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Analysis of Individual Natural Killer Cell Responses

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Book cover Innate Immunity

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 415))

Summary

Typical assays for natural killer (NK) cell function assess the responses of entire NK cell populations. It is now possible to determine the responses of individual NK cells. Herein, two representative assays are described along with examples of how they have helped clarify current understanding of NK cell biology.

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References

  1. Trinchieri, G. (1989) Biology of natural killer cells. Adv Immunol 47, 187–376.

    Article  CAS  PubMed  Google Scholar 

  2. Yokoyama, W. M. (1999) Natural killer cells, Chapter 17. In Fundamental Immunology (Paul, W. E., Ed.), pp. 575–603, Lippincott-Raven, New York.

    Google Scholar 

  3. Herberman, R. (1982) NK Cells and Other Natural Effector Cells, Academic Press, New York.

    Google Scholar 

  4. Rosenberg, S. A., Lotze, M. T., Muul, L. M., Leitman, S., Chang, A. E., Ettinghausen, S. E., Matory, Y. L., Skibber, J. M., Shiloni, E., Vetto, J. T., Seipp, C. A., Simpson, C., and Reichert, C. M. (1985) Observations on the systemic administration of autologous lymphokine-activated killer cells and recombinant interleukin-2 to patients with metastatic cancer. N Engl J Med 313, 1485–92.

    Article  CAS  PubMed  Google Scholar 

  5. Bancroft, G. J. (1993) The role of natural killer cells in innate resistance to infection. Curr Opin Immunol 5, 503–10.

    Article  CAS  PubMed  Google Scholar 

  6. Biron, C. A., Byron, K. S., and Sullivan, J. L. (1989) Severe herpesvirus infections in an adolescent without natural killer cells. N Engl J Med 320, 1731–5.

    Article  CAS  PubMed  Google Scholar 

  7. Shellam, G. R., Allan, J. E., Papadimitriou, J. M., and Bancroft, G. J. (1981) Increased susceptibility to cytomegalovirus infection in beige mutant mice. Proc Natl Acad Sci USA 78, 5104–08.

    Article  CAS  PubMed  Google Scholar 

  8. Bukowski, J. F., Woda, B. A., Habu, S., Okumura, K., and Welsh, R. M. (1983) Natural killer cell depletion enhances virus synthesis and virus-induced hepatitis in vivo. J Immunol 131, 1531–8.

    CAS  PubMed  Google Scholar 

  9. Bukowski, J. F., Woda, B. A., and Welsh, R. M. (1984) Pathogenesis of murine cytomegalovirus infection in natural killer cell-depleted mice. J Virol 52, 119–28.

    CAS  PubMed  Google Scholar 

  10. Welsh, R. M., Dundon, P. L., Eynon, E. E., Brubaker, J. O., Koo, G. C., and O’Donnell, C. L. (1990) Demonstration of the antiviral role of natural killer cells in vivo with a natural killer cell-specific monoclonal antibody (NK 1.1). Nat Immun Cell Growth Regul 9, 112–20.

    CAS  PubMed  Google Scholar 

  11. Orange, J. S., and Biron, C. A. (1996) An absolute and restricted requirement for IL-12 in natural killer cell IFN-gamma production and antiviral defense. Studies of natural killer and T cell responses in contrasting viral infections. J Immunol 156, 1138–42.

    CAS  PubMed  Google Scholar 

  12. Loh, J., Chu, D. T., O’Guin, A. K., Yokoyama, W. M., and Virgin, H. W. t. (2005) Natural killer cells utilize both perforin and gamma interferon to regulate murine cytomegalovirus infection in the spleen and liver. J Virol 79, 661–7.

    Article  CAS  PubMed  Google Scholar 

  13. Kennedy, M. K., Glaccum, M., Brown, S. N., Butz, E. A., Viney, J. L., Embers, M., Matsuki, N., Charrier, K., Sedger, L., Willis, C. R., Brasel, K., Morrissey, P. J., Stocking, K., Schuh, J. C., Joyce, S., and Peschon, J. J. (2000) Reversible defects in natural killer and memory CD8 T cell lineages in interleukin 15-deficient mice. J Exp Med 191, 771–80.

