In situ Detection of Stage-Specific Genes and Enhancers in B Cell Differentiation via Gene-Search Retroviruses

  • William G. Kerr
  • Garry P. Nolan
  • Jeffrey B. Johnsen
  • Leonard A. Herzenberg
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 292)


We demonstrate that infection of an LPS-responsive pre-B cell line with transcription-ally-defective retroviruses containing a reporter gene (lacZ) can result in viral integrations where expression of lacZ is differentiation stage-dependent. Because expression of lacZ is dependent upon flanking cellular sequences these retroviral integrations represent in situ gene fusions with cellular enhancers (Enhsrl) and genes (Gensrl) which are either induced or repressed during LPS-stimulated differentiation. One of the well-documented effects of LPS upon pre-B cells is the induction of κ light chain transcription via NF-κB. The identification of LPS-stimulated gene repression during B cell differentiation indicates that LPS has multiple effects upon gene expression during the pre-B to B cell transition. The identification of cellular enhancers and genes which are downregulated during the transition from the pre-B to the B cell stage indicates that other transcription factors, in addition to NF-ACB, are required for this step in differentiation. Finally, we present some initial experiments which indicate the gene-search retroviruses can introduce expression of lacZ into normal hematopoietic cells in vitro and in vivo.


Cellular Gene lacZ Expression Fetal Liver Cell Light Chain Expression Enhancer Detection 
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  1. 1.
    M. D. Cooper and P. D. Burrows, B-cell differentiation, in: “Immunoglobulin Genes”, T. Honjo, F. Alt, and T. Rabbitts, eds., Academic Press Limited, London (1989).Google Scholar
  2. 2.
    W. G. Kerr, M. D. Cooper, L. Feng, P. D. Burrows, and L. M. Hendershot, Mu heavy chains can associate with a pseudo-light chain complex (L) in human pre-B cell lines, Internat. Immunol. 1:355 (1989).CrossRefGoogle Scholar
  3. 3.
    M. E. Koshland, The immunoglobulin helper: the J chain, in “Immunoglobulin Genes”,T. Honjo, F. Alt, and T. Rabbitts, eds., Academic Press Limited, London (1989).Google Scholar
  4. 4.
    W. G. Kerr, G. P. Nolan, and L. A. Herzenberg, In situ detection of transcriptionally-active chromatin and genetic regulatory elements in individual viable mammalian cells, Immunology 68 (Suppl. 2):74 (1989).PubMedGoogle Scholar
  5. 5.
    W. G. Kerr, G. P. Nolan, A. T. Serafini, and L. A. Herzenberg, Transcriptionally-defective retroviruses containing lacZ for the in situ detection of endogenous genes and develapmentally-regulated chromatin, CSHSQB 54:767 (1989).Google Scholar
  6. 6.
    N. D. Allen, D. G. Can, S. C. Barton, S. Hettle, and M. A. Surami, Transgenes as probes for active chromosomal domains in mouse development, Nature 333:852 (1988).PubMedCrossRefGoogle Scholar
  7. 7.
    A. Gossler, A. L. Joyner, J. Rossant, and W. C. Skarnes, Mouse embryonic stem cells and reporter constructs to detect developmentally regulated genes, Science 244:463 (1989).PubMedCrossRefGoogle Scholar
  8. 8.
    C. J. O’Kane and W. J. Gehring, Detection in situ of genomic regulatory elements in Drosophilia, Proc. Natl. Acad. Sci. USA 84:9123 (1987).PubMedCrossRefGoogle Scholar
  9. 9.
    H. J. Bellen, C. J. O’Kane, C. Wilson, U. Grossniklaus, R. K. Pearson, and W. J. Gehring, P-element-mediated enhancer detection: a versatile method to study development in Drosophilia, Genes and Devel. 3:1288 (1989).CrossRefGoogle Scholar
  10. 10.
    C. Wilson, R. K. Pearson, H. J. Bellen, C. J. O’Kane, U. Grossniklaus, and W. J. Gehring, P-element-mediated enhancer detection: an efficient method for isolating and characterizing developmentally regulated genes in Drosophilia, Genes and Devel. 3:1301 (1989).CrossRefGoogle Scholar
  11. 11.
    S. F. Yu, T. von Ruden, P. W. Kantoff, C. Garber, M. Sieberg, U. Ruther, W. F. Anderson, E. F. Wagner, and E. Gilboa, Self-inactivating retroviral vectors designed for transfer of whole genes into mammalian cells, Proc. Natl. Acad. Sci. USA 83:3194 (1986).PubMedCrossRefGoogle Scholar
  12. 12.
    H. E. Varmus, Form and function of retroviral proviruses, Science 216:812 (1982).PubMedCrossRefGoogle Scholar
  13. 13.
    R. Mann and D. Baltimore, Varying the position of a retrovirus packaging sequence results in the encapsidation of both unspliced and spliced RNAs, J. Virol. 54:401 (1985).PubMedGoogle Scholar
  14. 14.
    P. A. Lazo, V. Prasad, and P. N. Tsichlis, Splice acceptor site for the env message of Moloney murine leukemia virus, J. Virol. 61:2038 (1987).PubMedGoogle Scholar
  15. 15.
    C. C. Shih, J. P. Stoye, and J. M. Coffin, Highly preferred targets for retrovirus integration, Cell 53:531 (1988).PubMedCrossRefGoogle Scholar
  16. 16.
    A. Winoto and D. Baltimore, a lineage-specific expression of the a T cell receptor gene by nearby silencers, Cell 59:649 (1989).PubMedCrossRefGoogle Scholar
  17. 17.
    F. Melchers, The many roles of immunoglobulin molecules and growth control of the B-lymphocyte lineage, in “Immunoglobulin Genes”, T. Honjo, F. Alt, and T. Rabbitts, eds., Academic Press Limited, London (1989).Google Scholar
  18. 18.
    A. N. Shakhov, M. A. Collart, P. Vassali, S. A. Nedospasov, and C. V. Jongeneel, /cB-type enhancers are involved in lipopolysaccharide-mediated transcriptional activation of the tumor necrosis factor a gene in primary macrophages, J. Exp. Med. 171:35 (1990).PubMedCrossRefGoogle Scholar
  19. 19.
    R. Sen and D. Baltimore, Multiple nuclear factors interact with the immunoglobulin enhancer sequences, Cell 46:705 (1986).PubMedCrossRefGoogle Scholar
  20. 20.
    R. Sen and D. Baltimore, Inducibility of k ,immunoglobulin enhancer-binding protein NF-k. by a post-translational mechanism, Cell 47:921 (1986).PubMedCrossRefGoogle Scholar
  21. 21.
    R. R. Hardy, K. Hayakawa, D. R. Parks, L. A. Herzenberg, and L. A. Herzenberg, Murine B cell differentiation lineages, J. Exp. Med. 159:1169 (1984).PubMedCrossRefGoogle Scholar
  22. 22.
    R. L. Coffman and I. L. Weissman, B220, a B cell specific member of the T200 glycoprotein, Nature 289:681 (1981).PubMedCrossRefGoogle Scholar
  23. 23.
    M. Briskin, M. Kuwabara, D. Sigman, and R. Wall, Induction of k transcription by interferon-without activation of NF-kB, Science 242:1036 (1988).PubMedCrossRefGoogle Scholar
  24. 24.
    G. Lee, L. R. Ellingsworth, S. Gillis, R. Wall, and P. W. Kincade, transforming growth factors are potential regulators of B lymphocytes, J. Exp. Med. 166:1290 (1987).PubMedCrossRefGoogle Scholar
  25. 25.
    T. Honjo, A. Shimuzu, and Y. Yaoita, Constant-region genes of the immunoglobulin heavy chain and the molecular mechanism of heavy chain class switching, in “Immunoglobulin Genes”, T. Honjo, F. Alt, and T. Rabbitts, eds., Academic Press Limited, London (1989).Google Scholar
  26. 26.
    M. R. Lieber, J. E. Hesse, K. Mizuuchi, and M. Gellert, Developmental stage specificity of the lymphoid V(D)J recombination activity, Genes and Devel. 1:451 (1987).Google Scholar
  27. 27.
    S. Desiderio, G. Yancopolous, M. Paskind, E. Thomas, M. Boss, N. Landau, F. Alt, and D. Baltimore, Insertion of N-regions into heavy-chain genes is correlated with expression of terminal deoxynucleotide transferase in B cells, Nature 311:752 (1984).PubMedCrossRefGoogle Scholar
  28. 28.
    D. G. Schatz, M. A. Oettinger, and D. Baltimore, the V(D)J recombination activating gene, RAG-1, Cell 59:1035 (1989).Google Scholar
  29. 29.
    J. Hombach, T. Tsubata, L. Leclerq, H. Stappert, and M. Reth, Molecular components of the B-cell antigen receptor complex of the IgM class, Nature 343:760 (1990).PubMedCrossRefGoogle Scholar
  30. 30.
    J. Wienands, J. Hombach, A. Radbruch, C. Riesterer, and M. Reth, Molecular components of the B cell antigen receptor complex of IgD differ partly from those of IgM, EMBO 9:449 (1990).Google Scholar
  31. 31.
    J. Chen, A. M. Stall, L. A. Herzenberg, and L. A. Herzenberg, Differences in glycoproteins complexes associated with IgM and IgD on normal murine B cells potentially enable transduction of different signals (1990, submitted).Google Scholar
  32. 32.
    G. P. Nolan, S. Fiering, J. F. Nicolas, and L. A. Herzenberg, Fluorescence-activated cell analysis and sorting of viable mammalian cells based on ;-D-galactosidase activity after transduction of Escherichia coli lacZ, Proc. Natl. Acad. Sci. USA 85:2603 (1988).PubMedCrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1991

Authors and Affiliations

  • William G. Kerr
    • 1
  • Garry P. Nolan
    • 1
  • Jeffrey B. Johnsen
    • 1
  • Leonard A. Herzenberg
    • 1
  1. 1.Department of GeneticsStanford UniversityStanfordUSA

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