Skip to main content
NCBI home page
The Journal of Experimental Medicine logoLink to The Journal of Experimental Medicine
. 1993 Apr 1;177(4):915–923. doi: 10.1084/jem.177.4.915

Distinctive growth requirements and gene expression patterns distinguish progenitor B cells from pre-B cells

PMCID: PMC2190993  PMID: 7681470

Abstract

Long-term bone marrow cultures have been useful in determining gene expression patterns in pre-B cells and in the identification of cytokines such as interleukin 7 (IL-7). We have developed a culture system to selectively grow populations of B lineage restricted progenitors (pro-B cells) from murine bone marrow. Pro-B cells do not grow in response to IL-7, Steel locus factor (SLF), or a combination of the two. c-kit, the SLF receptor, and the IL-7 receptor are both expressed by pro-B cells, indicating that the lack of response is not simply due to the absence of receptors. Furthermore, SLF is not necessary for the growth of pro-B cells since they could be expanded on a stromal line derived from Steel mice that produces no SLF. IL-7 responsiveness in pre-B cells is associated with an increase in n-myc expression and is correlated with immunoglobulin (Ig) gene rearrangements. Although members of the ets family of transcription factors and the Pim-1 kinase are expressed by pro-B cells, n-myc is not expressed. Pro-B cells maintain Ig genes in the germline configuration, which is correlated with a low level of recombination activating genes 1 and 2 (Rag-1 and 2) mRNA expression, but high expression of sterile mu and terminal deoxynucleotidyl transferase. Pro-B cells are unable to grow separated from the stromal layer by a porous membrane, indicating that stromal contact is required for growth. These results suggest that pro-B cells are dependent on alternative growth signals derived from bone marrow stroma and can be distinguished from pre-B cells by specific patterns of gene expression.

Full Text

The Full Text of this article is available as a PDF (1.2 MB).

Selected References

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

  1. Alt F. W., Blackwell T. K., Yancopoulos G. D. Development of the primary antibody repertoire. Science. 1987 Nov 20;238(4830):1079–1087. doi: 10.1126/science.3317825. [DOI] [PubMed] [Google Scholar]
  2. 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]
  3. Anderson D. M., Williams D. E., Tushinski R., Gimpel S., Eisenman J., Cannizzaro L. A., Aronson M., Croce C. M., Huebner K., Cosman D. Alternate splicing of mRNAs encoding human mast cell growth factor and localization of the gene to chromosome 12q22-q24. Cell Growth Differ. 1991 Aug;2(8):373–378. [PubMed] [Google Scholar]
  4. Bauer S. R., Kudo A., Melchers F. Structure and pre-B lymphocyte restricted expression of the VpreB in humans and conservation of its structure in other mammalian species. EMBO J. 1988 Jan;7(1):111–116. doi: 10.1002/j.1460-2075.1988.tb02789.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ben-David Y., Giddens E. B., Letwin K., Bernstein A. Erythroleukemia induction by Friend murine leukemia virus: insertional activation of a new member of the ets gene family, Fli-1, closely linked to c-ets-1. Genes Dev. 1991 Jun;5(6):908–918. doi: 10.1101/gad.5.6.908. [DOI] [PubMed] [Google Scholar]
  6. Billips L. G., Petitte D., Dorshkind K., Narayanan R., Chiu C. P., Landreth K. S. Differential roles of stromal cells, interleukin-7, and kit-ligand in the regulation of B lymphopoiesis. Blood. 1992 Mar 1;79(5):1185–1192. [PubMed] [Google Scholar]
  7. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  8. Collins L. S., Dorshkind K. A stromal cell line from myeloid long-term bone marrow cultures can support myelopoiesis and B lymphopoiesis. J Immunol. 1987 Feb 15;138(4):1082–1087. [PubMed] [Google Scholar]
  9. Cumano A., Paige C. J., Iscove N. N., Brady G. Bipotential precursors of B cells and macrophages in murine fetal liver. Nature. 1992 Apr 16;356(6370):612–615. doi: 10.1038/356612a0. [DOI] [PubMed] [Google Scholar]
  10. Davidson W. F., Fredrickson T. N., Rudikoff E. K., Coffman R. L., Hartley J. W., Morse H. C., 3rd A unique series of lymphomas related to the Ly-1+ lineage of B lymphocyte differentiation. J Immunol. 1984 Aug;133(2):744–753. [PubMed] [Google Scholar]
  11. 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]
  12. Goodwin R. G., Friend D., Ziegler S. F., Jerzy R., Falk B. A., Gimpel S., Cosman D., Dower S. K., March C. J., Namen A. E. Cloning of the human and murine interleukin-7 receptors: demonstration of a soluble form and homology to a new receptor superfamily. Cell. 1990 Mar 23;60(6):941–951. doi: 10.1016/0092-8674(90)90342-c. [DOI] [PubMed] [Google Scholar]
  13. Gunther C. V., Nye J. A., Bryner R. S., Graves B. J. Sequence-specific DNA binding of the proto-oncoprotein ets-1 defines a transcriptional activator sequence within the long terminal repeat of the Moloney murine sarcoma virus. Genes Dev. 1990 Apr;4(4):667–679. doi: 10.1101/gad.4.4.667. [DOI] [PubMed] [Google Scholar]
  14. Hara K., Suda T., Suda J., Eguchi M., Ihle J. N., Nagata S., Miura Y., Saito M. Bipotential murine hemopoietic cell line (NFS-60) that is responsive to IL-3, GM-CSF, G-CSF, and erythropoietin. Exp Hematol. 1988 May;16(4):256–261. [PubMed] [Google Scholar]
  15. Hardy R. R., Carmack C. E., Shinton S. A., Kemp J. D., Hayakawa K. Resolution and characterization of pro-B and pre-pro-B cell stages in normal mouse bone marrow. J Exp Med. 1991 May 1;173(5):1213–1225. doi: 10.1084/jem.173.5.1213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hayashi S., Kunisada T., Ogawa M., Sudo T., Kodama H., Suda T., Nishikawa S., Nishikawa S. Stepwise progression of B lineage differentiation supported by interleukin 7 and other stromal cell molecules. J Exp Med. 1990 May 1;171(5):1683–1695. doi: 10.1084/jem.171.5.1683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hermanson G. G., Eisenberg D., Kincade P. W., Wall R. B29: a member of the immunoglobulin gene superfamily exclusively expressed on beta-lineage cells. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6890–6894. doi: 10.1073/pnas.85.18.6890. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hombach J., Tsubata T., Leclercq L., Stappert H., Reth M. Molecular components of the B-cell antigen receptor complex of the IgM class. Nature. 1990 Feb 22;343(6260):760–762. doi: 10.1038/343760a0. [DOI] [PubMed] [Google Scholar]
  19. Hunt P., Robertson D., Weiss D., Rennick D., Lee F., Witte O. N. A single bone marrow-derived stromal cell type supports the in vitro growth of early lymphoid and myeloid cells. Cell. 1987 Mar 27;48(6):997–1007. doi: 10.1016/0092-8674(87)90708-2. [DOI] [PubMed] [Google Scholar]
  20. Ikuta K., Weissman I. L. Evidence that hematopoietic stem cells express mouse c-kit but do not depend on steel factor for their generation. Proc Natl Acad Sci U S A. 1992 Feb 15;89(4):1502–1506. doi: 10.1073/pnas.89.4.1502. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kierney P. C., Dorshkind K. B lymphocyte precursors and myeloid progenitors survive in diffusion chamber cultures but B cell differentiation requires close association with stromal cells. Blood. 1987 Nov;70(5):1418–1424. [PubMed] [Google Scholar]
  22. Klemsz M. J., McKercher S. R., Celada A., Van Beveren C., Maki R. A. The macrophage and B cell-specific transcription factor PU.1 is related to the ets oncogene. Cell. 1990 Apr 6;61(1):113–124. doi: 10.1016/0092-8674(90)90219-5. [DOI] [PubMed] [Google Scholar]
  23. Landau N. R., St John T. P., Weissman I. L., Wolf S. C., Silverstone A. E., Baltimore D. Cloning of terminal transferase cDNA by antibody screening. Proc Natl Acad Sci U S A. 1984 Sep;81(18):5836–5840. doi: 10.1073/pnas.81.18.5836. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Landreth K. S., Narayanan R., Dorshkind K. Insulin-like growth factor-I regulates pro-B cell differentiation. Blood. 1992 Sep 1;80(5):1207–1212. [PubMed] [Google Scholar]
  25. Lee G., Namen A. E., Gillis S., Ellingsworth L. R., Kincade P. W. Normal B cell precursors responsive to recombinant murine IL-7 and inhibition of IL-7 activity by transforming growth factor-beta. J Immunol. 1989 Jun 1;142(11):3875–3883. [PubMed] [Google Scholar]
  26. Lüscher B., Eisenman R. N. New light on Myc and Myb. Part I. Myc. Genes Dev. 1990 Dec;4(12A):2025–2035. doi: 10.1101/gad.4.12a.2025. [DOI] [PubMed] [Google Scholar]
  27. Matsui Y., Toksoz D., Nishikawa S., Nishikawa S., Williams D., Zsebo K., Hogan B. L. Effect of Steel factor and leukaemia inhibitory factor on murine primordial germ cells in culture. Nature. 1991 Oct 24;353(6346):750–752. doi: 10.1038/353750a0. [DOI] [PubMed] [Google Scholar]
  28. McKearn J. P., McCubrey J., Fagg B. Enrichment of hematopoietic precursor cells and cloning of multipotential B-lymphocyte precursors. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7414–7418. doi: 10.1073/pnas.82.21.7414. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. McNiece I. K., Langley K. E., Zsebo K. M. The role of recombinant stem cell factor in early B cell development. Synergistic interaction with IL-7. J Immunol. 1991 Jun 1;146(11):3785–3790. [PubMed] [Google Scholar]
  30. Meeker T. C., Loeb J., Ayres M., Sellers W. The human Pim-1 gene is selectively transcribed in different hemato-lymphoid cell lines in spite of a G + C-rich housekeeping promoter. Mol Cell Biol. 1990 Apr;10(4):1680–1688. doi: 10.1128/mcb.10.4.1680. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Morrow M. A., Lee G., Gillis S., Yancopoulos G. D., Alt F. W. Interleukin-7 induces N-myc and c-myc expression in normal precursor B lymphocytes. Genes Dev. 1992 Jan;6(1):61–70. doi: 10.1101/gad.6.1.61. [DOI] [PubMed] [Google Scholar]
  32. Namen A. E., Lupton S., Hjerrild K., Wignall J., Mochizuki D. Y., Schmierer A., Mosley B., March C. J., Urdal D., Gillis S. Stimulation of B-cell progenitors by cloned murine interleukin-7. Nature. 1988 Jun 9;333(6173):571–573. doi: 10.1038/333571a0. [DOI] [PubMed] [Google Scholar]
  33. Nishikawa S., Ogawa M., Nishikawa S., Kunisada T., Kodama H. B lymphopoiesis on stromal cell clone: stromal cell clones acting on different stages of B cell differentiation. Eur J Immunol. 1988 Nov;18(11):1767–1771. doi: 10.1002/eji.1830181117. [DOI] [PubMed] [Google Scholar]
  34. Nocka K., Tan J. C., Chiu E., Chu T. Y., Ray P., Traktman P., Besmer P. Molecular bases of dominant negative and loss of function mutations at the murine c-kit/white spotting locus: W37, Wv, W41 and W. EMBO J. 1990 Jun;9(6):1805–1813. doi: 10.1002/j.1460-2075.1990.tb08305.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Oettinger M. A., Schatz D. G., Gorka C., Baltimore D. RAG-1 and RAG-2, adjacent genes that synergistically activate V(D)J recombination. Science. 1990 Jun 22;248(4962):1517–1523. doi: 10.1126/science.2360047. [DOI] [PubMed] [Google Scholar]
  36. Palacios R., Karasuyama H., Rolink A. Ly1+ PRO-B lymphocyte clones. Phenotype, growth requirements and differentiation in vitro and in vivo. EMBO J. 1987 Dec 1;6(12):3687–3693. doi: 10.1002/j.1460-2075.1987.tb02702.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Palacios R., Samaridis J. Fetal liver pro-B and pre-B lymphocyte clones: expression of lymphoid-specific genes, surface markers, growth requirements, colonization of the bone marrow, and generation of B lymphocytes in vivo and in vitro. Mol Cell Biol. 1992 Feb;12(2):518–530. doi: 10.1128/mcb.12.2.518. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pleiman C. M., Gimpel S. D., Park L. S., Harada H., Taniguchi T., Ziegler S. F. Organization of the murine and human interleukin-7 receptor genes: two mRNAs generated by differential splicing and presence of a type I-interferon-inducible promoter. Mol Cell Biol. 1991 Jun;11(6):3052–3059. doi: 10.1128/mcb.11.6.3052. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Reith A. D., Ellis C., Lyman S. D., Anderson D. M., Williams D. E., Bernstein A., Pawson T. Signal transduction by normal isoforms and W mutant variants of the Kit receptor tyrosine kinase. EMBO J. 1991 Sep;10(9):2451–2459. doi: 10.1002/j.1460-2075.1991.tb07784.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Rettenmier C. W., Roussel M. F. Differential processing of colony-stimulating factor 1 precursors encoded by two human cDNAs. Mol Cell Biol. 1988 Nov;8(11):5026–5034. doi: 10.1128/mcb.8.11.5026. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rolink A., Melchers F. Molecular and cellular origins of B lymphocyte diversity. Cell. 1991 Sep 20;66(6):1081–1094. doi: 10.1016/0092-8674(91)90032-t. [DOI] [PubMed] [Google Scholar]
  42. Sakaguchi N., Kashiwamura S., Kimoto M., Thalmann P., Melchers F. B lymphocyte lineage-restricted expression of mb-1, a gene with CD3-like structural properties. EMBO J. 1988 Nov;7(11):3457–3464. doi: 10.1002/j.1460-2075.1988.tb03220.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Saris C. J., Domen J., Berns A. The pim-1 oncogene encodes two related protein-serine/threonine kinases by alternative initiation at AUG and CUG. EMBO J. 1991 Mar;10(3):655–664. doi: 10.1002/j.1460-2075.1991.tb07994.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Schatz D. G., Oettinger M. A., Baltimore D. The V(D)J recombination activating gene, RAG-1. Cell. 1989 Dec 22;59(6):1035–1048. doi: 10.1016/0092-8674(89)90760-5. [DOI] [PubMed] [Google Scholar]
  45. Schatz D. G., Oettinger M. A., Schlissel M. S. V(D)J recombination: molecular biology and regulation. Annu Rev Immunol. 1992;10:359–383. doi: 10.1146/annurev.iy.10.040192.002043. [DOI] [PubMed] [Google Scholar]