    Article  CAS  PubMed  Google Scholar 

  14. Henkart, P. A. (1994) Lymphocyte-mediated cytotoxicity: two pathways and multiple effector molecules. Immunity 1, 343–46.

    Article  CAS  PubMed  Google Scholar 

  15. Biron, C. A., Nguyen, K. B., Pien, G. C., Cousens, L. P., and Salazar-Mather, T. P. (1999) Natural killer cells in antiviral defense: function and regulation by innate cytokines. Annu Rev Immunol 17, 189–220.

    Article  CAS  PubMed  Google Scholar 

  16. Grimm, E. A., Robb, R. J., Roth, J. A., Neckers, L. M., Lachman, L. B., Wilson, D. J., and Rosenberg, S. A. (1983) Lymphokine-activated killer cell phenomenon. III. Evidence that IL-2 is sufficient for direct activation of peripheral blood lymphocytes into lymphokine-activated killer cells. J Exp Med 158, 1356–61.

    Article  CAS  PubMed  Google Scholar 

  17. Orange, J. S., and Biron, C. A. (1996) Characterization of early IL-12, IFN-alphabeta, and TNF effects on antiviral state and NK cell responses during murine cytomegalovirus infection. J Immunol 156, 4746–56.

    CAS  PubMed  Google Scholar 

  18. Carnaud, C., Lee, D., Donnars, O., Park, S. H., Beavis, A., Koezuka, Y., and Bendelac, A. (1999) Cutting edge: cross-talk between cells of the innate immune system: NKT cells rapidly activate NK cells. J Immunol 163, 4647–50.

    CAS  PubMed  Google Scholar 

  19. Nguyen, K. B., Salazar-Mather, T. P., Dalod, M. Y., Van Deusen, J. B., Wei, X. Q., Liew, F. Y., Caligiuri, M. A., Durbin, J. E., and Biron, C. A. (2002) Coordinated and distinct roles for IFN-alphabeta, IL-12, and IL-15 regulation of NK cell responses to viral infection. J Immunol 169, 4279–87.

    CAS  PubMed  Google Scholar 

  20. Andrews, D. M., Scalzo, A. A., Yokoyama, W. M., Smyth, M. J., and Degli-Esposti, M. A. (2003) Functional interactions between dendritic cells and NK cells during viral infection. Nat Immunol 4, 175–81.

    Article  CAS  PubMed  Google Scholar 

  21. Krug, A., French, A. R., Barchet, W., Fischer, J. A. A., Dzionek, A., Pingel, J. T., Orihuela, M. M., Akira, S., Yokoyama, W. M., and Colonna, M. (2004) TLR9-dependent recognition of MCMV by IPC and DC generates coordinated cytokine responses that activate antiviral NK cell function. Immunity 21, 107–19.

    Article  CAS  PubMed  Google Scholar 

  22. Lanier, L. L., Phillips, J. H., Hackett, J., Jr., Tutt, M., and Kumar, V. (1986) Natural killer cells: definition of a cell type rather than a function. J Immunol 137, 2735–9.

    CAS  PubMed  Google Scholar 

  23. Ljunggren, H. G., and Karre, K. (1990) In search of the ‘missing self’: MHC molecules and NK cell recognition. Immunol Today 11, 237–44.

    Article  CAS  PubMed  Google Scholar 

  24. Karlhofer, F. M., Ribaudo, R. K., and Yokoyama, W. M. (1992) MHC class I alloantigen specificity of Ly-49+ IL-2-activated natural killer cells. Nature 358, 66–70.

    Article  CAS  PubMed  Google Scholar 

  25. Yokoyama, W. M. (1997) What goes up must come down: the emerging spectrum of inhibitory receptors. J Exp Med 186, 1803–08.

