Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1989 Dec 25;17(24):10455–10472. doi: 10.1093/nar/17.24.10455

Effects of anti-IgM suppression on polyclonally activated murine B cells: analysis of immunoglobulin mRNA, gene specific nuclear factors and cell cycle distribution.

A Marcuzzi 1, B Van Ness 1, T Rouse 1, D Lafrenz 1
PMCID: PMC335312  PMID: 2481271

Abstract

Polyclonal activation of murine B cells with bacterial lipopolysaccharide (LPS) and dextran sulfate (DxS) results in cell proliferation as well as increased immunoglobulin gene transcription and antibody secretion. When added to B cell cultures during mitogen activation, anti-mu antibody suppresses the rate of proliferation and immunoglobulin gene expression. Using this model system we show that co-cultures of B cells with LPS/DxS and anti-mu resulted in a decrease of both mu and kappa chain mRNA. Suppression did not prevent B cell entry into cycle nor a significant alteration in the distribution of cells in phases of cell cycle, although it did prolong the cycle transit time in a dose dependent fashion as determined by bromodeoxyuridine pulse labelling. Analysis of B cell specific nuclear binding factors, which previously have been shown to be important in regulating immunoglobulin gene transcription were examined. Results show that the kappa-specific enhancer binding activity of NF-kappa B was induced in activated as well as suppressed cultures. The lymphoid specific factor NF-A2, which recognizes the octamer sequence motif in the promoters of immunoglobulin genes, was induced by the polyclonal activation but was selectively lost in extracts from suppressed cells. Thus, specific regulation of the nuclear factor which plays a critical role in both heavy and light chain immunoglobulin gene expression may contribute to the transcriptional suppression observed in anti-mu treated B cells.

Full text

PDF
10455

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Andersson J., Coutinho A., Melchers F. Stimulation of murine B lymphocytes to IgG synthesis and secretion by the mitogens lipopolysaccharide and lipoprotein and its inhibition by anti-immunoglobulin antibodies. Eur J Immunol. 1978 May;8(5):336–343. doi: 10.1002/eji.1830080509. [DOI] [PubMed] [Google Scholar]
  2. Andersson J., Sjöberg O., Möller G. Mitogens as probes for immunocyte activation and cellular cooperation. Transplant Rev. 1972;11:131–177. doi: 10.1111/j.1600-065x.1972.tb00048.x. [DOI] [PubMed] [Google Scholar]
  3. Atchison M. L., Perry R. P. The role of the kappa enhancer and its binding factor NF-kappa B in the developmental regulation of kappa gene transcription. Cell. 1987 Jan 16;48(1):121–128. doi: 10.1016/0092-8674(87)90362-x. [DOI] [PubMed] [Google Scholar]
  4. Baeuerle P. A., Baltimore D. I kappa B: a specific inhibitor of the NF-kappa B transcription factor. Science. 1988 Oct 28;242(4878):540–546. doi: 10.1126/science.3140380. [DOI] [PubMed] [Google Scholar]
  5. Bergman Y., Rice D., Grosschedl R., Baltimore D. Two regulatory elements for immunoglobulin kappa light chain gene expression. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7041–7045. doi: 10.1073/pnas.81.22.7041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chen-Bettecken U., Wecker E., Schimpl A. IgM RNA switch from membrane to secretory form is prevented by adding antireceptor antibody to bacterial lipopolysaccharide-stimulated murine primary B-cell cultures. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7384–7388. doi: 10.1073/pnas.82.21.7384. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Darzynkiewicz Z., Sharpless T., Staiano-Coico L., Melamed M. R. Subcompartments of the G1 phase of cell cycle detected by flow cytometry. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6696–6699. doi: 10.1073/pnas.77.11.6696. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Darzynkiewicz Z., Traganos F., Sharpless T., Melamed M. R. Lymphocyte stimulation: a rapid multiparameter analysis. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2881–2884. doi: 10.1073/pnas.73.8.2881. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dignam J. D., Lebovitz R. M., Roeder R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. doi: 10.1093/nar/11.5.1475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Falkner F. G., Zachau H. G. Correct transcription of an immunoglobulin kappa gene requires an upstream fragment containing conserved sequence elements. Nature. 1984 Jul 5;310(5972):71–74. doi: 10.1038/310071a0. [DOI] [PubMed] [Google Scholar]
  11. Flahart R. E., Lawton A. R. Anti-mu antibody blocks LPS driven B cell differentiation by suppressing specific mRNAs. J Mol Cell Immunol. 1987;3(2):61–68. [PubMed] [Google Scholar]
  12. Hromas R., Pauli U., Marcuzzi A., Lafrenz D., Nick H., Stein J., Stein G., Van Ness B. Inducible DNA-protein interactions of the murine kappa immunoglobulin enhancer in intact cells: comparison with in vitro interactions. Nucleic Acids Res. 1988 Feb 11;16(3):953–967. doi: 10.1093/nar/16.3.953. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hromas R., Van Ness B. Nuclear factors bind to regulatory regions of the mouse kappa immunoglobulin gene. Nucleic Acids Res. 1986 Jun 25;14(12):4837–4848. doi: 10.1093/nar/14.12.4837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kearney J. F., Klein J., Bockman D. E., Cooper M. D., Lawton A. R. B cell differentiation induced by lipopolysaccharide. V. Suppression of plasma cell maturation by anti-mu: mode of action and characteristics of suppressed cells. J Immunol. 1978 Jan;120(1):158–166. [PubMed] [Google Scholar]
  15. Kettman J., Wetzel M. Antibody synthesis in vitro, a marker of B cell differentiation. J Immunol Methods. 1980;39(3):203–222. doi: 10.1016/0022-1759(80)90056-3. [DOI] [PubMed] [Google Scholar]
  16. Landolfi N. F., Capra J. D., Tucker P. W. Protein-nucleotide contacts in the immunoglobulin heavy-chain promoter region. Proc Natl Acad Sci U S A. 1987 Jun;84(11):3851–3855. doi: 10.1073/pnas.84.11.3851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Leanderson T., Hsu E. Anti-IgM treatment influences immunoglobulin heavy and light chain mRNA levels in mitogen-stimulated B lymphocytes. Eur J Immunol. 1985 Jun;15(6):641–643. doi: 10.1002/eji.1830150621. [DOI] [PubMed] [Google Scholar]
  18. Mason J. O., Williams G. T., Neuberger M. S. Transcription cell type specificity is conferred by an immunoglobulin VH gene promoter that includes a functional consensus sequence. Cell. 1985 Jun;41(2):479–487. doi: 10.1016/s0092-8674(85)80021-0. [DOI] [PubMed] [Google Scholar]
  19. Parker D. C. Induction and suppression of polyclonal antibody responses by anti-Ig reagents and antigen-nonspecific helper factors: a comparison of the effects of anti-Fab, anti-IgM, and anti IgD on murine B cells. Immunol Rev. 1980;52:115–139. doi: 10.1111/j.1600-065x.1980.tb00333.x. [DOI] [PubMed] [Google Scholar]
  20. Parker D. C. Stimulation of mouse lymphocytes by insoluble anti-mouse immunoglobulin. Nature. 1975 Nov 27;258(5533):361–363. doi: 10.1038/258361a0. [DOI] [PubMed] [Google Scholar]
  21. Parslow T. G., Blair D. L., Murphy W. J., Granner D. K. Structure of the 5' ends of immunoglobulin genes: a novel conserved sequence. Proc Natl Acad Sci U S A. 1984 May;81(9):2650–2654. doi: 10.1073/pnas.81.9.2650. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Phillips N. E., Parker D. C. Fc-dependent inhibition of mouse B cell activation by whole anti-mu antibodies. J Immunol. 1983 Feb;130(2):602–606. [PubMed] [Google Scholar]
  23. Picard D., Schaffner W. Cell-type preference of immunoglobulin kappa and lambda gene promoters. EMBO J. 1985 Nov;4(11):2831–2838. doi: 10.1002/j.1460-2075.1985.tb04011.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Queen C., Baltimore D. Immunoglobulin gene transcription is activated by downstream sequence elements. Cell. 1983 Jul;33(3):741–748. doi: 10.1016/0092-8674(83)90016-8. [DOI] [PubMed] [Google Scholar]
  25. Sasaki K., Murakami T., Ogino T., Takahashi M., Kawasaki S. Flow cytometric estimation of cell cycle parameters using a monoclonal antibody to bromodeoxyuridine. Cytometry. 1986 Jul;7(4):391–395. doi: 10.1002/cyto.990070415. [DOI] [PubMed] [Google Scholar]
  26. Scheidereit C., Heguy A., Roeder R. G. Identification and purification of a human lymphoid-specific octamer-binding protein (OTF-2) that activates transcription of an immunoglobulin promoter in vitro. Cell. 1987 Dec 4;51(5):783–793. doi: 10.1016/0092-8674(87)90101-2. [DOI] [PubMed] [Google Scholar]
  27. Sen R., Baltimore D. Inducibility of kappa immunoglobulin enhancer-binding protein Nf-kappa B by a posttranslational mechanism. Cell. 1986 Dec 26;47(6):921–928. doi: 10.1016/0092-8674(86)90807-x. [DOI] [PubMed] [Google Scholar]
  28. Severson C. D., Burg D. L., Lafrenz D. E., Feldbush T. L. An alternative method of panning for rat B lymphocytes. Immunol Lett. 1987 Aug;15(4):291–295. doi: 10.1016/0165-2478(87)90130-1. [DOI] [PubMed] [Google Scholar]
  29. Sidman C. L., Unanue E. R. Control of B-lymphocyte function. I. Inactivation of mitogenesis by interactions with surface immunoglobulin and Fc-receptor molecules. J Exp Med. 1976 Oct 1;144(4):882–896. doi: 10.1084/jem.144.4.882. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sidman C. L., Unanue E. R. Requirements for mitogenic stimulation of murine B cells by soluble anti-IgM antibodies. J Immunol. 1979 Feb;122(2):406–413. [PubMed] [Google Scholar]
  31. Van Ness B. G., Coleclough C., Perry R. P., Weigert M. DNA between variable and joining gene segments of immunoglobulin kappa light chain is frequently retained in cells that rearrange the kappa locus. Proc Natl Acad Sci U S A. 1982 Jan;79(2):262–266. doi: 10.1073/pnas.79.2.262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Vitetta E., Puré E., Isakson P., Buck L., Uhr J. The activation of murine B cells: the role of surface immunoglobulins. Immunol Rev. 1980;52:211–231. doi: 10.1111/j.1600-065x.1980.tb00336.x. [DOI] [PubMed] [Google Scholar]
  33. Wirth T., Staudt L., Baltimore D. An octamer oligonucleotide upstream of a TATA motif is sufficient for lymphoid-specific promoter activity. Nature. 1987 Sep 10;329(6135):174–178. doi: 10.1038/329174a0. [DOI] [PubMed] [Google Scholar]
  34. Yuan D. Molecular basis for the inhibition of LPS induced differentiation by anti-immunoglobulin. J Mol Cell Immunol. 1987;3(3):133–144. [PubMed] [Google Scholar]
  35. Yuan D., Tucker P. W. Transcriptional regulation of the mu-delta heavy chain locus in normal murine B lymphocytes. J Exp Med. 1984 Aug 1;160(2):564–583. doi: 10.1084/jem.160.2.564. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES