Critical functions for c-Myb at three checkpoints during thymocyte development

Article metrics


The transcription factor c-Myb is expressed throughout T cell development in the thymus. However, little is understood about c-Myb function because of the embryonic lethality of traditional Myb-null mutations. Using tissue-specific deletion to abrogate c-Myb expression at distinct stages of T cell development, we identify three points at which c-Myb activity is required for normal T cell differentiation: transition through the double-negative 3 stage, survival of preselection CD4+CD8+ thymocytes, and differentiation of CD4 thymocytes. Thus, c-Myb is essential at several stages during T cell development in the thymus.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Expression of Myb mRNA in the thymus and conditional targeting of the mouse Myb locus.
Figure 2: Tissue- and differentiation-specific deletion of the mouse Myb locus.
Figure 3: Impaired thymocyte development in Mybf/d LckCre and Mybf/f cwLckCre mice.
Figure 4: Differentiation and proliferation but decreased intracellular TCRβ expression in c-Myb-deficient DN3 thymocytes.
Figure 5: Inefficient V(D)J recombination at the Tcrb locus in c-Myb-deficient DN3 thymocytes.
Figure 6: c-Myb is required for DP thymocyte survival and is more important for differentiation of CD4 SP than CD8 SP cells.
Figure 7: Decreased Vα→Jα rearrangement at the Tcra locus in c-Myb-deficient DP thymocytes.


  1. 1

    Fehling, H.J., Gilfillan, S. & Ceredig, R. αβ/γδ lineage commitment in the thymus of normal and genetically manipulated mice. Adv. Immunol. 71, 1–76 (1999).

  2. 2

    Godfrey, D.I., Kennedy, J., Suda, T. & Zlotnik, A. A developmental pathway involving four phenotypically and functionally distinct subsets of CD3CD4CD8 triple-negative adult mouse thymocytes defined by CD44 and CD25 expression. J. Immunol. 150, 4244–4252 (1993).

  3. 3

    Guidos, C.J., Weissman, I.L. & Adkins, B. Intrathymic maturation of murine T lymphocytes from CD8+ precursors. Proc. Natl. Acad. Sci. USA 86, 7542–7546 (1989).

  4. 4

    von Boehmer, H. et al. Pleiotropic changes controlled by the pre-T-cell receptor. Curr. Opin. Immunol. 11, 135–142 (1999).

  5. 5

    Germain, R.N. T-cell development and the CD4-CD8 lineage decision. Nat. Rev. Immunol. 2, 309–322 (2002).

  6. 6

    Oh, I.H. & Reddy, E.P. The myb gene family in cell growth, differentiation and apoptosis. Oncogene 18, 3017–3033 (1999).

  7. 7

    Akashi, K., Traver, D., Miyamoto, T. & Weissman, I.L. A clonogenic common myeloid progenitor that gives rise to all myeloid lineages. Nature. 404, 193–197 (2000).

  8. 8

    Westin, E.H. et al. Differential expression of the amv gene in human hematopoietic cells. Proc. Natl. Acad. Sci. USA 79, 2194–2198 (1982).

  9. 9

    Mucenski, M.L. et al. A functional c-myb gene is required for normal murine fetal hepatic hematopoiesis. Cell 65, 677–689 (1991).

  10. 10

    Ess, K.C., Witte, D.P., Bascomb, C.P. & Aronow, B.J. Diverse developing mouse lineages exhibit high-level c-Myb expression in immature cells and loss of expression upon differentiation. Oncogene 18, 1103–1111 (1999).

  11. 11

    Stern, J.B. & Smith, K.A. Interleukin-2 induction of T-cell G1 progression and c-myb expression. Science 233, 203–206 (1986).

  12. 12

    Allen, R.D., Bender, T.P. & Siu, G. c-Myb is essential for T cell development. Genes Dev. 13, 1073–1078 (1999).

  13. 13

    Bender, T.P., Thompson, C.B. & Kuehl, W.M. Differential expression of c-myb mRNA in murine B lymphomas by a block to transcription elongation. Science 237, 1473–1476 (1987).

  14. 14

    Torres, R.M. & Kuhn, R. Laboratory Protocols for Conditional Gene Targeting (Oxford University Press, 1997).

  15. 15

    Lavu, S. & Reddy, E.P. Structural organization and nucleotide sequence of mouse c-myb oncogene: activation in ABPL tumors is due to viral integration in an intron which results in the deletion of the 5′ coding sequences. Nucleic Acids Res. 14, 5309–5320 (1986).

  16. 16

    Jacobs, S.M., Gorse, K.M. & Westin, E.H. Identification of a second promoter in the human c-myb proto-oncogene. Oncogene 9, 227–235 (1994).

  17. 17

    Hennet, T., Hagen, F.K., Tabak, L.A. & Marth, J.D. T-cell-specific deletion of a polypeptide N-acetylgalactosaminyl-transferase gene by site-directed recombination. Proc. Natl. Acad. Sci. USA 92, 12070–12074 (1995).

  18. 18

    Lee, P.P. et al. A critical role for Dnmt1 and DNA methylation in T cell development, function, and survival. Immunity. 15, 763–774 (2001).

  19. 19

    Luscher, B. & Eisenman, R.N. c-myc and c-myb protein degradation: effect of metabolic inhibitors and heat shock. Mol. Cell. Biol. 8, 2504–2512 (1988).

  20. 20

    McMurry, M.T., Hernandez-Munain, C., Lauzurica, P. & Krangel, M.S. Enhancer control of local accessibility to V(D)J recombinase. Mol. Cell. Biol. 17, 4553–4561 (1997).

  21. 21

    Hsiang, H.H., Goldman, J.P. & Raulet, D.H. The role of c-Myb or a related factor in regulating the T cell receptor γ gene enhancer. J. Immunol. 154, 5195–5204 (1995).

  22. 22

    Penit, C., Lucas, B. & Vasseur, F. Cell expansion and growth arrest phases during the transition from precursor (CD48) to immature (CD4+8+) thymocytes in normal and genetically modified mice. J. Immunol. 154, 5103–5113 (1995).

  23. 23

    Sleckman, B.P., Gorman, J.R. & Alt, F.W. Accessibility control of antigen-receptor variable-region gene assembly: role of cis-acting elements. Ann. Rev. Immunol. 14, 459–481 (1996).

  24. 24

    Reizis, B. & Leder, P. The upstream enhancer is necessary and sufficient for the expression of the pre-T cell receptor α gene in immature T lymphocytes. J. Exp. Med. 194, 979–990 (2001).

  25. 25

    Shinkai, Y. & Alt, F.W. CD3 epsilon-mediated signals rescue the development of CD4+CD8+ thymocytes in RAG-2−/− mice in the absence of TCR β chain expression. Int. Immunol. 6, 995–1001 (1994).

  26. 26

    Sun, Z. et al. Requirement for RORγ in thymocyte survival and lymphoid organ development. Science. 288, 2369–2373 (2000).

  27. 27

    Gartner, F. et al. Immature thymocytes employ distinct signaling pathways for allelic exclusion versus differentiation and expansion. Immunity. 10, 537–546 (1999).

  28. 28

    Barnden, M.J., Allison, J., Heath, W.R. & Carbone, F.R. Defective TCR expression in transgenic mice constructed using cDNA-based α- and β-chain genes under the control of heterologous regulatory elements. Immunol. Cell. Biol. 76, 34–40 (1998).

  29. 29

    Bluthmann, H. et al. T-cell-specific deletion of T-cell receptor transgenes allows functional rearrangement of endogenous α- and β-genes. Nature 334, 156–159 (1988).

  30. 30

    Huesmann, M., Scott, B., Kisielow, P. & von Boehmer, H. Kinetics and efficacy of positive selection in the thymus of normal and T cell receptor transgenic mice. Cell 66, 533–540 (1991).

  31. 31

    Anderson, S.J., Abraham, K.M., Nakayama, T., Singer, A. & Perlmutter, R.M. Inhibition of T-cell receptor β-chain gene rearrangement by overexpression of the non-receptor protein tyrosine kinase p56lck. EMBO J. 11, 4877–4886 (1992).

  32. 32

    Pearson, R. & Weston, K. c-Myb regulates the proliferation of immature thymocytes following β-selection. EMBO J. 19, 6112–6120 (2000).

  33. 33

    Wang, Q.F., Lauring, J. & Schlissel, M.S. c-Myb binds to a sequence in the proximal region of the RAG-2 promoter and is essential for promoter activity in T-lineage cells. Mol. Cell. Biol. 20, 9203–9211 (2000).