    Article  CAS  PubMed  Google Scholar 

  26. Long, E. O. (1999) Regulation of immune responses through inhibitory receptors. Annu Rev Immunol 17, 875–904.

    Article  CAS  PubMed  Google Scholar 

  27. Yokoyama, W. M. (1995) Natural killer cell receptors. Curr Opin Immunol 7, 110–20.

    Article  CAS  PubMed  Google Scholar 

  28. Furukawa, H., Yabe, T., Watanabe, K., Miyamoto, R., Miki, A., Akaza, T., Tadokoro, K., Tohma, S., Inoue, T., Yamamoto, K., and Juji, T. (1999) Tolerance of NK and LAK activity for HLA class I-deficient targets in a TAP1-deficient patient (bare lymphocyte syndrome type I). Hum Immunol 60, 32–40.

    Article  CAS  PubMed  Google Scholar 

  29. Vitale, M., Zimmer, J., Castriconi, R., Hanau, D., Donato, L., Bottino, C., Moretta, L., de la Salle, H., and Moretta, A. (2002) Analysis of natural killer cells in TAP2-deficient patients: expression of functional triggering receptors and evidence for the existence of inhibitory receptor(s) that prevent lysis of normal autologous cells. Blood 99, 1723–9.

    Article  CAS  PubMed  Google Scholar 

  30. Bix, M., Liao, N. S., Zijlstra, M., Loring, J., Jaenisch, R., and Raulet, D. (1991) Rejection of class I MHC-deficient haemopoietic cells by irradiated MHC-matched mice. Nature 349, 329–31.

    Article  CAS  PubMed  Google Scholar 

  31. Hoglund, P., Ohlen, C., Carbone, E., Franksson, L., Ljunggren, H. G., Latour, A., Koller, B., and Kärre, K. (1991) Recognition of beta 2-microglobulin- negative (beta 2m-) T-cell blasts by natural killer cells from normal but not from beta 2m- mice: nonresponsiveness controlled by beta 2m- bone marrow in chimeric mice. Proc Natl Acad Sci USA 88, 10332–36.

    Article  CAS  PubMed  Google Scholar 

  32. Liao, N. S., Bix, M., Zijlstra, M., Jaenisch, R., and Raulet, D. (1991) MHC class I deficiency: susceptibility to natural killer (NK) cells and impaired NK activity. Science 253, 199–202.

    Article  CAS  PubMed  Google Scholar 

  33. Ljunggren, H. G., Van Kaer, L., Ploegh, H. L., and Tonegawa, S. (1994) Altered natural killer cell repertoire in Tap-1 mutant mice. Proc Natl Acad Sci USA 91, 6520–4.

    Article  CAS  PubMed  Google Scholar 

  34. Dorfman, J. R., Zerrahn, J., Coles, M. C., and Raulet, D. H. (1997) The basis for self-tolerance of natural killer cells in beta2-microglobulin- and TAP-1- mice. J Immunol 159, 5219–25.

    CAS  PubMed  Google Scholar 

  35. Ohlen, C., Kling, G., Hoglund, P., Hansson, M., Scangos, G., Bieberich, C., Jay, G., and Karre, K. (1989) Prevention of allogeneic bone marrow graft rejection by H-2 transgene in donor mice. Science 246, 666–8.

    Article  CAS  PubMed  Google Scholar 

  36. Bennett, M., Yu, Y. Y., Stoneman, E., Rembecki, R. M., Mathew, P. A., Lindahl, K. F., and Kumar, V. (1995) Hybrid resistance: ‘negative’ and ‘positive’ signaling of murine natural killer cells. Semin Immunol 7, 121–7.

    Article  CAS  PubMed  Google Scholar 

  37. Raulet, D. H., Vance, R. E., and McMahon, C. W. (2001) Regulation of the natural killer cell receptor repertoire. Annu Rev Immunol 19, 291–330.

    Article  CAS  PubMed  Google Scholar 

  38. Sentman, C. L., Olsson, M. Y., and Karre, K. (1995) Missing self recognition by natural killer cells in MHC class I transgenic mice. A ‘receptor calibration’ model for how effector cells adapt to self. Semin Immunol 7, 109–19.

    Article  CAS  PubMed  Google Scholar 

  39. Kim, S., Iizuka, K., Kang, H. S., Dokun, A., French, A. R., Greco, S., and Yokoyama, W. M. (2002) In vivo developmental stages in murine natural killer cell maturation. Nat Immunol 3, 523–8.

