Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1983 Aug;80(16):4997–5001. doi: 10.1073/pnas.80.16.4997

Recombination between antibody heavy chain variable-region genes: evidence for gene conversion.

U Krawinkel, G Zoebelein, M Brüggemann, A Radbruch, K Rajewsky
PMCID: PMC384175  PMID: 6308665

Abstract

The murine hybridoma line B1-8.delta 1 secretes monoclonal IgD lambda 1 antibodies specific for the hapten (4-hydroxy-3-nitrophenyl)acetyl (NP). The variable (V) region of these antibodies is defined by a characteristic pattern of idiotopes. A spontaneous V-region variant (B1-8.V1) with altered idiotope pattern was selected. The structural variation is confined to the V region of the heavy chain. It was shown previously that the variant V region is encoded by a gene that was generated by a crossover between the rearranged VDJ gene of the wild type (B1-8.delta 1) and a neighboring germ-line VH gene. In the present study the nucleotide sequence of coding and flanking regions of the VH gene expressed in variant B1-8.V1 was determined. Wild-type and variant VH genes differ at 15 positions in a region between leader sequence and codon 66. The sequence of the region carrying the substitutions is identical to the sequence of the corresponding region in a neighboring germ-line VH gene. This implies that the variant VH gene was generated by a mechanism of recombination more complicated than single crossover. Gene conversion as the mechanism of the recombination is discussed.

