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Analysis of Interleukin-4-Induced Class Switch Recombination in Mouse Myeloma CH12F3-2 Cells

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Immune Mediators in Cancer

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2108))

Abstract

Affinity maturation of B lymphocytes is a process that includes somatic hypermutation and class switch recombination. Class switch recombination is a fundamental factor of the human adaptive immunity. The perturbation of this process has an adverse effect on human health, and results in global chromosome rearrangements and cell transformation. Evaluation of the class switch recombination efficiency is an important component of laboratory diagnosis of immunotoxic components. Here we describe a method for testing the efficiency of the class switch recombination. Cultivation of mouse myeloma CH12F3-2 cell line with anti-CD40 antibodies, transforming growth factor beta, and recombinant interleukin-4 (IL-4) triggers a cascade of signal transduction network events that lead to switching the immunoglobulin isotypes from IgM to IgA. This chapter describes the methodology of class switch recombination assay for assessment of the effect of environmental pollutants in toxicological laboratory diagnostics.

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References

  1. Carroll MC, Isenman DE (2012) Regulation of humoral immunity by complement. Immunity 37(2):199–207. https://doi.org/10.1016/j.immuni.2012.08.002

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Kawakami T, Galli SJ (2002) Regulation of mast-cell and basophil function and survival by IgE. Nat Rev Immunol 2(10):773–786. https://doi.org/10.1038/nri914

    Article  CAS  PubMed  Google Scholar 

  3. Macpherson AJ, Geuking MB, McCoy KD (2012) Homeland security: IgA immunity at the frontiers of the body. Trends Immunol 33(4):160–167. https://doi.org/10.1016/j.it.2012.02.002

    Article  CAS  PubMed  Google Scholar 

  4. Nimmerjahn F, Ravetch JV (2011) FcgammaRs in health and disease. Curr Top Microbiol Immunol 350:105–125. https://doi.org/10.1007/82_2010_86

    Article  CAS  PubMed  Google Scholar 

  5. Pabst O (2012) New concepts in the generation and functions of IgA. Nat Rev Immunol 12(12):821–832. https://doi.org/10.1038/nri3322

    Article  CAS  PubMed  Google Scholar 

  6. Roopenian DC, Akilesh S (2007) FcRn: the neonatal Fc receptor comes of age. Nat Rev Immunol 7(9):715–725. https://doi.org/10.1038/nri2155

    Article  CAS  PubMed  Google Scholar 

  7. Nakamura M, Kondo S, Sugai M, Nazarea M, Imamura S, Honjo T (1996) High frequency class switching of an IgM+ B lymphoma clone CH12F3 to IgA+ cells. Int Immunol 8(2):193–201

    Article  CAS  Google Scholar 

  8. Pene J, Rousset F, Briere F, Chretien I, Paliard X, Banchereau J, Spits H, De Vries JE (1988) IgE production by normal human B cells induced by alloreactive T cell clones is mediated by IL-4 and suppressed by IFN-gamma. J Immunol 141(4):1218–1224

    CAS  PubMed  Google Scholar 

  9. Vercelli D, Jabara HH, Arai K, Yokota T, Geha RS (1989) Endogenous interleukin 6 plays an obligatory role in interleukin 4-dependent human IgE synthesis. Eur J Immunol 19(8):1419–1424. https://doi.org/10.1002/eji.1830190811

    Article  CAS  PubMed  Google Scholar 

  10. Zhang K, Clark EA, Saxon A (1991) CD40 stimulation provides an IFN-gamma-independent and IL-4-dependent differentiation signal directly to human B cells for IgE production. J Immunol 146(6):1836–1842

    CAS  PubMed  Google Scholar 

  11. Tran TH, Nakata M, Suzuki K, Begum NA, Shinkura R, Fagarasan S, Honjo T, Nagaoka H (2010) B cell-specific and stimulation-responsive enhancers derepress Aicda by overcoming the effects of silencers. Nat Immunol 11(2):148–154. https://doi.org/10.1038/ni.1829

    Article  CAS  PubMed  Google Scholar 

  12. Crouch EE, Li Z, Takizawa M, Fichtner-Feigl S, Gourzi P, Montano C, Feigenbaum L, Wilson P, Janz S, Papavasiliou FN, Casellas R (2007) Regulation of AID expression in the immune response. J Exp Med 204(5):1145–1156. https://doi.org/10.1084/jem.20061952

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Dedeoglu F, Horwitz B, Chaudhuri J, Alt FW, Geha RS (2004) Induction of activation-induced cytidine deaminase gene expression by IL-4 and CD40 ligation is dependent on STAT6 and NFkappaB. Int Immunol 16(3):395–404

    Article  CAS  Google Scholar 

  14. Huong le T, Kobayashi M, Nakata M, Shioi G, Miyachi H, Honjo T, Nagaoka H (2013) In vivo analysis of Aicda gene regulation: a critical balance between upstream enhancers and intronic silencers governs appropriate expression. PLoS One 8(4):e61433. https://doi.org/10.1371/journal.pone.0061433

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Poltoratsky V, Goodman MF, Scharff MD (2000) Error-prone candidates vie for somatic mutation. J Exp Med 192(10):F27–F30