    Article  PubMed  Google Scholar 

  40. Kim, S., Poursine-Laurent, J., Truscott, S. M., Lybarger, L., Song, Y. J., Yang, L., French, A. R., Sunwoo, J. B., Lemieux, S., Hansen, T. H., and Yokoyama, W. M. (2005) Licensing of natural killer cells by host major histocompatibility complex class I molecules. Nature 436, 709–13.

    Article  CAS  PubMed  Google Scholar 

  41. Brown, M. G., Dokun, A. O., Heusel, J. W., Smith, H. R., Beckman, D. L., Blattenberger, E. A., Dubbelde, C. E., Stone, L. R., Scalzo, A. A., and Yokoyama, W. M. (2001) Vital involvement of a natural killer cell activation receptor in resistance to viral infection. Science 292, 934–7.

    Article  CAS  PubMed  Google Scholar 

  42. Scalzo, A. A., Fitzgerald, N. A., Simmons, A., La Vista, A. B., and Shellam, G. R. (1990) Cmv-1, a genetic locus that controls murine cytomegalovirus replication in the spleen. J Exp Med 171, 1469–83.

    Article  CAS  PubMed  Google Scholar 

  43. Scalzo, A. A., Lyons, P. A., Fitzgerald, N. A., Forbes, C. A., Yokoyama, W. M., and Shellam, G. R. (1995) Genetic mapping of Cmv1 in the region of mouse chromosome 6 encoding the NK gene complex-associated loci Ly49 and musNKR-P1. Genomics 27, 435–41.

    Article  CAS  PubMed  Google Scholar 

  44. Forbes, C. A., Brown, M. G., Cho, R., Shellam, G. R., Yokoyama, W. M., and Scalzo, A. A. (1997) The Cmv1 host resistance locus is closely linked to the Ly49 multigene family within the natural killer cell gene complex on mouse chromosome 6. Genomics 41, 406–13.

    Article  CAS  PubMed  Google Scholar 

  45. Depatie, C., Muise, E., Lepage, P., Gros, P., and Vidal, S. M. (1997) High-resolution linkage map in the proximity of the host resistance locus CMV1. Genomics 39, 154–63.

    Article  CAS  PubMed  Google Scholar 

  46. Brown, M. G., Scalzo, A. A., Matsumoto, K., and Yokoyama, W. M. (1997) The natural killer gene complex-a genetic basis for understanding natural killer cell function and innate immunity. Immunol Rev 155, 53–65.

    Article  CAS  PubMed  Google Scholar 

  47. Scalzo, A. A., Fitzgerald, N. A., Wallace, C. R., Gibbons, A. E., Smart, Y. C., Burton, R. C., and Shellam, G. R. (1992) The effect of the Cmv-1 resistance gene, which is linked to the natural killer cell gene complex, is mediated by natural killer cells. J Immunol 149, 581–9.

    CAS  PubMed  Google Scholar 

  48. Lee, S. H., Girard, S., Macina, D., Busa, M., Zafer, A., Belouchi, A., Gros, P., and Vidal, S. M. (2001) Susceptibility to mouse cytomegalovirus is associated with deletion of an activating natural killer cell receptor of the C-type lectin superfamily. Nat Genet 28, 42–5.

    Article  CAS  PubMed  Google Scholar 

  49. Daniels, K. A., Devora, G., Lai, W. C., O’Donnell, C. L., Bennett, M., and Welsh, R. M. (2001) Murine cytomegalovirus is regulated by a discrete subset of natural killer cells reactive with monoclonal antibody to ly49h. J Exp Med 194, 29–44.

    Article  CAS  PubMed  Google Scholar 

  50. Lee, S. H., Zafer, A., de Repentigny, Y., Kothary, R., Tremblay, M. L., Gros, P., Duplay, P., Webb, J. R., and Vidal, S. M. (2003) Transgenic expression of the activating natural killer receptor Ly49H confers resistance to cytomegalovirus in genetically susceptible mice. J Exp Med 197, 515–26.