Full text

PDF
4997

Images in this article

4998Image
on p.4998

Selected References

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

  1. Alt F., Rosenberg N., Lewis S., Thomas E., Baltimore D. Organization and reorganization of immunoglobulin genes in A-MULV-transformed cells: rearrangement of heavy but not light chain genes. Cell. 1981 Dec;27(2 Pt 1):381–390. doi: 10.1016/0092-8674(81)90421-9. [DOI] [PubMed] [Google Scholar]
  2. Baltimore D. Gene conversion: some implications for immunoglobulin genes. Cell. 1981 Jun;24(3):592–594. doi: 10.1016/0092-8674(81)90082-9. [DOI] [PubMed] [Google Scholar]
  3. Bentley D. L., Rabbitts T. H. Evolution of immunoglobulin V genes: evidence indicating that recently duplicated human V kappa sequences have diverged by gene conversion. Cell. 1983 Jan;32(1):181–189. doi: 10.1016/0092-8674(83)90508-1. [DOI] [PubMed] [Google Scholar]
  4. Bentley D. L., Rabbitts T. H. Human V kappa immunoglobulin gene number: implications for the origin of antibody diversity. Cell. 1981 Jun;24(3):613–623. doi: 10.1016/0092-8674(81)90088-x. [DOI] [PubMed] [Google Scholar]
  5. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  6. Bernard O., Hozumi N., Tonegawa S. Sequences of mouse immunoglobulin light chain genes before and after somatic changes. Cell. 1978 Dec;15(4):1133–1144. doi: 10.1016/0092-8674(78)90041-7. [DOI] [PubMed] [Google Scholar]
  7. Bothwell A. L., Paskind M., Reth M., Imanishi-Kari T., Rajewsky K., Baltimore D. Heavy chain variable region contribution to the NPb family of antibodies: somatic mutation evident in a gamma 2a variable region. Cell. 1981 Jun;24(3):625–637. doi: 10.1016/0092-8674(81)90089-1. [DOI] [PubMed] [Google Scholar]
  8. Brack C., Hirama M., Lenhard-Schuller R., Tonegawa S. A complete immunoglobulin gene is created by somatic recombination. Cell. 1978 Sep;15(1):1–14. doi: 10.1016/0092-8674(78)90078-8. [DOI] [PubMed] [Google Scholar]
  9. Brüggemann M., Radbruch A., Rajewsky K. Immunoglobulin V region variants in hybridoma cells. I. Isolation of a variant with altered idiotypic and antigen binding specificity. EMBO J. 1982;1(5):629–634. doi: 10.1002/j.1460-2075.1982.tb01219.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Clarke S. H., Claflin J. L., Rudikoff S. Polymorphism in immunoglobulin heavy chains suggesting gene conversion. Proc Natl Acad Sci U S A. 1982 May;79(10):3280–3284. doi: 10.1073/pnas.79.10.3280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cohen J. B., Effron K., Rechavi G., Ben-Neriah Y., Zakut R., Givol D. Simple DNA sequences in homologous flanking regions near immunoglobulin VH genes: a role in gene interaction? Nucleic Acids Res. 1982 Jun 11;10(11):3353–3370. doi: 10.1093/nar/10.11.3353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Crews S., Griffin J., Huang H., Calame K., Hood L. A single VH gene segment encodes the immune response to phosphorylcholine: somatic mutation is correlated with the class of the antibody. Cell. 1981 Jul;25(1):59–66. doi: 10.1016/0092-8674(81)90231-2. [DOI] [PubMed] [Google Scholar]
  13. Denhardt D. T. A membrane-filter technique for the detection of complementary DNA. Biochem Biophys Res Commun. 1966 Jun 13;23(5):641–646. doi: 10.1016/0006-291x(66)90447-5. [DOI] [PubMed] [Google Scholar]
  14. Dildrop R., Brüggemann M., Radbruch A., Rajewsky K., Beyreuther K. Immunoglobulin V region variants in hybridoma cells. II. Recombination between V genes. EMBO J. 1982;1(5):635–640. doi: 10.1002/j.1460-2075.1982.tb01220.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Early P., Huang H., Davis M., Calame K., Hood L. An immunoglobulin heavy chain variable region gene is generated from three segments of DNA: VH, D and JH. Cell. 1980 Apr;19(4):981–992. doi: 10.1016/0092-8674(80)90089-6. [DOI] [PubMed] [Google Scholar]
  16. Egel R. Intergenic conversion and reiterated genes. Nature. 1981 Mar 19;290(5803):191–192. doi: 10.1038/290191a0. [DOI] [PubMed] [Google Scholar]
  17. Ernst J. F., Stewart J. W., Sherman F. The cyc1-11 mutation in yeast reverts by recombination with a nonallelic gene: composite genes determining the iso-cytochromes c. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6334–6338. doi: 10.1073/pnas.78.10.6334. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Evans G. A., Margulies D. H., Camerini-Otero R. D., Ozato K., Seidman J. G. Structure and expression of a mouse major histocompatibility antigen gene, H-2Ld. Proc Natl Acad Sci U S A. 1982 Mar;79(6):1994–1998. doi: 10.1073/pnas.79.6.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gearhart P. J., Johnson N. D., Douglas R., Hood L. IgG antibodies to phosphorylcholine exhibit more diversity than their IgM counterparts. Nature. 1981 May 7;291(5810):29–34. doi: 10.1038/291029a0. [DOI] [PubMed] [Google Scholar]
  20. Gough N. M., Bernard O. Sequences of the joining region genes for immunoglobulin heavy chains and their role in generation of antibody diversity. Proc Natl Acad Sci U S A. 1981 Jan;78(1):509–513. doi: 10.1073/pnas.78.1.509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hohn B., Murray K. Packaging recombinant DNA molecules into bacteriophage particles in vitro. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3259–3263. doi: 10.1073/pnas.74.8.3259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jackson J. A., Fink G. R. Gene conversion between duplicated genetic elements in yeast. Nature. 1981 Jul 23;292(5821):306–311. doi: 10.1038/292306a0. [DOI] [PubMed] [Google Scholar]