    Article  CAS  Google Scholar 

  16. Poltoratsky VP, Wilson SH, Kunkel TA, Pavlov YI (2004) Recombinogenic phenotype of human activation-induced cytosine deaminase. J Immunol 172(7):4308–4313

    Article  CAS  Google Scholar 

  17. Chang TP, Vancurova I (2013) NFkappaB function and regulation in cutaneous T-cell lymphoma. Am J Cancer Res 3(5):433–445

    PubMed  PubMed Central  Google Scholar 

  18. DiDonato JA, Mercurio F, Karin M (2012) NF-kappaB and the link between inflammation and cancer. Immunol Rev 246(1):379–400. https://doi.org/10.1111/j.1600-065X.2012.01099.x

    Article  CAS  PubMed  Google Scholar 

  19. Li MO, Wan YY, Sanjabi S, Robertson AK, Flavell RA (2006) Transforming growth factor-beta regulation of immune responses. Annu Rev Immunol 24:99–146. https://doi.org/10.1146/annurev.immunol.24.021605.090737

    Article  CAS  PubMed  Google Scholar 

  20. Massague J (2012) TGFbeta signalling in context. Nat Rev Mol Cell Biol 13(10):616–630. https://doi.org/10.1038/nrm3434

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Muramatsu M, Kinoshita K, Fagarasan S, Yamada S, Shinkai Y, Honjo T (2000) Class switch recombination and hypermutation require activation-induced cytidine deaminase (AID), a potential RNA editing enzyme. Cell 102(5):553–563. https://doi.org/10.1016/s0092-8674(00)00078-7

    Article  CAS  PubMed  Google Scholar 

  22. Petersen S, Casellas R, Reina-San-Martin B, Chen HT, Difilippantonio MJ, Wilson PC, Hanitsch L, Celeste A, Muramatsu M, Pilch DR, Redon C, Ried T, Bonner WM, Honjo T, Nussenzweig MC, Nussenzweig A (2001) AID is required to initiate Nbs1/gamma-H2AX focus formation and mutations at sites of class switching. Nature 414(6864):660–665

    Article  CAS  Google Scholar 

  23. Matthews AJ, Zheng S, DiMenna LJ, Chaudhuri J (2014) Regulation of immunoglobulin class-switch recombination: choreography of noncoding transcription, targeted DNA deamination, and long-range DNA repair. Adv Immunol 122:1–57. https://doi.org/10.1016/b978-0-12-800267-4.00001-8

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Battey J, Moulding C, Taub R, Murphy W, Stewart T, Potter H, Lenoir G, Leder P (1983) The human c-myc oncogene: structural consequences of translocation into the IgH locus in Burkitt lymphoma. Cell 34(3):779–787

    Article  CAS  Google Scholar 

  25. Neri A, Barriga F, Knowles DM, Magrath IT, Dalla-Favera R (1988) Different regions of the immunoglobulin heavy-chain locus are involved in chromosomal translocations in distinct pathogenetic forms of Burkitt lymphoma. Proc Natl Acad Sci U S A 85(8):2748–2752

    Article  CAS  Google Scholar 

  26. Shiramizu B, Barriga F, Neequaye J, Jafri A, Dalla-Favera R, Neri A, Guttierez M, Levine P, Magrath I (1991) Patterns of chromosomal breakpoint locations in Burkitt’s lymphoma: relevance to geography and Epstein-Barr virus association. Blood 77(7):1516–1526

    Article  CAS  Google Scholar 

  27. Akasaka H, Akasaka T, Kurata M, Ueda C, Shimizu A, Uchiyama T, Ohno H (2000) Molecular anatomy of BCL6 translocations revealed by long-distance polymerase chain reaction-based assays. Cancer Res 60(9):2335–2341

    CAS  PubMed  Google Scholar 

  28. Offit K, Louie DC, Parsa NZ, Roy P, Leung D, Lo Coco F, Zelenetz A, Dalla-Favera R, Chaganti RS (1995) BCL6 gene rearrangement and other cytogenetic abnormalities in diffuse large cell lymphoma. Leuk Lymphoma 20(1-2):85–89

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Pharmaceutical Sciences, St. John’s University, Queens, NY, USA

    Wenjun Wu, Zhihui Xiao & Vladimir Poltoratsky

  2. Department of Biological Sciences, St. John’s University, Queens, NY, USA

    Deon Buritis & Vladimir Poltoratsky

Authors
  1. Wenjun Wu
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  2. Zhihui Xiao
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  3. Deon Buritis
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  4. Vladimir Poltoratsky
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Corresponding author

Correspondence to Vladimir Poltoratsky .

Editor information

Editors and Affiliations

  1. Department of Biological Sciences, St. John’s University, Queens, NY, USA

    Ivana Vancurova

  2. Department of Biological Sciences, St. John’s University, Queens, NY, USA

    Yan Zhu

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Wu, W., Xiao, Z., Buritis, D., Poltoratsky, V. (2020). Analysis of Interleukin-4-Induced Class Switch Recombination in Mouse Myeloma CH12F3-2 Cells. In: Vancurova, I., Zhu, Y. (eds) Immune Mediators in Cancer. Methods in Molecular Biology, vol 2108. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0247-8_15

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  • DOI: https://doi.org/10.1007/978-1-0716-0247-8_15

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0246-1

  • Online ISBN: 978-1-0716-0247-8

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