  46. Scherle P. A., Dorshkind K., Witte O. N. Clonal lymphoid progenitor cell lines expressing the BCR/ABL oncogene retain full differentiative function. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1908–1912. doi: 10.1073/pnas.87.5.1908. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Schwartz R. C., Stanton L. W., Riley S. C., Marcu K. B., Witte O. N. Synergism of v-myc and v-Ha-ras in the in vitro neoplastic progression of murine lymphoid cells. Mol Cell Biol. 1986 Sep;6(9):3221–3231. doi: 10.1128/mcb.6.9.3221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Sudo T., Ito M., Ogawa Y., Iizuka M., Kodama H., Kunisada T., Hayashi S., Ogawa M., Sakai K., Nishikawa S. Interleukin 7 production and function in stromal cell-dependent B cell development. J Exp Med. 1989 Jul 1;170(1):333–338. doi: 10.1084/jem.170.1.333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Tidmarsh G. F., Heimfeld S., Whitlock C. A., Weissman I. L., Müller-Sieburg C. E. Identification of a novel bone marrow-derived B-cell progenitor population that coexpresses B220 and Thy-1 and is highly enriched for Abelson leukemia virus targets. Mol Cell Biol. 1989 Jun;9(6):2665–2671. doi: 10.1128/mcb.9.6.2665. [DOI] [PMC free article] [PubMed] [Google Scholar] [Retracted]
  50. Tsukada S., Sugiyama H., Oka Y., Kishimoto S. Estimation of D segment usage in initial D to JH joinings in a murine immature B cell line. Preferential usage of DFL16.1, the most 5' D segment and DQ52, the most JH-proximal D segment. J Immunol. 1990 May 15;144(10):4053–4059. [PubMed] [Google Scholar]
  51. Whitlock C. A., Robertson D., Witte O. N. Murine B cell lymphopoiesis in long term culture. J Immunol Methods. 1984 Mar 16;67(2):353–369. doi: 10.1016/0022-1759(84)90475-7. [DOI] [PubMed] [Google Scholar]
  52. Whitlock C. A., Witte O. N. Long-term culture of B lymphocytes and their precursors from murine bone marrow. Proc Natl Acad Sci U S A. 1982 Jun;79(11):3608–3612. doi: 10.1073/pnas.79.11.3608. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Whitlock C., Denis K., Robertson D., Witte O. In vitro analysis of murine B-cell development. Annu Rev Immunol. 1985;3:213–235. doi: 10.1146/annurev.iy.03.040185.001241. [DOI] [PubMed] [Google Scholar]
  54. Yamanashi Y., Mori S., Yoshida M., Kishimoto T., Inoue K., Yamamoto T., Toyoshima K. Selective expression of a protein-tyrosine kinase, p56lyn, in hematopoietic cells and association with production of human T-cell lymphotropic virus type I. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6538–6542. doi: 10.1073/pnas.86.17.6538. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Yancopoulos G. D., Desiderio S. V., Paskind M., Kearney J. F., Baltimore D., Alt F. W. Preferential utilization of the most JH-proximal VH gene segments in pre-B-cell lines. Nature. 1984 Oct 25;311(5988):727–733. doi: 10.1038/311727a0. [DOI] [PubMed] [Google Scholar]
  56. Young J. C., Gishizky M. L., Witte O. N. Hyperexpression of interleukin-7 is not necessary or sufficient for transformation of a pre-B lymphoid cell line. Mol Cell Biol. 1991 Feb;11(2):854–863. doi: 10.1128/mcb.11.2.854. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Zimmerman K. A., Yancopoulos G. D., Collum R. G., Smith R. K., Kohl N. E., Denis K. A., Nau M. M., Witte O. N., Toran-Allerand D., Gee C. E. Differential expression of myc family genes during murine development. 1986 Feb 27-Mar 5Nature. 319(6056):780–783. doi: 10.1038/319780a0. [DOI] [PubMed] [Google Scholar]
  58. Zsebo K. M., Wypych J., McNiece I. K., Lu H. S., Smith K. A., Karkare S. B., Sachdev R. K., Yuschenkoff V. N., Birkett N. C., Williams L. R. Identification, purification, and biological characterization of hematopoietic stem cell factor from buffalo rat liver--conditioned medium. Cell. 1990 Oct 5;63(1):195–201. doi: 10.1016/0092-8674(90)90300-4. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press

RESOURCES