    Article  CAS  PubMed  Google Scholar 

  51. Sjolin, H., Tomasello, E., Mousavi-Jazi, M., Bartolazzi, A., Karre, K., Vivier, E., and Cerboni, C. (2002) Pivotal role of KARAP/DAP12 adaptor molecule in the natural killer cell-mediated resistance to murine cytomegalovirus infection. J Exp Med 195, 825–34.

    Google Scholar 

  52. Smith, K. M., Wu, J., Bakker, A. B., Phillips, J. H., and Lanier, L. L. (1998) Cutting edge: Ly-49D and Ly-49H associate with mouse DAP12 and form activating receptors. J Immunol 161, 7–10.

    CAS  PubMed  Google Scholar 

  53. Smith, H. R., Heusel, J. W., Mehta, I. K., Kim, S., Dorner, B. G., Naidenko, O. V., Iizuka, K., Furukawa, H., Beckman, D. L., Pingel, J. T., Scalzo, A. A., Fremont, D. H., and Yokoyama, W. M. (2002) Recognition of a virus-encoded ligand by a natural killer cell activation receptor. Proc Natl Acad Sci USA 99, 8826–31.

    CAS  PubMed  Google Scholar 

  54. Arase, H., Mocarski, E. S., Campbell, A. E., Hill, A. B., and Lanier, L. L. (2002) Direct recognition of cytomegalovirus by activating and inhibitory NK cell receptors. Science 296, 1323–6.

    Article  CAS  PubMed  Google Scholar 

  55. Cuturi, M. C., Anegon, I., Sherman, F., Loudon, R., Clark, S. C., Perussia, B., and Trinchieri, G. (1989) Production of hematopoietic colony stimulating factors by human natural killer cells. J Exp Med 169, 569.

    Article  CAS  PubMed  Google Scholar 

  56. Kim, S., and Yokoyama, W. M. (1998) NK cell granule exocytosis and cytokine production inhibited by Ly-49A engagement. Cell Immunol 183, 106–12.

    Article  CAS  PubMed  Google Scholar 

  57. Dorner, B. G., Smith, H. R. C., French, A. R., Kim, S., Poursine-Laurent, J., Beckman, D. L., Pingel, J. T., Kroczek, R. A., and Yokoyama, W. M. (2004) Coordinate expression of cytokines and chemokines by natural killer cells during murine cytomegalovirus infection. J Immunol 172, 3119–31.

    CAS  PubMed  Google Scholar 

  58. Dokun, A. O., Kim, S., Smith, H. R., Kang, H. S., Chu, D. T., and Yokoyama, W. M. (2001) Specific and nonspecific NK cell activation during virus infection. Nat Immunol 2, 951–6.

    Article  CAS  PubMed  Google Scholar 

  59. Biron, C. A., Sonnenfeld, G., and Welsh, R. M. (1984) Interferon induces natural killer cell blastogenesis in vivo. J Leukoc Biol 35, 31–7.

    CAS  PubMed  Google Scholar 

  60. Tough, D. F., Borrow, P., and Sprent, J. (1996) Induction of bystander T cell proliferation by viruses and type I interferon in vivo. Science 272, 1947–50.

    Article  CAS  PubMed  Google Scholar 

  61. Kung, S. K., Su, R. C., Shannon, J., and Miller, R. G. (1999) The NKR-P1B gene product is an inhibitory receptor on SJL/J NK cells. J Immunol 162, 5876–87.

    CAS  PubMed  Google Scholar 

  62. Carlyle, J. R., Martin, A., Mehra, A., Attisano, L., Tsui, F. W., and Zuniga-Pflucker, J. C. (1999) Mouse NKR-P1B, a novel NK1.1 antigen with inhibitory function. J Immunol 162, 5917–23.

    CAS  PubMed  Google Scholar 

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Yokoyama, W.M., Kim, S. (2008). Analysis of Individual Natural Killer Cell Responses. In: Ewbank, J., Vivier, E. (eds) Innate Immunity. Methods in Molecular Biology™, vol 415. Humana Press. https://doi.org/10.1007/978-1-59745-570-1_11

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  • DOI: https://doi.org/10.1007/978-1-59745-570-1_11

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-746-4

  • Online ISBN: 978-1-59745-570-1

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