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Interactions among HCLS1, HAX1 and LEF-1 proteins are essential for G-CSF–triggered granulopoiesis

Nature Medicine volume 18, pages 15501559 (2012) | Download Citation


We found that hematopoietic cell–specific Lyn substrate 1 (HCLS1 or HS1) is highly expressed in human myeloid cells and that stimulation with granulocyte colony-stimulating factor (G-CSF) leads to HCLS1 phosphorylation. HCLS1 binds the transcription factor lymphoid-enhancer binding factor 1 (LEF-1), transporting LEF-1 into the nucleus upon G-CSF stimulation and inducing LEF-1 autoregulation. In patients with severe congenital neutropenia, inherited mutations in the gene encoding HCLS1-associated protein X-1 (HAX1) lead to profound defects in G-CSF–triggered phosphorylation of HCLS1 and subsequently to reduced autoregulation and expression of LEF-1. Consistent with these results, HCLS1-deficient mice are neutropenic. In bone marrow biopsies of the majority of tested patients with acute myeloid leukemia, HCLS1 protein expression is substantially elevated, associated with high levels of G-CSF synthesis and, in some individuals, a four-residue insertion in a proline-rich region of HCLS1 protein known to accelerate intracellular signaling. These data demonstrate the importance of HCLS1 in myelopoiesis in vitro and in vivo.

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We thank A. Gigina, K. Cherkaoui, A. Müller Brechlin, M. Reuter and A.-L. Hagemann for technical assistance. We thank J. Klupp for assistance in generation of LEF-1 Ala16 mutant; D.D. Billadeau (Department of Immunology, Schulze Center for Novel Therapeutics, College of Medicine, Mayo Clinic) for providing us with the rabbit polyclonal HCLS1-specific antibody; L. Naldini (San Raffaele Telethon Institute for Gene Therapy, Division of Regenerative Medicine, Gene Therapy and Stem Cells, San Raffaele Institute) for pRRL.PPT.PGK.GFPpre vector; A. Schambach (Department of Experimental Hematology, Hannover Medical School) for VSVg envelope plasmid; Thomas J. Hope (University of Illinois at Chicago) for pRSV-Rev plasmid; and O. Tetsu (Department of Otolaryngology—Head and Neck Surgery, University of California–San Francisco) for the CCND1 reporter construct. We also thank the physicians within the Severe Chronic Neutropenia International Registry for providing patient material. We thank study subjects for their cooperation. This work was partially supported by German José Carreras Leukemia Foundation (J.S., M.K.), by REBIRTH Cluster of Excellence of the Hannover Medical School (J.S.), by Madeleine Schickedanz-KinderKrebs-Stiftung (J.S.), and by the Deutsche Forschungsgemeinschaft (Z.L.; DFG grant Li 1608/2-1).

Author information

Author notes

    • Maxim Klimiankou
    •  & Olga Klimenkova

    These authors contributed equally to this work.


  1. Department of Molecular Hematopoiesis, Hannover Medical School, Hannover, Germany.

    • Julia Skokowa
    • , Maxim Klimiankou
    • , Olga Klimenkova
    • , Dan Lan
    • , Kshama Gupta
    • , Inna Kusnetsova
    • , Cornelia Zeidler
    •  & Karl Welte
  2. Department of Pediatrics, The First Affiliated Hospital of GuangXi Medical University, NanNing, China.

    • Dan Lan
  3. Department of Pathology, Hannover Medical School, Hannover, Germany.

    • Kais Hussein
    •  & Hans-Heinrich Kreipe
  4. Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia and University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA.

    • Esteban Carrizosa
    •  & Janis Burkhardt
  5. Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany.

    • Zhixiong Li
  6. Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany.

    • Arnold Ganser
  7. Max Planck Institute of Immunobiology, Department of Cellular and Molecular Immunology, Freiburg, Germany.

    • Claudio Sustmann
    •  & Rudolf Grosschedl


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K.W. and J.S. made initial observations, designed the experiments, analyzed the data, supervised experimentation and wrote the manuscript; J.S., D.L., M.K., O.K., A.G. and I.K. performed the main experiments; K.G. introduced mutations into LEF-1 and C/EBPα reporter gene constructs and performed reporter gene assays in HEK293T cells; J.B. and E.C. performed blood cell counting in Hcls1−/− mice and provided bone marrow and blood material of these mice; K.H. and H.-H.K. performed tissue microarray analysis; Z.L. and A.G. provided some of the AML samples; C.S. and R.G. introduced mutations in LEF-1 cDNA corresponding to HCLS1-binding site; C.Z. provided patient material.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Julia Skokowa or Karl Welte.

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    Microarray data of CD34+ cells transduced with HCLS1 shRNA vs ctrl shRNA

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