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Integration of Notch and Wnt signaling in hematopoietic stem cell maintenance

Nature Immunology volume 6, pages 314–322 (2005)Cite this article

Abstract

A fundamental question in hematopoietic stem cell (HSC) biology is how self-renewal is controlled. Here we show that the molecular regulation of two critical elements of self-renewal, inhibition of differentiation and induction of proliferation, can be uncoupled, and we identify Notch signaling as a key factor in inhibiting differentiation. Using transgenic Notch reporter mice, we found that Notch signaling was active in HSCs in vivo and downregulated as HSCs differentiated. Inhibition of Notch signaling led to accelerated differentiation of HSCs in vitro and depletion of HSCs in vivo. Finally, intact Notch signaling was required for Wnt-mediated maintenance of undifferentiated HSCs but not for survival or entry into the cell cycle in vitro. These data suggest that Notch signaling has a dominant function in inhibiting differentiation and provide a model for how HSCs may integrate multiple signals to maintain the stem cell state.

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Figure 1: Notch signaling is highly active in HSCs and reduced in differentiated cells.
Figure 2: Notch signaling is downregulated during differentiation and defines a more primitive subset within the HSC population.
Figure 3: Inhibition of Notch signaling in HSCs leads to accelerated differentiation.
Figure 4: Inhibition of Notch signaling in vivo leads to depletion of the HSC population after long-term reconstitution.
Figure 5: Inhibition of Notch signaling disrupts ability of Wnt signaling to maintain HSCs in an undifferentiated state.

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Acknowledgements

We thank C. Kintner for the dnXSu(H) construct; S. Artavanis-Tsakonas for the dominant negative MAML1 construct; R. Nusse for help with Wnt3A purification; G. Nolan for providing the retrovirus packaging system; A. Bhandoola, R. Storms and C. Voermans for advice and help; D. Stemmle for lab management; D. McDonnell for use of the iCycler; and I. Weissman, G. Fishell, W. Pear, A. Means and B. Hogan for critical review of this manuscript. Supported by American Heart Association predoctoral fellowships (A.W.D. and K.L.C.), Burroughs Wellcome Fund Cancer Award (N.G.), Kimmel Foundation Scholar Award (N.G.), Cancer Research Institute Investigator Award (T.R.), Ellison Medical Foundation Scholar Award (T.R.) and National Institutes of Health (DK63031 to T.R.).

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  1. Andrew W Duncan and Frédérique M Rattis: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, 27710, North Carolina, USA

    Andrew W Duncan, Frédérique M Rattis, Leah N DiMascio, Kendra L Congdon, Gregory Pazianos, Chen Zhao, J Michael Cook, Karl Willert & Tannishtha Reya

  2. Department of Neurology and Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, 21205, Maryland, USA

    Keejung Yoon & Nicholas Gaiano

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Supplementary information

Supplementary Fig. 1

GFP protein expression reflects active Notch signaline in TNR hematopoietic cells. (PDF 113 kb)

Supplementary Fig. 2

Control staining for cKti and of bone sections from WT mice. (PDF 197 kb)

Supplementary Fig. 3

Analysis of Notch reporter activity in lineage positive cells. (PDF 632 kb)

Supplementary Fig. 4

Pattern of Notch signaling activation in subpopulations of CD4−CD8− double negative thymocytes. (PDF 251 kb)

Supplementary Fig. 5

Analysis of individual lineage markers on in vitro differentiated KLS GFP+ cells. (PDF 57 kb)

Supplementary Fig. 6

Expression of dnXSu(H) in virally-transduced HSCs. (PDF 141 kb)

Supplementary Fig. 7

Lineage markers are equivalently expressed in uninfected KTLS cells transduced with control or dnXSu(H) retroviruses. (PDF 81 kb)

Supplementary Fig. 8

Inhibition of Notch signaling in vivo leads to an increase in myeloid and B lineage cells and to vector silencing in double negative thymocytes in vivo. (PDF 228 kb)

Supplementary Methods (PDF 49 kb)

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Duncan, A., Rattis, F., DiMascio, L. et al. Integration of Notch and Wnt signaling in hematopoietic stem cell maintenance. Nat Immunol 6, 314–322 (2005). https://doi.org/10.1038/ni1164

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