultan, H. et al. Neoantigen-specific cytotoxic Tr1 CD4 T cells suppress cancer immunotherapy. Nature 632, 182–191 (2024).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  23. Pot, C., Apetoh, L. & Kuchroo, V. K. Type 1 regulatory T cells (Tr1) in autoimmunity. Semin. Immunol. 23, 202–208 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Bahl, K. et al. Preclinical and clinical demonstration of immunogenicity by mRNA vaccines against H10N8 and H7N9 influenza viruses. Mol. Ther. 25, 1316–1327 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Suzuki, Y. et al. Design and lyophilization of lipid nanoparticles for mRNA vaccine and its robust immune response in mice and nonhuman primates. Mol. Ther. Nucleic Acids 30, 226–240 (2022).

    Article  CAS  PubMed  Google Scholar 

  26. Hassett, K. J. et al. mRNA vaccine trafficking and resulting protein expression after intramuscular administration. Mol. Ther. Nucleic Acids 35, 102083 (2024).

    Article  CAS  PubMed  Google Scholar 

  27. Buckley, M. et al. Visualizing lipid nanoparticle trafficking for mRNA vaccine delivery in non-human primates. Mol. Ther. 33, 1105–1117 (2025).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Durai, V. et al. Cryptic activation of an Irf8 enhancer governs cDC1 fate specification. Nat. Immunol. 20, 1161–1173 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Liu, T.-T. et al. Ablation of cDC2 development by triple mutations within the Zeb2 enhancer. Nature 607, 142–148 (2022).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  30. Dolan, B. P., Gibbs, K. D. Jr & Ostrand-Rosenberg, S. Dendritic cells cross-dressed with peptide MHC class I complexes prime CD8+ T cells. J. Immunol. 177, 6018–6024 (2006).

    Article  CAS  PubMed  Google Scholar 

  31. Wakim, L. M. & Bevan, M. J. Cross-dressed dendritic cells drive memory CD8+ T-cell activation after viral infection. Nature 471, 629–632 (2011).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  32. Harding, C. V. & Unanue, E. R. Quantitation of antigen-presenting cell MHC class II/peptide complexes necessary for T-cell stimulation. Nature 346, 574–576 (1990).

    Article  CAS  PubMed  ADS  Google Scholar 

  33. Christinck, E. R., Luscher, M. A., Barber, B. H. & Williams, D. B. Peptide binding to class I MHC on living cells and quantitation of complexes required for CTL lysis. Nature 352, 67–70 (1991).

    Article  CAS  PubMed  ADS  Google Scholar 

  34. Sykulev, Y., Joo, M., Vturina, I., Tsomides, T. J. & Eisen, H. N. Evidence that a single peptide-MHC complex on a target cell can elicit a cytolytic T cell response. Immunity 4, 565–571 (1996).

    Article  CAS  PubMed  Google Scholar 

  35. Sprent, J., Miller, J. F. & Mitchell, G. F. Antigen-induced selective recruitment of circulating lymphocytes. Cell. Immunol. 2, 171–181 (1971).

    Article  CAS  PubMed  Google Scholar 

  36. Matloubian, M. et al. Lymphocyte egress from thymus and peripheral lymphoid organs is dependent on S1P receptor 1. Nature 427, 355–360 (2004).

    Article  CAS  PubMed  ADS  Google Scholar 

  37. Brinkman, C. C., Rouhani, S. J., Srinivasan, N. & Engelhard, V. H. Peripheral tissue homing receptors enable T cell entry into lymph nodes and affect the anatomical distribution of memory cells. J. Immunol. 191, 2412–2425 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Bénéchet, A. P. et al. Dynamics and genomic landscape of CD8+ T cells undergoing hepatic priming. Nature 574, 200–205 (2019).

