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LEF-1 is crucial for neutrophil granulocytopoiesis and its expression is severely reduced in congenital neutropenia

Nature Medicine volume 12, pages 1191–1197 (2006)Cite this article

An Erratum to this article was published on 01 November 2006

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Abstract

We demonstrate here that lymphoid enhancer-binding factor 1 (LEF-1) mediates the proliferation, survival and differentiation of granulocyte progenitor cells. We initially documented the importance of this transcription factor in the bone marrow of individuals with severe congenital neutropenia (CN) with a 'differentiation block' at the promyelocytic stage of myelopoiesis1. LEF-1 expression was greatly reduced or even absent in CN arrested promyelocytes, resulting in defective expression of the LEF-1 target genes CCND1, MYC and BIRC5, encoding cyclin D1 (ref. 2), c-Myc3 and survivin4, respectively. In contrast, healthy individuals showed highest LEF-1 expression in promyelocytes. Reconstitution of LEF-1 in early hematopoietic progenitors of two individuals with CN corrected the defective myelopoiesis and resulted in the differentiation of these progenitors into mature granulocytes. Repression of endogenous LEF-1 by specific short hairpin RNA inhibited proliferation and induced apoptosis of CD34+ progenitors from healthy individuals and of cells from two myeloid lines (HL-60 and K562). C/EBPα, a key transcription factor in granulopoiesis5,6, was directly regulated by LEF-1. These observations indicate that LEF-1 is an instructive factor regulating neutrophilic granulopoiesis whose absence plays a critical role in the defective maturation program of myeloid progenitors in individuals with CN. NOTE: In the version of this article initially published, the DOI was incorrect. The correct DOI is 10.1038/nm1474. The error has been corrected in the HTML and PDF versions of the article.

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Figure 1: Expression of LEF-1 and LEF-1 target genes is abrogated in CN myeloid precursors and is upregulated in LEF-1–rescued CN CD34+ cells.
Figure 2: Restoration of defective LEF-1 expression promotes granulocytic differentiation of CN CD34+ progenitors.
Figure 3: Anti-proliferative and pro-apoptotic effects of LEF-1 inhibition in CD34+ progenitors.
Figure 4: LEF-1 overexpression in CD34+ cells promoted upregulation of LEF-1 target genes and increased cell proliferation.

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Change history

  • 19 October 2006

    In the version of this article initially published, the DOI was incorrect. The correct DOI is 10.1038/nm1474. The error has been corrected in the HTML and PDF versions of the article.

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Acknowledgements

We thank M. Morgan for critically reading the manuscript; V. Bucan and D. Lan for sequencing, G. Asgedom and A. Müller-Brechlin for excellent technical assistance, and M. Ballmaier and C. Reimer for assistance in cell sorting. We also thank the study subjects and their families for cooperation. This work was supported by the German Network “Congenital Bone Marrow Failure Syndromes” of the Federal Ministry of Education and Research (BMBF), Deutsche Forschungsgemeinschaft (SFB 566), Madeleine-Schickedanz-Kinderkrebsstiftung, and José Carreras Leukämie-Stiftung e.V.

Author information

Author notes

  1. Gunnar Cario

    Present address: Department of Pediatrics, University Hospital Schleswig-Holstein, Campus Kiel, 24105, Kiel, Germany

  2. Gunnar Cario and Murat Uenalan: These authors contributed equally to this work.

Authors and Affiliations

  1. Departments of Pediatric Hematology and Oncology, Carl-Neuberg-Str. 1, Hannover, 30625, Germany

    Julia Skokowa, Gunnar Cario, Murat Uenalan, Axel Schambach, Manuela Germeshausen, Cornelia Zeidler, Martin Stanulla & Karl Welte

  2. Hematology, Hemostaseology and Oncology, Carl-Neuberg-Str. 1, Hannover, 30625, Germany

    Axel Schambach, Karin Battmer, Matthias Eder, Christopher Baum & Michaela Scherr

  3. Pathology, Hannover Medical School, Carl-Neuberg-Str. 1, Hannover, 30625, Germany

    Ulrich Lehmann

  4. Department of Cellular and Molecular Immunology, Max Planck Institute of Immunobiology, Freiburg, 79108, Germany

    Rudolf Grosschedl

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Contributions

K.W., G.C., J.S. and M.St. made initial observations. J.S. designed and performed the main experiments, analyzed the data and wrote the manuscript. M.U. performed western blot analysis and wrote the manuscript. M.Sch., M.E. and K.B. designed shRNA constructs and transduced cells. A.S. and C.B. designed lv constructs that contained full-length LEF-1 or dnLEF-1 and produced viral supernatants. M.G. performed sequence analysis of ELA2 gene. C.Z. provided patients data. U.L. provided laser-assisted microscope. K.W. supervised experimentation, coordinated the project and wrote the manuscript.

Corresponding authors

Correspondence to Julia Skokowa or Karl Welte.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

mRNA expression of secretory granule proteins; LEF-1 mRNA/protein expression as well as TCF-3,-4 mRNA expression. (PDF 220 kb)

Supplementary Fig. 2

mRNA expression of indicated genes in sorted GFP− CD34+ CN cells from the LEF-1 lv and dnLEF-1 lv experiments. (PDF 26 kb)

Supplementary Fig. 3

mRNA expression of indicated genes in sorted RFP− cells from the shRNA transduction experiments. Morphology of LEF-1 and β-catenin shRNA transduced cells. (PDF 65 kb)

Supplementary Fig. 4

Effects of LEF-1 inhibition in two myeloid cell lines HL-60 and K562. (PDF 160 kb)

Supplementary Fig. 5

mRNA expression of indicated genes in sorted RFP− cells from the shRNA transduction experiments in K562 and HL-60 cell lines. (PDF 26 kb)

Supplementary Fig. 6

mRNA expression of indicated genes in sorted GFP− cells from the experiments in LEF-1 lv transduced CD34+ cells of healthy controls. (PDF 23 kb)

Supplementary Table 1

CN patients' characteristics and ELA2 mutations status. (PDF 15 kb)

Supplementary Table 2

Oligonucleotide sequences used for NoShift assay. (PDF 12 kb)

Supplementary Methods (PDF 106 kb)

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Skokowa, J., Cario, G., Uenalan, M. et al. LEF-1 is crucial for neutrophil granulocytopoiesis and its expression is severely reduced in congenital neutropenia. Nat Med 12, 1191–1197 (2006). https://doi.org/10.1038/nm1474

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