  34. 34

    Chen, F., Rowen, L., Hood, L. & Rothenberg, E.V. Differential transcriptional regulation of individual TCR Vβ segments before gene rearrangement. J. Immunol. 166, 1771–1780 (2001).

  35. 35

    Krangel, M.S. Gene segment selection in V(D)J recombination: accessibility and beyond. Nat. Immunol. 4, 624–630 (2003).

  36. 36

    McClinton, D., Stafford, J., Brents, L., Bender, T.P. & Kuehl, W.M. Differentiation of mouse erythroleukemia cells is blocked by late up-regulation of a c-myb transgene. Mol. Cell. Biol. 10, 705–710 (1990).

  37. 37

    Gonda, T.J. & Metcalf, D. Expression of myb, myc and fos proto-oncogenes during the differentiation of a murine myeloid leukaemia. Nature 310, 249–251 (1984).

  38. 38

    Taylor, D., Badiani, P. & Weston, K. A dominant interfering Myb mutant causes apoptosis in T cells. Genes Dev. 10, 2732–2744 (1996).

  39. 39

    Ma, A. et al. Bclx regulates the survival of double-positive thymocytes. Proc. Natl. Acad. Sci. USA 92, 4763–4767 (1995).

  40. 40

    Guo, J. et al. Regulation of the TCRα repertoire by the survival window of CD4+CD8+ thymocytes. Nat. Immunol. 3, 469–476 (2002).

  41. 41

    Siu, G., Wurster, A.L., Lipsick, J.S. & Hedrick, S.M. Expression of the CD4 gene requires a Myb transcription factor. Mol. Cell. Biol. 12, 1592–1604 (1992).

  42. 42

    Allen, R.D., III, Kim, H.K., Sarafova, S.D. & Siu, G. Negative regulation of CD4 gene expression by a HES-1-c-Myb complex. Mol. Cell. Biol. 21, 3071–3082 (2001).

  43. 43

    Luscher, B., Christenson, E., Litchfield, D.W., Krebs, E.G. & Eisenman, R.N. Myb DNA binding inhibited by phosphorylation at a site deleted during oncogenic activation. Nature 344, 517–522 (1990).

  44. 44

    Miglarese, M.R., Richardson, A.F., Aziz, N. & Bender, T.P. Differential regulation of c-Myb-induced transcription activation by a phosphorylation site in the negative regulatory domain. J. Biol. Chem. 271, 22697–22705 (1996).

  45. 45

    Ramsay, R.G. et al. Regulation of c-Myb through protein phosphorylation and leucine zipper interactions. Oncogene 11, 2113–2120 (1995).

  46. 46

    Emambokus, N. et al. Progression through key stages of haemopoiesis is dependent on distinct threshold levels of c-Myb. EMBO J. 22, 4478–4488 (2003).

  47. 47

    Zhang, L., Camerini, V., Bender, T.P. & Ravichandran, K.S. A nonredundant role for the adapter protein Shc in thymic T cell development. Nat. Immunol. 3, 749–755 (2002).

Download references


The authors thank A. Roth (University of Cologne, Cologne, Germany) for doing the blastocyst injections; C. Goettlinger (University of Cologne) and J. Lannigan (University of Virginia, Charlottesville, Virginia) for help with cell sorting; and M. McDuffie, V. Engelhard, U. Lorenz and K. Ravichandran for critical review of the manuscript. Supported by National Institutes of Health (CA85842 to T.P.B.), Fogarty International Center (TW02297 to T.P.B.), Deutsche Forschungsgemeinschaft (SFB 243 to K.R.) and the European Union (BIO4-CT96-0077 to K.R.).

Author information

Correspondence to Timothy P Bender.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Expression of c-Myb protein in Mybf/f lckCre thymocytes. (PDF 169 kb)

Supplementary Fig. 2

Semi-quantitative RT-PCR analysis of Rag1 and Rag2 expression in c-Myb-deficient thymocytes. (PDF 232 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Bender, T., Kremer, C., Kraus, M. et al. Critical functions for c-Myb at three checkpoints during thymocyte development. Nat Immunol 5, 721–729 (2004) doi:10.1038/ni1085

Download citation

Further reading