    Article  PubMed  PubMed Central  ADS  Google Scholar 

  39. Kim, M. et al. Exogenous RNA surveillance by proton-sensing TRIM25. Science 388, eads4539 (2025).

    Article  CAS  PubMed  Google Scholar 

  40. Theisen, D. J. et al. WDFY4 is required for cross-presentation in response to viral and tumor antigens. Science 362, 694–699 (2018).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  41. Chatterjee, F. & Spranger, S. MHC-dressing on dendritic cells: boosting anti-tumor immunity via unconventional tumor antigen presentation. Semin. Immunol. 66, 101710 (2023).

    Article  CAS  PubMed  Google Scholar 

  42. Bernabeu, C., van de Rijn, M., Lerch, P. G. & Terhorst, C. P. β2-microglobulin from serum associates with MHC class I antigens on the surface of cultured cells. Nature 308, 642–645 (1984).

    Article  CAS  PubMed  ADS  Google Scholar 

  43. Duong, E. et al. Type I interferon activates MHC class I-dressed CD11b+ conventional dendritic cells to promote protective anti-tumor CD8+ T cell immunity. Immunity 55, 308–323 (2022).

    Article  CAS  PubMed  Google Scholar 

  44. Kobiyama, K. & Ishii, K. J. Making innate sense of mRNA vaccine adjuvanticity. Nat. Immunol. 23, 474–476 (2022).

    Article  CAS  PubMed  Google Scholar 

  45. Zinkernagel, R. M. & Althage, A. On the role of thymic epithelium vs. bone marrow-derived cells in repertoire selection of T cells. Proc. Natl Acad. Sci. USA 96, 8092–8097 (1999).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  46. Sakoda, Y. et al. Donor-derived thymic-dependent T cells cause chronic graft-versus-host disease. Blood 109, 1756–1764 (2007).

    Article  CAS  PubMed  Google Scholar 

  47. Ferris, S. T. et al. cDC1 prime and are licensed by CD4+ T cells to induce anti-tumour immunity. Nature 584, 624–629 (2020).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  48. Grippin, A. J. et al. SARS-CoV-2 mRNA vaccines sensitize tumours to immune checkpoint blockade. Nature 647, 488–497 (2025).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  49. Krawczyk, P. S. et al. Re-adenylation by TENT5A enhances efficacy of SARS-CoV-2 mRNA vaccines. Nature 641, 984–992 (2025).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Lobb, T. A. et al. Type I interferon restricts mRNA vaccine efficacy through suppression of antigen uptake in cDCs. npj Vaccines 11, 41 (2026).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Li, C. et al. Intravenous injection of Coronavirus disease 2019 (COVID-19) mRNA vaccine can induce acute myopericarditis in mouse model. Clin. Infect. Dis. 74, 1933–1950 (2022).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  52. Mandl, J. N. et al. Quantification of lymph node transit times reveals differences in antigen surveillance strategies of naïve CD4+ and CD8+ T cells. Proc. Natl Acad. Sci. USA 109, 18036–18041 (2012).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  53. Mashayekhi, M. et al. CD8α+ dendritic cells are the critical source of interleukin-12 that controls acute infection by Toxoplasma gondii tachyzoites. Immunity 35, 249–259 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. MacNabb, B. W. et al. Dendritic cells can prime anti-tumor CD8+ T cell responses through major histocompatibility complex cross-dressing. Immunity 55, 982–997 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Nakayama, M. Antigen presentation by MHC-dressed cells. Front. Immunol. 5, 672 (2014).

    PubMed  Google Scholar 

  56. Lybarger, L., Wang, X., Harris, M. R., Virgin, H. W. IV & Hansen, T. H. Virus subversion of the MHC class I peptide-loading complex. Immunity 18, 121–130 (2003).

    Article  CAS  PubMed  Google Scholar 

  57. Theisen, D. J. et al. Batf3-dependent genes control tumor rejection induced by dendritic cells independently of cross-presentation. Cancer Immunol. Res. 7, 29–39 (2019).

    Article  CAS  PubMed  Google Scholar 

  58. Lee, D. D. et al. Correction: ADAPT-3D: accelerated deep adaptable processing of tissue for 3-dimensional fluorescence tissue imaging for research and clinical settings. Sci. Rep. 15, 32625 (2025).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  59. Matsushita, H. et al. Cancer exome analysis reveals a T-cell-dependent mechanism of cancer immunoediting. Nature 482, 400–404 (2012).

    Article  CAS  PubMed  PubMed Central  ADS  Google Scholar 

  60. Hao, Y. et al. Dictionary learning for integrative, multimodal and scalable single-cell analysis. Nat. Biotechnol. 42, 293–304 (2024).

    Article  CAS  PubMed  ADS  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from the US National Institute of Health (NIH) to K.M.M. (R01AI150297, R01CA248919, R01AI162643 and R21AI163421) and to W.E.G. (R01CA240983); and grants from the Washington University’s Pancreatic Cancer SPORE to W.E.G. (P50CA196510-05). Additional support to W.E.G. was provided through a sponsored research agreement with Innovac Therapeutics. We thank the staff at the NIH Tetramer Core Facility (contract number 75N93020D00005) for providing APC- and PE-conjugated H-2Kb chicken ova 257–264 SIINFEKL tetramers; the staff at GTAC@MGI at WashU Medicine for sequencing services. The centre is partially supported by NCI Cancer Center Support Grant P30 CA91842 to the Siteman Cancer Center from the National Center for Research Resources (NCRR), a component of the NIH, and NIH Roadmap for Medical Research. Graphics were created in BioRender; Jo, S. https://BioRender.com/62s9j87 (2026). This publication is solely the responsibility of the authors and does not necessarily represent the official view of NCRR or NIH.

Author information

Authors and Affiliations

  1. Department of Pathology and Immunology, Washington University in St Louis School of Medicine, St Louis, MO, USA

    Suin Jo, Kevin A. Telfer, Hussein Sultan, Ray A. Ohara, Michelle He, Giri Nam, Jing Chen, Feiya Ou, Robert D. Schreiber, Gwendalyn J. Randolph, Naresha Saligrama, Theresa L. Murphy & Kenneth M. Murphy

  2. Department of Surgery, Washington University in St Louis School of Medicine, St Louis, MO, USA

    Lijin Li & William E. Gillanders

  3. Department of Neurology, Washington University School of Medicine, St Louis, MO, USA

    Chandrani Thakur & Naresha Saligrama

  4. The Andrew M. and Jane M. Bursky Center for Human Immunology and Immunotherapy Programs, Washington University in St Louis School of Medicine, St Louis, MO, USA

    Hussein Sultan, Robert D. Schreiber, Naresha Saligrama & William E. Gillanders

  5. Innovac Therapeutics, Cambridge, MA, USA

    Monia Draghi & Nicholas M. Valiante

  6. The Alvin J. Siteman Cancer Center at Barnes–Jewish Hospital and Washington University School of Medicine, St Louis, MO, USA

    Robert D. Schreiber & William E. Gillanders

Authors
  1. Suin Jo
    View author publications

    Search author on:PubMed Google Scholar

  2. Lijin Li
    View author publications

    Search author on:PubMed Google Scholar

  3. Chandrani Thakur
    View author publications

    Search author on:PubMed Google Scholar

  4. Kevin A. Telfer
    View author publications

    Search author on:PubMed Google Scholar

  5. Hussein Sultan
    View author publications

    Search author on:PubMed Google Scholar

  6. Ray A. Ohara
    View author publications

    Search author on:PubMed Google Scholar

  7. Michelle He
    View author publications

    Search author on:PubMed Google Scholar

  8. Giri Nam
    View author publications

    Search author on:PubMed Google Scholar

  9. Jing Chen
    View author publications

    Search author on:PubMed Google Scholar

  10. Feiya Ou
    View author publications

    Search author on:PubMed Google Scholar

  11. Monia Draghi
    View author publications

    Search author on:PubMed Google Scholar

  12. Nicholas M. Valiante
    View author publications

    Search author on:PubMed Google Scholar

  13. Robert D. Schreiber
    View author publications

    Search author on:PubMed Google Scholar

  14. Gwendalyn J. Randolph
    View author publications

    Search author on:PubMed Google Scholar

  15. Naresha Saligrama
    View author publications

    Search author on:PubMed Google Scholar

  16. Theresa L. Murphy
    View author publications

    Search author on:PubMed Google Scholar

  17. William E. Gillanders
    View author publications

    Search author on:PubMed Google Scholar

  18. Kenneth M. Murphy
    View author publications

    Search author on:PubMed Google Scholar

Contributions

S.J., L.L., W.E.G., T.L.M. and K.M.M. designed the study. S.J., L.L., K.A.T., H.S., R.A.O., M.H., G.N., J.C. and F.O. conducted experiments and analysed the data. S.J., R.D.S., G.J.R., N.S., T.L.M. and W.E.G. generated and maintained mice and supervised experiments. M.D. and N.M.V. manufactured and tested the mRNA–LNPs. S.J. executed the scRNA-seq experiments, and S.J., C.T. and N.S. conducted their analysis. S.J., T.L.M. and K.M.M. wrote the manuscript with input from all of the authors.

Corresponding authors

Correspondence to William E. Gillanders or Kenneth M. Murphy.

Ethics declarations

Competing interests

N.M.V. holds stock in Glyde Bio and Innovac Therapeutics. G.J.R. reports a patent pending (PCT/US2025/038758) and an associated product licensed to Leinco Technologies. R.D.S. is a co-founder, scientific advisory board member and stockholder of Asher Biotherapeutics, and a scientific advisory board member of A2 Biotherapeutics, BioLegend (Revvity) and Neuvogen. Work in the Schreiber laboratory was supported by research grants from the National Cancer Institute and the Parker Institute for Cancer Immunotherapy. The other authors declare no competing interests.

Peer review

Peer review information

Nature thanks the anonymous reviewers for their contribution to the peer review of this work. Peer reviewer reports are available.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data figures and tables

Extended Data Fig. 1 Localized mRNA-LNP vaccination can systemically prime CD8 T cells.

a–d, in vivo OT-I proliferation assay at day 0, 2, 3 and 7 after OVA-mRNA immunization. CTV-labelled CD45.1+ naïve OT-I cells were sorted and adoptively transferred to WT mice. 1 day later, mice were i.m. immunized with 10 μg OVA mRNA-LNP. On day 0, 2, 3, or 7 after immunization, spleen, peripheral blood, draining LN, contralateral LN, and mesentery LN were analysed for OT-I proliferation analysis. a, Schematic of experimental setup for a–c. Data is representative of biologically independent samples per day (0, 2, 3, and 7) from two independent experiments (n = 4 per group). b, Representative flow plots of OT-I proliferation. On each day, plots from different tissues are paired from the same mouse. c, Representative histograms of OT-I cells stained with CTV. d, CTV dilution (mean MFI) with different timepoints. (n = 4 each). Two-way ANOVA with multiple comparisons. ns = not significant. e, in vivo OT-I proliferation assay with inhibition of T cell egress from lymphoid tissues. 1 day after OT-I transfer, mice were intraperitoneally (i.p.) injected with PBS or 1 mg/kg body weight FTY720. 6 h later, mice were injected with OVA mRNA-LNP. (day 2: n = 2; day 3: n = 4 per group) two-tailed t-test. ***P = 0.0006. f, OT-I cell trafficking to lymphoid organs was blocked by splenectomy and anti-CD62L treatment every two days. 6 h after the first dose of anti-CD62L, splenectomized mice received CTV-labelled naïve OT-I cells followed by OVA mRNA-LNP immunization (n = 6 per genotype). two-tailed t-test. Data are represented as mean values ± s.d. of pooled biologically independent samples from two independent experiments. The diagram in a was created using BioRender; Jo, S. https://BioRender.com/62s9j87 (2026).

Extended Data Fig. 2 mRNA-LNP systemically primes CD8 T cells in lymphoid tissues.

a, Gating strategy for OT-I in the in vivo OT-I proliferation assay. b, Representative pregating flow plots of OT-I cells from peripheral blood, showing the transient absence of CD8a+ Va2+ CD45.1+ cells specifically on day 2, but not at other timepoints. c, CTV dilution (mean MFI) at the indicated timepoints, following in vivo OT-I proliferation assay at day 2 post OVA-mRNA immunization with different doses. One day after OT-I transfer, mice were immunized i.m. with 0, 0.1, 0.32, 1, 3.2, or 10 μg of OVA mRNA-LNP. Data are represented as mean values of pooled biologically independent samples from one representative experiment from two independent experiments. (n = 2 for each condition). Two-way ANOVA with multiple comparisons. d, e, in vivo OT-I proliferation assay at day 0, 1 and 2 after OVA-mRNA immunization as described in Fig. 1a. d, Representative flow plots showing CD69 expression by OT-I cells at indicated timepoints. e, Frequencies of CD69+ OT-I cells in different tissues across time. Data are represented as mean values of pooled biologically independent samples from two independent experiments. (n = 2 for day 0, n = 4 for day 1, and n = 3 for day 2).

Extended Data Fig. 3 CD8 T cell entry to lymphoid organs is critical for their priming.

a, Representative flow plots showing OT-I proliferation from Fig. 1f. Gating strategy for cDC1, cDC2, and B cells from LNs after mRNA-LNP immunization. b, c, in vivo OT-I proliferation assay with inhibition of T cell egress from LNs. CD45.1+ CTV+ naïve OT-I cells were adoptively transferred to CD45.2+ wild type mice. 1 day later, mice were intraperitoneally injected with PBS or 1 mg/kg body weight FTY720. 6 h later, mice were i.m. injected with OVA mRNA-LNP. b, Representative flow plots showing OT-I proliferation on day 2. c, Frequencies of proliferating OT-I cells. Data are represented as mean values ± s.d. of pooled biologically independent samples from two independent experiments. (n = 4 for each condition). Two-tailed t-tests. ***P = 0.0006, ns=not significant. d–f, OT-I cells in muscle are migrated from lymphoid organs. OT-I cell trafficking to lymphoid organs was blocked by splenectomy and anti-CD62L treatment every two days. 6 h after the first dose of anti-CD62L, splenectomized mice received CTV-labelled naïve OT-I cells followed by OVA mRNA-LNP immunization. Data are represented as mean values ± s.d. of pooled biologically independent samples from three independent experiments. (n = 6 for each condition). Two-way ANOVA. *P = 0.0486, ns=not significant.

Extended Data Fig. 4 Both cDC1 and cDC2 prime CD8 T cells after mRNA vaccination.

a, Gating strategy for cDC1, cDC2, and B cells from LNs after mRNA-LNP immunization. b, Gating strategy for tetramer+ CD8 T cells from spleen. c, ELISpot results from cDNA vaccination experiment of Fig. 2d,e. P** = 0.0037, P*** = 0.0008, ns=0.8135. d, e, WT and Δ32 mice were injected i.m. with 10 μg OVA adjuvanted with AddaVax on day 0 and 7. Spleens were stained for SIINFEKL-Kb tetramer+ CD8 T cells and CD44/CD62L expression on day 11. Data represents pooled biologically independent samples from three independent experiments. (n = 15 WT. n = 9 Δ32). One-way ANOVA with Tukey’s multiple comparisons. P** = 0.0081. f, IFNg ELISpot results from Fig. 3e,f. (n = 7 Cre−, n = 8 Cre+). One-way ANOVA with Tukey’s multiple comparisons. g, SIINFEKL-Kb-PE MFI from tetramer+ CD8 T cells from the experiment shown in Fig. 2f,g. (n = 9 WT, n = 11 Δ32, and n = 11 Δ1+2+3). One-way ANOVA with Tukey’s multiple comparisons. h, SIINFEKL-Kb tetramer+ CD8 T cells on day 7 after mLama4 mRNA-LPX vaccination in WT, Δ32, and Δ1+2+3 mice. (unimmunized: n = 4; immunized: n = 8 WT, n = 7 Δ32, and n = 7 Δ1+2+3). One-way ANOVA with Tukey’s multiple comparisons. P*** = 0.0003. Data represented as mean values ± s.d. and pooled biologically independent samples from two independent experiments. ns = not significant.

Extended Data Fig. 5 cDCs are required to prime CD8 T cells.

a, Verification of WT, Δ32, and Δ1+2+3 mice were injected i.m. with OVA mRNA-LNP. 7 days later, spleens were analysed for the presence of SIINFEKL-Kb tetramer+ CD8 T cells. One-way ANOVA b, c, Verification of splenic cDC depletion after DT treatment in CD11c-DTR mice from the experiments shown in Fig. 2h,i. One-way ANOVA. d, Verification of splenic cDC depletion after DT treatment in CD11c-DTR bone marrow chimera mice from the experiments shown in Fig. 2j. Two-tailed t-test. e, f, Quantification of macrophage population after DT treatment in CD11c-DTR mice from the experiments shown in Fig. 2h,i. One-way ANOVA. g, h, Quantification of inflammatory monocyte population after DT treatment in CD11c-DTR bone marrow chimeras from the experiments shown in Fig. 2j. Two-tailed t-test. i, Representative images of inguinal LN with or without DT treatment in CD11c-DTR BM chimeras from two independent experiments. CD11c is shown in turquoise, CD169 in red, F4/80 in green, and DAPI in blue. j–l, WT and Wdfy4−/− mice were injected with OVA mRNA-LNP on day 0 and 7. Spleens were stained for the presence of SIINFEKL-Kb tetramer+ CD8 T cells and their CD44 and CD62L surface expression on day 11. (n = 6 for WT, n = 7 for Wdfy4−/−). j, Representative flow plots of CD8 T cells. k, Quantification of tetramer+ CD44+ CD62L+ CD8 T cells as a percentage of CD8 T cells. Two-tailed t-test. l, IFNg ELISpot results. Two-tailed t-test. Data represented as mean values ± s.d. and pooled biologically independent samples from two independent experiments P**** <0.0001, ns = not significant.

Extended Data Fig. 6 cDCs and BM progenitors were exclusively derived from donor.

a, Gating strategy for splenic cDC1 and cDC2 to examine BM chimerism, from a representative MHC-I TKO → SJL BM chimera. b, Quantification of splenic cDC chimerism. c, Gating strategy for CDP, MDP, pre-cDC1, and pre-cDC2 populations to examine bone marrow chimerism. (n = 6 each). d, Quantification of BM progenitor chimerism. (n = 6 each). Data represented as mean values ± s.d. and pooled biologically independent samples from two independent experiments.

Extended Data Fig. 7 Type I IFN signalling mediates cross-dressing following mRNA vaccines.

a–d, Tetramer staining of CD8 T cells in peripheral blood (a, b) and draining LN (c, d) from IFNAR1 blockade experiment from Fig. 5. b, d, One-way ANOVA with Tukey’s multiple comparisons. P**** < 0.0001, P** = 0.0061. e, f, Itgax-Cre+ β2mfl/fl mice were immunized with OVA mRNA-LNP, with or without 2 mg anti-IFNAR1 treatment one day before immunization. 4 days after immunization, splenocytes were analysed for H2-Kb expression on cDCs. (n = 4 per genotype). Two-tailed t-test. Data are represented as mean values ± s.d. from pooled biologically independent samples from two independent experiments.

Extended Data Fig. 8 cDCs acquire non-self MHC-I from non-hematopoietic cells.

a, Acquisition of H2-Kb expression in CD45.2+ splenic cDCs from BM chimera of Fig. 3h,i. Data shown is representative flow plots from two independent experiments. b, Gating strategy for splenic cDC1 and cDC2. c, d, BM chimerism of B6 and BALB/c allogeneic BM chimeras from Fig. 4. c, Representative flow plots of lineage- cKithigh BM progenitors. d, Quantification of lineage- cKithigh BM progenitor chimerism. e, f, Acquisition of H2-Kb expression in Balb/c+ splenic cDCs from B6 and BALB/c allogeneic BM chimera shown in Fig. 4. Data shown is representative flow plots from two independent experiments. Two-tailed t-test. P* = 0.033. g, h Tetramer response from B6 and BALB/c allogeneic BM chimeras from Fig. 4. (For unimmunized control: n = 1–2 per condition. For OVA mRNA-LNP, n = 7 for BALB/c → B6, n = 3 for BALB/c → BALB/c, n = 6 for B6 → B6, and n = 6 for B6 → BALB/c mice). Data are represented as mean values ± s.d. from pooled biologically independent samples from two independent experiments.

Extended Data Fig. 9 CD8 T cell memory and functions.

a–c, Memory SIINFEKL-Kb tetramer+ CD8 T cells after OVA mRNA–LNP vaccination. WT, Δ32 and Δ1+2+3 mice were immunized on days 0 and 7 with OVA mRNA–LNP, and peripheral blood was analysed 6 weeks after the second dose. a, Frequencies of SIINFEKL-Kb tetramer+ CD8 T cells. One-way ANOVA with Tukey’s multiple comparisons test. b, Representative flow plots of tetramer+ CD8 T cells showing CD127 and KLRG1 expression. c, Quantification of CD127+ cells among tetramer+ CD8 T cells. (n = 8 WT, n = 7 Δ32, and n = 6 Δ1+2+3). One-way ANOVA. *P (left) = 0.0125, *P (right) = 0.0215. d–g, In vivo cytotoxicity assay. Six weeks after the second dose of OVA mRNA-LNP, mice received a 1:1 mixture of irrelevant peptide-loaded (CTV-low) or SIINFEKL-loaded (CTV-high) CD45.1+ splenocytes. d, Representative CTV histograms of transferred target cells. e, Killing efficiency normalized to WT. (n = 10 WT, n = 13 Δ32, and n = 9 Δ1+2+3). One-way ANOVA with Tukey’s multiple comparisons test. *P = 0.0474. f, g, Frequencies of SIINFEKL-Kb tetramer+ CD8 T cells in spleen (f) and LNs (g) (n = 8 WT, n = 9 Δ32, and n = 9 Δ1+2+3). ****P < 0.0001, **P = 0.0014, *P = 0.0429. Data are represented as mean values ± s.d. from pooled biologically independent samples from two independent experiments. ns = not significant.

Extended Data Fig. 10 Identification of clusters in single cell analysis of CD8 T cells.

Heatmap displaying pseudobulk expression profiles of the top 50 differentially expressed genes across experimental conditions, stratified by cell cluster. Gene expression was aggregated at the cluster level per condition to highlight transcriptional shifts in response to immunization.

Supplementary information

Reporting Summary (download PDF )

Supplementary Table 1 (download XLSX )

SIINFEKL-specific hyperexpanded (clone size > 20) clonotypes identified by scRNA-seq in individual mice.

Supplementary Table 2 (download XLSX )

Metadata associated with CD8+ T cell scRNA-seq and TCR-seq datasets available in GEO.

Peer Review File (download PDF )

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Jo, S., Li, L., Thakur, C. et al. mRNA vaccines engage unconventional pathways in CD8+ T cell priming. Nature (2026). https://doi.org/10.1038/s41586-026-10353-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Version of record:

  • DOI: https://doi.org/10.1038/s41586-026-10353-6

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing