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Activation of the canonical Wnt pathway leads to loss of hematopoietic stem cell repopulation and multilineage differentiation block

Abstract

Wnt signaling increases hematopoietic stem cell self-renewal and is activated in both myeloid and lymphoid malignancies, indicating involvement in both normal and malignant hematopoiesis. We report here activated canonical Wnt signaling in the hematopoietic system through conditional expression of a stable form of β-catenin. This enforced expression led to hematopoietic failure associated with loss of myeloid lineage commitment at the granulocyte-macrophage progenitor stage; blocked erythrocyte differentiation; disruption of lymphoid development; and loss of repopulating stem cell activity. Loss of hematopoietic stem cell function was associated with decreased expression of Cdkn1a (encoding the cell cycle inhibitor p21cdk), Sfpi1, Hoxb4 and Bmi1 (encoding the transcription factors PU.1, HoxB4 and Bmi-1, respectively) and altered integrin expression in LinSca-1+c-Kit+ cells, whereas PU.1 was upregulated in erythroid progenitors. Constitutive activation of canonical Wnt signaling therefore causes multilineage differentiation block and compromised hematopoietic stem cell maintenance.

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Figure 1: 'Knock-in' of the gene encoding β-catS33Y into the R26 locus and its inducible expression.
Figure 2: Induced expression of activated β-catS33Y in adult bone marrow results in severe pancytopenia.
Figure 3: Bone marrow cells expressing activated β-catS33Y show multiple lineage defects.
Figure 4: Lymphoid phenotype induced by β-catS33Y expression.
Figure 5: Immunophenotypic and functional analysis of HSCs expressing activated β-catS33Y.
Figure 6: Deregulation of integrins in response to expression of β-catS33Y.
Figure 7: Gene expression changes induced by β-catS33Y in LSK and erythroid cells.

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Acknowledgements

We thank E. Fearon (University of Michigan, Ann Arbor, Michigan), P. Soriano (Fred Hutchinson Cancer Research Center, Seattle, Washington), K. Rajewsky and D.G. Tenen (Harvard Medical School, Boston, Massachusetts) and F. Stewart (University of Technology, Dresden, Germany) for providing mouse strains and reagents; O. Ermakova for the generation of PU.1-YFP 'knock-in' mice; and the FACS facility at the Lund Stem Cell Center for cell sorting. Supported by the European Commission (EuroStemCell integrated project), the Association for International Cancer Research, the Swedish Research Council, and the Swedish Foundation for Strategic Research (for the Lund Stem Cell Center).

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Authors and Affiliations

Authors

Contributions

P.K., phenotypic analysis of R26-LNL-βcatS33Y 'knock-in' mice, bone marrow transplantation and analysis of transplanted mice; K.A., cell sorting, bone marrow transplantation and analysis of transplanted mice; B.T.P., generation of the targeting construct and targeted embryonic stem cells lines used to produce R26-LNL-βcatS33Y 'knock-in' mice; S.E.W.J., contributions to the experimental design and writing of the manuscript; C.N., experimental design and writing of the manuscript.

Corresponding author

Correspondence to Claus Nerlov.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Viability of mice expressing activated β-cat-S33Y. (PDF 310 kb)

Supplementary Fig. 2

Decreased apoptosis in LSK cells expressing β-cat-S33Y. (PDF 349 kb)

Supplementary Fig. 3

Increased frequency of LSK cells in BM expressing β-cat-S33Y. (PDF 564 kb)

Supplementary Fig. 4

Competitive BM transplantation assay. (PDF 576 kb)

Supplementary Fig. 5

Non-competitive BM transplantation assay. (PDF 411 kb)

Supplementary Fig. 6

Competitive BM transplantation prior to β-catenin induction. (PDF 785 kb)

Supplementary Table 1

Primers for qRT-PCR. (PDF 72 kb)

Supplementary Methods (PDF 72 kb)

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Kirstetter, P., Anderson, K., Porse, B. et al. Activation of the canonical Wnt pathway leads to loss of hematopoietic stem cell repopulation and multilineage differentiation block. Nat Immunol 7, 1048–1056 (2006). https://doi.org/10.1038/ni1381

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