Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Notch signaling maintains bone marrow mesenchymal progenitors by suppressing osteoblast differentiation

Abstract

Postnatal bone marrow houses mesenchymal progenitor cells that are osteoblast precursors. These cells have established therapeutic potential, but they are difficult to maintain and expand in vitro, presumably because little is known about the mechanisms controlling their fate decisions. To investigate the potential role of Notch signaling in osteoblastogenesis, we used conditional alleles to genetically remove components of the Notch signaling system during skeletal development. We found that disruption of Notch signaling in the limb skeletogenic mesenchyme markedly increased trabecular bone mass in adolescent mice. Notably, mesenchymal progenitors were undetectable in the bone marrow of mice with high bone mass. As a result, these mice developed severe osteopenia as they aged. Moreover, Notch signaling seemed to inhibit osteoblast differentiation through Hes or Hey proteins, which diminished Runx2 transcriptional activity via physical interaction. These results support a model wherein Notch signaling in bone marrow normally acts to maintain a pool of mesenchymal progenitors by suppressing osteoblast differentiation. Thus, mesenchymal progenitors may be expanded in vitro by activating the Notch pathway, whereas bone formation in vivo may be enhanced by transiently suppressing this pathway.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Skeletal phenotype of PPS and Psen2-null mice at 8 weeks of age.
Figure 2: Skeletal phenotype of PNN and wild type (WT) mice at 8 weeks of age.
Figure 3: Molecular and histological analyses in PNN and WT embryos.
Figure 4: Notch regulation of bone marrow mesenchymal progenitors.
Figure 5: Progressive bone loss in mature WT and PNN mice.
Figure 6: Mechanisms for Notch function in bone.

References

  1. Artavanis-Tsakonas, S., Rand, M.D. & Lake, R.J. Notch signaling: cell fate control and signal integration in development. Science 284, 770–776 (1999).

    Article  CAS  PubMed  Google Scholar 

  2. Chiba, S. Notch signaling in stem cell systems. Stem Cells 24, 2437–2447 (2006).

    Article  CAS  PubMed  Google Scholar 

  3. Schroeter, E.H., Kisslinger, J.A. & Kopan, R. Notch-1 signalling requires ligand-induced proteolytic release of intracellular domain. Nature 393, 382–386 (1998).

    Article  CAS  PubMed  Google Scholar 

  4. Honjo, T. The shortest path from the surface to the nucleus: RBP-Jκ/Su(H) transcription factor. Genes Cells 1, 1–9 (1996).

    Article  CAS  PubMed  Google Scholar 

  5. Kopan, R. & Goate, A. A common enzyme connects Notch signaling and Alzheimer's disease. Genes Dev. 14, 2799–2806 (2000).

    Article  CAS  PubMed  Google Scholar 

  6. Donoviel, D.B. et al. Mice lacking both presenilin genes exhibit early embryonic patterning defects. Genes Dev. 13, 2801–2810 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Herreman, A. et al. Presenilin 2 deficiency causes a mild pulmonary phenotype and no changes in amyloid precursor protein processing but enhances the embryonic lethal phenotype of presenilin 1 deficiency. Proc. Natl. Acad. Sci. USA 96, 11872–11877 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Krebs, L.T. et al. Notch signaling is essential for vascular morphogenesis in mice. Genes Dev. 14, 1343–1352 (2000).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Hamada, Y. et al. Mutation in ankyrin repeats of the mouse Notch2 gene induces early embryonic lethality. Development 126, 3415–3424 (1999).

    CAS  PubMed  Google Scholar 

  10. Swiatek, P.J., Lindsell, C.E., del Amo, F.F., Weinmaster, G. & Gridley, T. Notch1 is essential for postimplantation development in mice. Genes Dev. 8, 707–719 (1994).

    Article  CAS  PubMed  Google Scholar 

  11. Domenga, V. et al. Notch3 is required for arterial identity and maturation of vascular smooth muscle cells. Genes Dev. 18, 2730–2735 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Hrabe de Angelis, M., McIntyre, J. II & Gossler, A. Maintenance of somite borders in mice requires the Delta homologue DII1. Nature 386, 717–721 (1997).

    Article  CAS  PubMed  Google Scholar 

  13. Duarte, A. et al. Dosage-sensitive requirement for mouse Dll4 in artery development. Genes Dev. 18, 2474–2478 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Gale, N.W. et al. Haploinsufficiency of delta-like 4 ligand results in embryonic lethality due to major defects in arterial and vascular development. Proc. Natl. Acad. Sci. USA 101, 15949–15954 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Xue, Y. et al. Embryonic lethality and vascular defects in mice lacking the Notch ligand Jagged1. Hum. Mol. Genet. 8, 723–730 (1999).

    Article  CAS  PubMed  Google Scholar 

  16. Jiang, R. et al. Defects in limb, craniofacial, and thymic development in Jagged2 mutant mice. Genes Dev. 12, 1046–1057 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Dunwoodie, S.L. et al. Axial skeletal defects caused by mutation in the spondylocostal dysplasia/pudgy gene Dll3 are associated with disruption of the segmentation clock within the presomitic mesoderm. Development 129, 1795–1806 (2002).

    CAS  PubMed  Google Scholar 

  18. Shen, J. et al. Skeletal and CNS defects in presenilin-1–deficient mice. Cell 89, 629–639 (1997).

    Article  CAS  PubMed  Google Scholar 

  19. Wong, P.C. et al. Presenilin 1 is required for Notch1 and DII1 expression in the paraxial mesoderm. Nature 387, 288–292 (1997).

    Article  CAS  PubMed  Google Scholar 

  20. Pan, Y., Liu, Z., Shen, J. & Kopan, R. Notch1 and 2 cooperate in limb ectoderm to receive an early Jagged2 signal regulating interdigital apoptosis. Dev. Biol. 286, 472–482 (2005).

    Article  CAS  PubMed  Google Scholar 

  21. Logan, M. et al. Expression of CrerRecombinase in the developing mouse limb bud driven by a Prxl enhancer. Genesis 33, 77–80 (2002).

    Article  CAS  PubMed  Google Scholar 

  22. Miao, D. et al. Osteoblast-derived PTHrP is a potent endogenous bone anabolic agent that modifies the therapeutic efficacy of administered PTH 1–34. J. Clin. Invest. 115, 2402–2411 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Owen, M. & Friedenstein, A.J. Stromal stem cells: marrow-derived osteogenic precursors. Ciba Found. Symp. 136, 42–60 (1988).

    CAS  PubMed  Google Scholar 

  24. Kuznetsov, S.A. et al. The interplay of osteogenesis and hematopoiesis: expression of a constitutively active PTH/PTHrP receptor in osteogenic cells perturbs the establishment of hematopoiesis in bone and of skeletal stem cells in the bone marrow. J. Cell Biol. 167, 1113–1122 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Nichols, A.M. et al. Notch pathway is dispensable for adipocyte specification. Genesis 40, 40–44 (2004).

    Article  CAS  PubMed  Google Scholar 

  26. Karsenty, G. & Wagner, E.F. Reaching a genetic and molecular understanding of skeletal development. Dev. Cell 2, 389–406 (2002).

    Article  CAS  PubMed  Google Scholar 

  27. Bai, S. et al. Notch1 regulates osteoclastogenesis directly in osteoclast precursors and indirectly via osteoblast lineage cells. J. Biol. Chem. published online, doi:10.1074/jbc.M707000200 (22 December 2007).

  28. Iso, T., Kedes, L. & Hamamori, Y. HES and HERP families: multiple effectors of the Notch signaling pathway. J. Cell. Physiol. 194, 237–255 (2003).

    Article  CAS  PubMed  Google Scholar 

  29. Duncan, A.W. et al. Integration of Notch and Wnt signaling in hematopoietic stem cell maintenance. Nat. Immunol. 6, 314–322 (2005).

    Article  CAS  PubMed  Google Scholar 

  30. Vooijs, M. et al. Mapping the consequence of Notch1 proteolysis in vivo with NIP-CRE. Development 134, 535–544 (2007).

    Article  CAS  PubMed  Google Scholar 

  31. Soriano, P. Generalized lacZ expression with the ROSA26 Cre reporter strain. Nat. Genet. 21, 70–71 (1999).

    Article  CAS  PubMed  Google Scholar 

  32. Deregowski, V., Gazzerro, E., Priest, L., Rydziel, S. & Canalis, E. Notch 1 overexpression inhibits osteoblastogenesis by suppressing Wnt/β-catenin but not bone morphogenetic protein signaling. J. Biol. Chem. 281, 6203–6210 (2006).

    Article  CAS  PubMed  Google Scholar 

  33. Sciaudone, M., Gazzerro, E., Priest, L., Delany, A.M. & Canalis, E. Notch 1 impairs osteoblastic cell differentiation. Endocrinology 144, 5631–5639 (2003).

    Article  CAS  PubMed  Google Scholar 

  34. Shindo, K. et al. Osteogenic differentiation of the mesenchymal progenitor cells, Kusa is suppressed by Notch signaling. Exp. Cell Res. 290, 370–380 (2003).

    Article  CAS  PubMed  Google Scholar 

  35. Nobta, M. et al. Critical regulation of bone morphogenetic protein-induced osteoblastic differentiation by Delta1/Jagged1-activated Notch1 signaling. J. Biol. Chem. 280, 15842–15848 (2005).

    Article  CAS  PubMed  Google Scholar 

  36. Tezuka, K. et al. Stimulation of osteoblastic cell differentiation by Notch. J. Bone Miner. Res. 17, 231–239 (2002).

    Article  CAS  PubMed  Google Scholar 

  37. Garg, V. et al. Mutations in NOTCH1 cause aortic valve disease. Nature 437, 270–274 (2005).

    Article  CAS  PubMed  Google Scholar 

  38. Zamurovic, N., Cappellen, D., Rohner, D. & Susa, M. Coordinated activation of notch, Wnt, and transforming growth factor-β signaling pathways in bone morphogenic protein 2–induced osteogenesis. Notch target gene Hey1 inhibits mineralization and Runx2 transcriptional activity. J. Biol. Chem. 279, 37704–37715 (2004).

    Article  CAS  PubMed  Google Scholar 

  39. Conlon, R.A., Reaume, A.G. & Rossant, J. Notch1 is required for the coordinate segmentation of somites. Development 121, 1533–1545 (1995).

    CAS  PubMed  Google Scholar 

  40. Pan, Y. et al. Gamma-secretase functions through Notch signaling to maintain skin appendages but is not required for their patterning or initial morphogenesis. Dev. Cell 7, 731–743 (2004).

    Article  CAS  PubMed  Google Scholar 

  41. McCright, B., Lozier, J. & Gridley, T. Generation of new Notch2 mutant alleles. Genesis 44, 29–33 (2006).

    Article  CAS  PubMed  Google Scholar 

  42. Yu, H. et al. APP processing and synaptic plasticity in presenilin-1 conditional knockout mice. Neuron 31, 713–726 (2001).

    Article  CAS  PubMed  Google Scholar 

  43. Hilton, M.J., Tu, X., Cook, J., Hu, H. & Long, F. Ihh controls cartilage development by antagonizing Gli3, but requires additional effectors to regulate osteoblast and vascular development. Development 132, 4339–4351 (2005).

    Article  CAS  PubMed  Google Scholar 

  44. Tu, X. et al. Noncanonical Wnt signaling through G protein–linked PKCδ activation promotes bone formation. Dev. Cell 12, 113–127 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Huppert, S.S. et al. Embryonic lethality in mice homozygous for a processing-deficient allele of Notch1. Nature 405, 966–970 (2000).

    Article  CAS  PubMed  Google Scholar 

  46. Ong, C.T. et al. Target selectivity of vertebrate notch proteins. Collaboration between discrete domains and CSL-binding site architecture determines activation probability. J. Biol. Chem. 281, 5106–5119 (2006).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported in part by US National Institutes of Health grants DK065789 (F.L.), HD044056 (R.K.), AR046852 (F.P.R.), AR046523 (S.L.T.), DK11794 (H.M.K.) and 5T32AR07033 (M.J.H.). We thank R. Civitelli (Washington University) for pCMV-Runx2 and p60SE2-Luc, R. Kageyama (Kyoto University) for pSV2-CMV-Hes1, P. Robey and S. Kuznetsov for advice on bone marrow CFU-F assays and J. Shen (Harvard Medical School) and Thomas Gridley (Jackson Laboratory) for mouse strains. We also thank D. Towler and M. Silva for their help with the response to reviewers' comments.

Author information

Authors and Affiliations

Authors

Contributions

M.J.H., animal studies and figure preparation; X.T., cell culture work and animal studies; X.W. coimmunoprecipitation experiments; S.B., H.Z., F.P.R. and S.L.T., TRAP staining, serum CTX1 assays and discussion; T.K. and H.M.K., Col1-Cre mice; R.K., Notch reagents and manuscript editing; F.L., project directing, figure preparation and manuscript writing.

Corresponding author

Correspondence to Fanxin Long.

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–5 and Supplementary Table 1 (PDF 937 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hilton, M., Tu, X., Wu, X. et al. Notch signaling maintains bone marrow mesenchymal progenitors by suppressing osteoblast differentiation. Nat Med 14, 306–314 (2008). https://doi.org/10.1038/nm1716

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm1716

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing