Skip to main content
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
. 1976 Mar;73(3):727–731. doi: 10.1073/pnas.73.3.727

No detectable reiteration of genes coding for mouse MOPC 41 immunoglobulin light-chain mRNA.

M G Farace, M F Aellen, P A Briand, C H Faust, P Vassalli, B Mach
PMCID: PMC335991  PMID: 815907

Abstract

RNA fractions rich in immunoglobulin light (L)-chain mRNA were isolated from mouse myeloma MOPC 41 by procedures previously described, and chemically labeled with 125I. These RNA fractions were hybridized with MOPC 41 DNA under conditions of DNA excess. Hybridization conditions were chosen under which the entire sequence of the L-chain mRNA probe, thus including the variable region, remains available for hybridization throughout the reaction. The hybridization (C0t) curve showed double transition kinetics, with one component corresponding to about 250 gene copies and the other to about two to four copies. In contrast, when MOPC 41 L-chain mRNA was further purified as a single band by gel elecptrophoresis in 99% formamide, the hybridization curve showed only a single transition, corresponding to about two to four genes, with the disappearance of the "reiterated" component. That component resulted therefore from contaminating RNA species. The data indicate that no reiteration can be detected by RNase or by hydroxylapatite for the genes corresponding to the entire sequence of MOPC 41 L-chain mRNA, including the untranslated segments, within the limits of detectability of short reiterated segments. It thus appears that there is only one or very few genes corresponding to the 41 L-chain variable region "subgroup" in MOPC 41 DNA. The possibility that the variable genes of plasmocytes might result frm a combination of several nonreiterated germline genes is discussed.

Full text

PDF
729

Images in this article

Selected References

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

  1. Bernardini A., Tonegawa S. Hybridization studies with an antibody heavy chain mRNA. FEBS Lett. 1974 Apr 15;41(1):73–77. doi: 10.1016/0014-5793(74)80957-9. [DOI] [PubMed] [Google Scholar]
  2. Bishop J. O. Molecular hybridization of ribonucleic acid with a large excess of deoxyribonucleic acid. Biochem J. 1972 Jan;126(1):171–185. doi: 10.1042/bj1260171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Campo M. S., Bishop J. O. Two classes of messenger RNA in cultured rat cells: repetitive sequence transcripts and unique sequence transcripts. J Mol Biol. 1974 Dec 25;90(4):649–663. doi: 10.1016/0022-2836(74)90530-0. [DOI] [PubMed] [Google Scholar]
  4. Delovitch T. L., Baglioni C. Estimation of light-chain gene reiteration of mouse immunoglobulin by DNA-RNA hybridization. Proc Natl Acad Sci U S A. 1973 Jan;70(1):173–178. doi: 10.1073/pnas.70.1.173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dina D., Meza I., Crippa M. Relative positions of the "repetitive", "unique" and poly(A) fragments of mRNA. Nature. 1974 Apr 5;248(448):486–490. doi: 10.1038/248486a0. [DOI] [PubMed] [Google Scholar]
  6. Dube S. K. Use of 125I in fingerprinting RNA. Nature. 1973 Dec 21;246(5434):483–483. doi: 10.1038/246483a0. [DOI] [PubMed] [Google Scholar]
  7. Eichmann K. Idiotype expression and the inheritance of mouse antibody clones. J Exp Med. 1973 Mar 1;137(3):603–621. doi: 10.1084/jem.137.3.603. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Faust C. H., Diggelmann H., Mach B. Estimation of the number of genes coding for the constant part of the mouse immunoglobulin kappa light chain. Proc Natl Acad Sci U S A. 1974 Jun;71(6):2491–2495. doi: 10.1073/pnas.71.6.2491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Faust C. H., Jr, Diggelmann H., Mach B. Isolation of poly(adenylic acid)-rich ribonucleic acid from mouse myeloma and synthesis of complementary deoxyribonucleic acid. Biochemistry. 1973 Feb 27;12(5):925–931. doi: 10.1021/bi00729a021. [DOI] [PubMed] [Google Scholar]
  10. Gally J. A., Edelman G. M. The genetic control of immunoglobulin synthesis. Annu Rev Genet. 1972;6:1–46. doi: 10.1146/annurev.ge.06.120172.000245. [DOI] [PubMed] [Google Scholar]
  11. Gambino R., Kacian D., O'Donnell J., Ramirez F., Marks P. A., Bank A. A limited number of globin genes in human DNA. Proc Natl Acad Sci U S A. 1974 Oct;71(10):3966–3970. doi: 10.1073/pnas.71.10.3966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gould H. J., Hamlyn P. H. The molecular weight of rabbit globin messenger RNA's. FEBS Lett. 1973 Mar 15;30(3):301–304. doi: 10.1016/0014-5793(73)80674-x. [DOI] [PubMed] [Google Scholar]
  13. Harrison P. R., Birnie G. D., Hell A., Humphries S., Young B. D., Paul J. Kinetic studies of gene frequency. I. Use of a DNA copy of reticulocyte 9 S RNA to estimate globin gene dosage in mouse tissues. J Mol Biol. 1974 Apr 25;84(4):539–554. doi: 10.1016/0022-2836(74)90115-6. [DOI] [PubMed] [Google Scholar]
  14. Honjo T., Packman S., Swan D., Nau M., Leder P. Organization of immunoglobulin genes: reiteration frequency of the mouse kappa chain constant region gene. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3659–3663. doi: 10.1073/pnas.71.9.3659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hood L., McKean D., Farnsworth V., Potter M. Mouse immunoglobulin chains. A survey of the amino-terminal sequences of kappa chains. Biochemistry. 1973 Feb;12(4):741–749. doi: 10.1021/bi00728a026. [DOI] [PubMed] [Google Scholar]
  16. Kindt T. J., Seide R. K., Bokisch V. A., Krause R. M. Detection of idiotypic cross-reactions among streptococcal antisera from related rabbits. J Exp Med. 1973 Sep 1;138(3):522–537. doi: 10.1084/jem.138.3.522. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Klein W. H., Murphy W., Attardi G., Britten R. J., Davidson E. H. Distribution of repetitive and nonrepetivite sequence transcripts in HeLa mRNA. Proc Natl Acad Sci U S A. 1974 May;71(5):1785–1789. doi: 10.1073/pnas.71.5.1785. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Knöchel W. Biological activity of 125iodine labelled globin mRNA in an Ehrlich ascites cell-free system. Mol Biol Rep. 1974 Mar;1(6):311–320. doi: 10.1007/BF00309564. [DOI] [PubMed] [Google Scholar]
  19. Leder P., Honjo T., Packman S., Swan D., Nau M., Norman B. The organization and diversity of immunoglobulin genes. Proc Natl Acad Sci U S A. 1974 Dec;71(12):5109–5115. doi: 10.1073/pnas.71.12.5109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mach B., Faust C., Vassalli P. Purification of 14S messenger RNA of immunoglobulin light chain that codes for a possible light-chain precursor. Proc Natl Acad Sci U S A. 1973 Feb;70(2):451–455. doi: 10.1073/pnas.70.2.451. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McCarthy B. J., Church R. B. The specificity of molecular hybridization reactions. Annu Rev Biochem. 1970;39:131–150. doi: 10.1146/annurev.bi.39.070170.001023. [DOI] [PubMed] [Google Scholar]
  22. Milstein C., Brownlee G. G., Cartwright E. M., Jarvis J. M., Proudfoot N. J. Sequence analysis of immunoglobulin light chain messenger RNA. Nature. 1974 Nov 29;252(5482):354–359. doi: 10.1038/252354a0. [DOI] [PubMed] [Google Scholar]
  23. Neiman P. E., Wright S. E., McMillin C., MacDonnell D. Nucleotide sequence relationships of avian RNA tumor viruses: measurement of the deletion in a transformation-defective mutant of Rous sarcoma virus. J Virol. 1974 Apr;13(4):837–846. doi: 10.1128/jvi.13.4.837-846.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Pinder J. C., Staynov D. Z., Gratzer W. B. Electrophoresis of RNA in formamide. Biochemistry. 1974 Dec 17;13(26):5373–5378. doi: 10.1021/bi00723a019. [DOI] [PubMed] [Google Scholar]
  25. Premkumar E., Shoyab M., Williamson A. R. Germ line basis for antibody diversity: immunoglobulin VH-and CH-gene frequencies measured by DNA-RNA hybridization. Proc Natl Acad Sci U S A. 1974 Jan;71(1):99–103. doi: 10.1073/pnas.71.1.99. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Prensky W., Steffensen D. M., Hughes W. L. The use of iodinated RNA for gene localization. Proc Natl Acad Sci U S A. 1973 Jun;70(6):1860–1864. doi: 10.1073/pnas.70.6.1860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Rabbitts T. H., Bishop J. O., Milstein C., Brownlee G. G. Comparative hybridization studies iwth an immunoglobulin light chain mRNA fraction and non-immunoglobulin mRNA of mouse. FEBS Lett. 1974 Mar 15;40(1):157–160. doi: 10.1016/0014-5793(74)80917-8. [DOI] [PubMed] [Google Scholar]
  28. Rabbitts T. H. Hybridization characteristics of enzymatically synthesised DNA complementary to mouse immunoglobulin messenger RNA. FEBS Lett. 1974 Jun 15;42(3):323–326. doi: 10.1016/0014-5793(74)80756-8. [DOI] [PubMed] [Google Scholar]
  29. Rabbitts T. H., Jarvis J. M., Milstein C. Demonstration that a mouse immunoglobulin light chain messenger RNA hybridizes exclusively with unique DNA. Cell. 1975 Sep;6(1):5–12. doi: 10.1016/0092-8674(75)90067-7. [DOI] [PubMed] [Google Scholar]
  30. Rabbitts T. H., Milstein C. Mouse immunoglobulin genes: studies on the reiteration frequency of light-chain genes by hybridisation procedures. Eur J Biochem. 1975 Mar 3;52(1):125–133. doi: 10.1111/j.1432-1033.1975.tb03980.x. [DOI] [PubMed] [Google Scholar]
  31. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. STUDIER F. W. SEDIMENTATION STUDIES OF THE SIZE AND SHAPE OF DNA. J Mol Biol. 1965 Feb;11:373–390. doi: 10.1016/s0022-2836(65)80064-x. [DOI] [PubMed] [Google Scholar]
  33. Stavnezer J., Huang R. C., Stavnezer E., Bishop J. M. Isolation of messenger RNA for an immunoglobulin kappa chain and enumeration of the genes for the constatn region of kappa chain in the mouse. J Mol Biol. 1974 Sep 5;88(1):43–63. doi: 10.1016/0022-2836(74)90294-0. [DOI] [PubMed] [Google Scholar]
  34. Storb U. Evidence for multiple immunoglobulin genes. Biochem Biophys Res Commun. 1974 Mar 15;57(1):31–38. doi: 10.1016/s0006-291x(74)80353-0. [DOI] [PubMed] [Google Scholar]
  35. Straus N. A., Bonner T. I. Temperature dependence of RNA-DNA hybridization kinetics. Biochim Biophys Acta. 1972 Aug 16;277(1):87–95. doi: 10.1016/0005-2787(72)90355-3. [DOI] [PubMed] [Google Scholar]
  36. Tereba A., McCarthy B. J. Hybridization of 125I-labeled ribonucleic acid. Biochemistry. 1973 Nov 6;12(23):4675–4679. doi: 10.1021/bi00747a020. [DOI] [PubMed] [Google Scholar]
  37. Tonegawa S., Steinberg C., Dube S., Bernardini A. Evidence for somatic generation of antibody diversity. Proc Natl Acad Sci U S A. 1974 Oct;71(10):4027–4031. doi: 10.1073/pnas.71.10.4027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Wu T. T., Kabat E. A. An analysis of the sequences of the variable regions of Bence Jones proteins and myeloma light chains and their implications for antibody complementarity. J Exp Med. 1970 Aug 1;132(2):211–250. doi: 10.1084/jem.132.2.211. [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