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Cross-presentation by intercellular peptide transfer through gap junctions


Major histocompatibility complex (MHC) class I molecules present peptides that are derived from endogenous proteins1. These antigens can also be transferred to professional antigen-presenting cells in a process called cross-presentation, which precedes initiation of a proper T-cell response2,3; but exactly how they do this is unclear. We tested whether peptides can be transferred directly from the cytoplasm of one cell into the cytoplasm of its neighbour through gap junctions. Here we show that peptides with a relative molecular mass of up to 1,800 diffuse intercellularly through gap junctions unless a three-dimensional structure is imposed. This intercellular peptide transfer causes cytotoxic T-cell recognition of adjacent, innocent bystander cells as well as activated monocytes. Gap-junction-mediated peptide transfer is restricted to a few coupling cells owing to the high cytosolic peptidase activity. We present a mechanism of antigen acquisition for cross-presentation that couples the antigen presentation system of two adjacent cells and is lost in most tumours: gap-junction-mediated intercellular peptide coupling for presentation by bystander MHC class I molecules and transfer to professional antigen presenting cells for cross-priming.

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  1. 1

    Yewdell, J. W., Reits, E. & Neefjes, J. Making sense of mass destruction: quantitating MHC class I antigen presentation. Nature Rev. Immunol. 3, 952–961 (2003)

  2. 2

    Rock, K. L. The ins and outs of cross-presentation. Nature Immunol. 4, 941–943 (2003)

  3. 3

    Ackerman, A. L. & Cresswell, P. Cellular mechanisms governing cross-presentation of exogenous antigens. Nature Immunol. 5, 678–684 (2004)

  4. 4

    Heath, W. R. et al. Cross-presentation, dendritic cell subsets, and the generation of immunity to cellular antigens. Immunol. Rev. 199, 9–26 (2004)

  5. 5

    Segretain, D. & Falk, M. M. Regulation of connexin biosynthesis, assembly, gap junction formation, and removal. Biochim. Biophys. Acta 1662, 3–21 (2004)

  6. 6

    Goldberg, G. S., Valiunas, V. & Brink, P. R. Selective permeability of gap junction channels. Biochim. Biophys. Acta 1662, 96–101 (2004)

  7. 7

    Warn-Cramer, B. J. & Lau, A. F. Regulation of gap junctions by tyrosine protein kinases. Biochim. Biophys. Acta 1662, 81–95 (2004)

  8. 8

    Oviedo-Orta, E. & Evans, W. H. Gap junctions and connexin-mediated communication in the immune system. Biochim. Biophys. Acta 1662, 102–112 (2004)

  9. 9

    Goldberg, G. S. & Lau, A. F. Dynamics of connexin43 phosphorylation in pp60v-src-transformed cells. Biochem. J. 295, 735–742 (1993)

  10. 10

    Esinduy, C. B., Chang, C. C., Trosko, J. E. & Ruch, R. J. In vitro growth inhibition of neoplastically transformed cells by non-transformed cells: requirement for gap junctional intercellular communication. Carcinogenesis 16, 915–921 (1995)

  11. 11

    Fischer, N. O., Mbuy, G. N. & Woodruff, R. I. HSV-2 disrupts gap junctional intercellular communication by mammalian cells in vitro. J. Virol. Methods 91, 157–166 (2001)

  12. 12

    Oelze, I., Kartenbeck, J., Crusius, K. & Alonso, A. Human papilomavirus type 16 E5 protein affects cell-cell communication in an epithelial cell line. J. Virol. 69, 4489–4494 (1995)

  13. 13

    Mesnil, M. & Yamasaki, H. Bystander effect in herpes simplex virus–thymidine kinase/ganciclovir cancer gene therapy: role of gap-junctional intercellular communication. Cancer Res. 60, 3989–3999 (2000)

  14. 14

    Giepmans, B. N., Hengeveld, T., Postma, F. R. & Moolenaar, W. H. Interaction of c-Src with gap junction protein connexin-43. Role in the regulation of cell-cell communication. J. Biol. Chem. 276, 8544–8549 (2001)

  15. 15

    Reits, E. et al. Peptide diffusion, protection, and degradation in nuclear and cytoplasmic compartments before antigen presentation by MHC class I. Immunity 18, 97–108 (2003)

  16. 16

    Harks, E. G. et al. Besides affecting the intracellular calcium signaling, 2-APB reversibly blocks gap junctional coupling in confluent monolayers. FASEB J. 17, 941–943 (2003)

  17. 17

    Princiotta, M. F. et al. Quantitating protein synthesis, degradation, and endogenous antigen processing. Immunity 18, 343–354 (2003)

  18. 18

    Eugenin, E. A., Branes, M. C., Berman, J. W. & Saez, J. C. TNF-alpha plus IFN-gamma induce connexin43 expression and formation of gap junctions between human monocytes/macrophages that enhance physiological responses. J. Immunol. 170, 1320–1328 (2003)

  19. 19

    Savinov, A. Y., Wong, F. S., Stonebraker, A. C. & Chervonsky, A. V. Presentation of antigen by endothelial cells and chemoattraction are required for homing of insulin-specific CD8 + T cells. J. Exp. Med. 197, 643–656 (2003)

  20. 20

    Kurts, C., Heath, W. R., Carbone, F. R., Kosaka, H. & Miller, J. F. Cross-presentation of self antigens to CD8 + T cells: the balance between tolerance and autoimmunity. Novartis Found. Symp. 215, 172–181 (1998)

  21. 21

    Li, Z., Menoret, A. & Srivastava, P. Roles of heat-shock proteins in antigen presentation and cross-presentation. Curr. Opin. Immunol. 14, 45–51 (2002)

  22. 22

    Gromme, M. et al. Recycling MHC class I molecules and endosomal peptide loading. Proc. Natl Acad. Sci. USA 96, 10326–10331 (1999)

  23. 23

    Gil-Torregrosa, B. C. et al. Control of cross-presentation during dendritic cell maturation. Eur. J. Immunol. 34, 398–407 (2004)

  24. 24

    Guermonprez, P. et al. ER-phagosome fusion defines an MHC class I cross-presentation compartment in dendritic cells. Nature 425, 397–402 (2003)

  25. 25

    Houde, M. et al. Phagosomes are competent organelles for antigen cross-presentation. Nature 425, 402–406 (2003)

  26. 26

    Serna, A., Ramirez, M. C., Soukhanova, A. & Sigal, L. J. Cutting edge: efficient MHC class I cross-presentation during early vaccinia infection requires the transfer of proteasomal intermediates between antigen donor and presenting cells. J. Immunol. 171, 5668–5672 (2003)

  27. 27

    Wolkers, M. C., Brouwenstijn, N., Bakker, A. H., Toebes, M. & Schumacher, T. N. Antigen bias in T cell cross-priming. Science 304, 1314–1317 (2004)

  28. 28

    Norbury, C. C. et al. CD8 + T cell cross-priming via transfer of proteasome substrates. Science 304, 1318–1321 (2004)

  29. 29

    Shen, L. & Rock, K. L. Cellular protein is the source of cross-priming antigen in vivo. Proc. Natl Acad. Sci. USA 101, 3035–3040 (2004)

  30. 30

    Verra, N. C. et al. Human telomerase reverse transcriptase-transduced human cytotoxic T cells suppress the growth of human melanoma in immunodeficient mice. Cancer Res. 64, 2153–2161 (2004)

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We thank R. van Beem and E. Sellink for human monocyte and HUVEC isolations, W. Moolenaar, B. Giepmans and L. van Zeijl for A431/Cx43 cells and Cx43 reagents, R. Luiten and H. Spits for T-cell clone (InfA13TGA), W. E. Benckhuijsen for peptide synthesis, E. Mesman and M. Tjin-A-Koeng for immunohistochemistry, K. Jalink for experimental support, and H. Pickersgill, A. Griekspoor and M. Wolkers for critical reading. This work was supported by grants from the Dutch Cancer Society KWF.

Authors' contributions. J.N. and C.H. performed most experiments with support from E.R. Constructs were made by L.J. and peptides by J.W.D. Supervision by J.N.

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Correspondence to Joost Neijssen.

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The authors declare that they have no competing financial interests.

Supplementary information

Supplementary Movie

The fluorescence in the bottom-right cell was photo-inactivated (bleached) and recovered over time to the cost of fluorescence in the micro-injected cell. (AVI 3568 kb)

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Figure 1: Gap-junction-dependent intercellular peptide transfer.
Figure 2: Size and structure dependency for Cx43-mediated intercellular peptide transfer.
Figure 3: Gap-junction-mediated intercellular peptide transfer for cross-presentation by HLA-A2 molecules.
Figure 4: Gap-junction-mediated transfer of influenza virus antigens to APCs and CTL stimulation.


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