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M-CSF instructs myeloid lineage fate in single haematopoietic stem cells

Nature volume 497, pages 239–243 (2013)Cite this article

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Abstract

Under stress conditions such as infection or inflammation the body rapidly needs to generate new blood cells that are adapted to the challenge. Haematopoietic cytokines are known to increase output of specific mature cells by affecting survival, expansion and differentiation of lineage-committed progenitors1,2, but it has been debated whether long-term haematopoietic stem cells (HSCs) are susceptible to direct lineage-specifying effects of cytokines. Although genetic changes in transcription factor balance can sensitize HSCs to cytokine instruction3, the initiation of HSC commitment is generally thought to be triggered by stochastic fluctuation in cell-intrinsic regulators such as lineage-specific transcription factors4,5,6,7, leaving cytokines to ensure survival and proliferation of the progeny cells8,9. Here we show that macrophage colony-stimulating factor (M-CSF, also called CSF1), a myeloid cytokine released during infection and inflammation, can directly induce the myeloid master regulator PU.1 and instruct myeloid cell-fate change in mouse HSCs, independently of selective survival or proliferation. Video imaging and single-cell gene expression analysis revealed that stimulation of highly purified HSCs with M-CSF in culture resulted in activation of the PU.1 promoter and an increased number of PU.1+ cells with myeloid gene signature and differentiation potential. In vivo, high systemic levels of M-CSF directly stimulated M-CSF-receptor-dependent activation of endogenous PU.1 protein in single HSCs and induced a PU.1-dependent myeloid differentiation preference. Our data demonstrate that lineage-specific cytokines can act directly on HSCs in vitro and in vivo to instruct a change of cell identity. This fundamentally changes the current view of how HSCs respond to environmental challenge and implicates stress-induced cytokines as direct instructors of HSC fate.

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Figure 1: M-CSF activates the myeloid master regulator PU.1 in HSCs.
Figure 2: Continuous video imaging of PU.1+ cell generation from individual PU.1− HSCs.
Figure 3: M-CSF activates PU.1 and instructs myeloid identity in single HSCs.
Figure 4: M-CSF directly induces endogenous PU.1 protein in single HSCs in vivo and stimulates a reversible, PU.1-dependent myeloid differentiation preference.

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Acknowledgements

We acknowledge grants from the ‘Association pour la recherche sur le Cancer’ (3422) and the ‘Agence nationale de la Recherche’ (BLAN07-1_205752). We thank P. Kastner and S. Chan for PU.1-GFP reporter mice; T. P. VuManh and J. Maurizio for bioinformatics; M. Barad, A. Zouine and M.-L. Thibult for flow cytometry support; L. Razafindramanana for animal handling; and J. Favret, P. Perrin and L. Chasson for tissue sectioning. E.R.S. is supported by NIH grant CA 32551. S.L.N. is an Australian Research Council Future Fellow and received Victorian State Government Operational and Australian Government NHMRC Independent Research Institute Infrastructure Support. M.H.S. is a ‘Fondation pour la Recherche Médicale’ (DEQ20071210559; DEQ20110421320) and INSERM-Helmholtz group leader.

Author information

Author notes

  1. Noushine Mossadegh-Keller and Sandrine Sarrazin: These authors contributed equally to this work.

Authors and Affiliations

  1. Centre d’Immunologie de Marseille-Luminy, Aix-Marseille Université, UM2, Campus de Luminy, Case 906, 13288 Marseille Cedex 09, France ,

    Noushine Mossadegh-Keller, Sandrine Sarrazin, Prashanth K. Kandalla & Michael H. Sieweke

  2. Institut National de la Santé et de la Recherche Médicale, U1104, Campus de Luminy, Case 906, 13288 Marseille Cedex 09, France ,

    Noushine Mossadegh-Keller, Sandrine Sarrazin, Prashanth K. Kandalla & Michael H. Sieweke

  3. Centre National de la Recherche Scientifique, UMR7280, Campus de Luminy, Case 906, Marseille Cedex 09, 13288, France

    Noushine Mossadegh-Keller, Sandrine Sarrazin, Prashanth K. Kandalla & Michael H. Sieweke

  4. Laboratoire de Chimie Bactérienne, Centre National de la Recherche Scientifique, UMR 7283, 31 Chemin Joseph Aiguier, 13009 Marseille, France ,

    Leon Espinosa

  5. Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, 10461, New York, USA

    E. Richard Stanley

  6. Walter and Eliza Hall Institute, 1G Royal Parade, Parkville, 3052, Victoria, Australia

    Stephen L. Nutt

  7. Fluidigm Corporation, South San Francisco, 94080, California, USA

    Jordan Moore

  8. Max-Delbrück-Centrum für Molekulare Medizin, Robert-Rössle-Str. 10, 13125 Berlin, Germany ,

    Michael H. Sieweke

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  1. Noushine Mossadegh-Keller
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Contributions

M.H.S. conceived the study, analysed and interpreted data and wrote the paper; S.S. performed experiments, analysed and interpreted data and contributed to the preparation of the manuscript; N.M.-K. performed most experiments and analysed data; P.K.K. performed and analysed video microscopy and contributed to other experiments; L.E. analysed and interpreted video microscopy data; J.M. provided expertise and service on Fluidigm experiments; E.R.S. and S.L.N. provided essential M-CSFR and PU.1-deficient haematopoietic cells. N.M.-K. and S.S. contributed equally to the study. N.M.-K., S.S., P.K.K., L.E. and M.H.S. jointly designed experiments and S.S. and M.H.S. coordinated the project.

Corresponding authors

Correspondence to Sandrine Sarrazin or Michael H. Sieweke.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-14, Supplementary Table 1, a Supplementary Discussion and Supplementary References. (PDF 748 kb)

GFP negative HSC

Time lapse microscopy recording bright field and GFP-fluorescence in 10 minute intervals of a representative GFP negative HSC sorted from PU.1-GFP reporter mice in M-CSF culture, showing no GFP activation. (AVI 259 kb)

GFP positive HSC 1

Time lapse microscopy recording bright field and GFP-fluorescence in 10 minute intervals of a representative GFP negative HSC sorted from PU.1-GFP reporter mice in M-CSF culture, showing GFP activation. (AVI 176 kb)

GFP positive HSC 2

Time lapse microscopy recording bright field and GFP-fluorescence in 10 minute intervals of a representative GFP negative HSC sorted from PU.1-GFP reporter mice in M-CSF culture, showing GFP activation. (AVI 252 kb)

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Mossadegh-Keller, N., Sarrazin, S., Kandalla, P. et al. M-CSF instructs myeloid lineage fate in single haematopoietic stem cells. Nature 497, 239–243 (2013). https://doi.org/10.1038/nature12026

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