  23. Kabat E. A., Wu T. T., Bilofsky H. Evidence supporting somatic assembly of the DNA segments (minigenes), coding for the framework, and complementarity-determining segments of immunoglobulin variable regions. J Exp Med. 1979 Jun 1;149(6):1299–1313. doi: 10.1084/jem.149.6.1299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kabat E. A., Wu T. T., Bilofsky H. Variable region genes for the immunoglobulin framework are assembled from small segments of DNA--a hypothesis. Proc Natl Acad Sci U S A. 1978 May;75(5):2429–2433. doi: 10.1073/pnas.75.5.2429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Klar A. J., Fogel S. Activation of mating type genes by transposition in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4539–4543. doi: 10.1073/pnas.76.9.4539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kurosawa Y., Tonegawa S. Organization, structure, and assembly of immunoglobulin heavy chain diversity DNA segments. J Exp Med. 1982 Jan 1;155(1):201–218. doi: 10.1084/jem.155.1.201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Laskey R. A., Mills A. D. Enhanced autoradiographic detection of 32P and 125I using intensifying screens and hypersensitized film. FEBS Lett. 1977 Oct 15;82(2):314–316. doi: 10.1016/0014-5793(77)80609-1. [DOI] [PubMed] [Google Scholar]
  28. Loh D. Y., Bothwell A. L., White-Scharf M. E., Imanishi-Kari T., Baltimore D. Molecular basis of a mouse strain-specific anti-hapten response. Cell. 1983 May;33(1):85–93. doi: 10.1016/0092-8674(83)90337-9. [DOI] [PubMed] [Google Scholar]
  29. Max E. E., Seidman J. G., Leder P. Sequences of five potential recombination sites encoded close to an immunoglobulin kappa constant region gene. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3450–3454. doi: 10.1073/pnas.76.7.3450. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Meselson M. S., Radding C. M. A general model for genetic recombination. Proc Natl Acad Sci U S A. 1975 Jan;72(1):358–361. doi: 10.1073/pnas.72.1.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Neuberger M. S., Rajewsky K. Switch from hapten-specific immunoglobulin M to immunoglobulin D secretion in a hybrid mouse cell line. Proc Natl Acad Sci U S A. 1981 Feb;78(2):1138–1142. doi: 10.1073/pnas.78.2.1138. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Ollo R., Rougeon F. Gene conversion and polymorphism: generation of mouse immunoglobulin gamma 2a chain alleles by differential gene conversion by gamma 2b chain gene. Cell. 1983 Feb;32(2):515–523. doi: 10.1016/0092-8674(83)90471-3. [DOI] [PubMed] [Google Scholar]
  33. Pech M., Höchtl J., Schnell H., Zachau H. G. Differences between germ-line and rearranged immunoglobulin V kappa coding sequences suggest a localized mutation mechanism. Nature. 1981 Jun 25;291(5817):668–670. doi: 10.1038/291668a0. [DOI] [PubMed] [Google Scholar]
  34. Reth M., Hämmerling G. J., Rajewsky K. Analysis of the repertoire of anti-NP antibodies in C57BL/6 mice by cell fusion. I. Characterization of antibody families in the primary and hyperimmune response. Eur J Immunol. 1978 Jun;8(6):393–400. doi: 10.1002/eji.1830080605. [DOI] [PubMed] [Google Scholar]
  35. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  36. Rimm D. L., Horness D., Kucera J., Blattner F. R. Construction of coliphage lambda Charon vectors with BamHI cloning sites. Gene. 1980 Dec;12(3-4):301–309. doi: 10.1016/0378-1119(80)90113-4. [DOI] [PubMed] [Google Scholar]
  37. Sablitzky F., Radbruch A., Rajewsky K. Spontaneous immunoglobulin class switching in myeloma and hybridoma cell lines differs from physiological class switching. Immunol Rev. 1982;67:59–72. doi: 10.1111/j.1600-065x.1982.tb01055.x. [DOI] [PubMed] [Google Scholar]
  38. Sakano H., Hüppi K., Heinrich G., Tonegawa S. Sequences at the somatic recombination sites of immunoglobulin light-chain genes. Nature. 1979 Jul 26;280(5720):288–294. doi: 10.1038/280288a0. [DOI] [PubMed] [Google Scholar]
  39. Sanger F., Coulson A. R., Barrell B. G., Smith A. J., Roe B. A. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. doi: 10.1016/0022-2836(80)90196-5. [DOI] [PubMed] [Google Scholar]
  40. Schreier P. H., Bothwell A. L., Mueller-Hill B., Baltimore D. Multiple differences between the nucleic acid sequences of the IgG2aa and IgG2ab alleles of the mouse. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4495–4499. doi: 10.1073/pnas.78.7.4495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schreier P. H., Cortese R. A fast and simple method for sequencing DNA cloned in the single-stranded bacteriophage M13. J Mol Biol. 1979 Mar 25;129(1):169–172. doi: 10.1016/0022-2836(79)90068-8. [DOI] [PubMed] [Google Scholar]
  42. Seidman J. G., Leder A., Nau M., Norman B., Leder P. Antibody diversity. Science. 1978 Oct 6;202(4363):11–17. doi: 10.1126/science.99815. [DOI] [PubMed] [Google Scholar]
  43. Slightom J. L., Blechl A. E., Smithies O. Human fetal G gamma- and A gamma-globin genes: complete nucleotide sequences suggest that DNA can be exchanged between these duplicated genes. Cell. 1980 Oct;21(3):627–638. doi: 10.1016/0092-8674(80)90426-2. [DOI] [PubMed] [Google Scholar]
  44. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  45. Steffen D., Weinberg R. A. The integrated genome of murine leukemia virus. Cell. 1978 Nov;15(3):1003–1010. doi: 10.1016/0092-8674(78)90284-2. [DOI] [PubMed] [Google Scholar]
  46. Szostak J. W., Orr-Weaver T. L., Rothstein R. J., Stahl F. W. The double-strand-break repair model for recombination. Cell. 1983 May;33(1):25–35. doi: 10.1016/0092-8674(83)90331-8. [DOI] [PubMed] [Google Scholar]
  47. Weigert M., Riblet R. Genetic control of antibody variable regions. Cold Spring Harb Symp Quant Biol. 1977;41(Pt 2):837–846. doi: 10.1101/sqb.1977.041.01.093. [DOI] [PubMed] [Google Scholar]
  48. Wu T. T., Kabat E. A. Fourteen nucleotides in the second complementarity-determining region of a human heavy-chain variable region gene are identical with a sequence in a human D minigene. Proc Natl Acad Sci U S A. 1982 Aug;79(16):5031–5032. doi: 10.1073/pnas.79.16.5031. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES