Wnt Signaling in Hematopoietic Stem Cell Specification and Leukemia

Summary

Principal Investigator: WILSON KENDRICK CLEMENTS
Abstract: Project Summary/Abstract Hematopoietic stem cells (HSCs) are progenitor cells that have the ability to both self-renew and regenerate all mature blood cell types, including red blood cells and immune cells over the lifetime of an individual. HSCs are used therapeutically in the treatment of numerous diseases including leukemia and congenital blood disorders, but obtaining suitable numbers of histocompatible cells for transplantation remains a problem. Determining how embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) are directed to become tissue specific stem cells such HSCs is a key goal of regenerative medicine. The most obvious approach to defining required regulatory networks is to determine the endogenous mechanisms used during embryonic development. Often, involved signal transduction pathways are also observed to be dysregulated in leukemia, making an understanding of their basic biology of further clinical relevance. I have recently shown that "non- canonical", [unreadable]-catenin/Tcf-independent signaling by the Wnt pathway, which was not previously known to be involved in HSC specification, is required for formation HSCs. The Wnt ligand, Wnt16, which is conserved across vertebrate phyla and was originally identified as a gene aberrantly upregulated in pre-B acute lymphocytic leukemia (ALL), is required for specification of the first HSCs. My preliminary results show that Wnt16 activates expression of two Notch ligands, deltaC and deltaD, and that these ligands are in turn required redundantly for HSC specification. Although cell-autonomous reception of a Notch signal in cells fated to become HSCs is an established requirement in vertebrates, the Notch signaling events regulated by DeltaC and DeltaD appear to be non-cell-autonomous and therefore represent a distinct, previously unappreciated requirement for Notch signaling. In the research proposed here, I will seek to determine the precise signal transduction pathway(s) that lie between Wnt16 and transcriptional activation of deltaC and deltaD, by identifying the required co-receptors and intracellular signal transduction proteins. Preliminary data suggest that absence of DeltaC and DeltaD leads to defects in the formation or behavior of a somite compartment, the sclerotome, which is adjacent to the primitive dorsal aorta, the tissue that gives rise to the first HSCs in vertebrates. During the time when endothelium of the dorsal aorta becomes hemogenic and commits to an HSC fate, sclerotomally derived cells emigrate from the somite to become smooth muscle cells surrounding the aorta and may also directly contribute "replacement" endothelium, suggesting that sclerotomal defects underlie failure of HSC specification. Using zebrafish, which are transparent during embryonic development and uniquely receptive to transgenesis, allowing direct visualization of fluorescently labeled tissues, I will generate transgenic animals with labeled sclerotome, to determine how this tissue behaves in normal and Wnt16/Notch-deficient animals. I will test the overall requirement for sclerotome in development by conditional ablation, and the requirement for specific proteins in sclerotomal cells by conditional expression of wild-type and dominant negative factors. The non-canonical Wnt receptor, Ryk appears to participate in Wnt16 signal transduction, but cannot explain all of the effects observed in Wnt16 deficient animals. These results suggest the presence of additional co-receptors. The strongest family of candidates for this co-receptor is the Ror family of non- canonical Wnt receptors. The C. elegans ortholog of Wnt16, EGL-20 interacts physically and functionally with the single worm Ror ortholog, CAM-1. Zebrafish have three Ror family members, MuSK, Ror1, and Ror2. I have determined that Ror2 is not the required Wnt16 co-receptor, and the phenotype of the unplugged mutant, which carries a null allele of the musk gene, suggests that MuSK is also unlikely to contribute to the Wnt16 hematopoietic phenotype. Thus, Ror1 is the strongest candidate for the Wnt16 co-receptor. Interestingly, ROR1 misexpression is strongly associated with chronic lympocytic leukemia (CLL) as well as some ALL. WNT16 is also misexpressed in forms of ALL and CLL. Taken together with the fact that tissue culture experiments suggest that WNT16 is causally involved in pre-B-ALL, these results suggest that WNT16 and ROR1 may cooperatively or independently contribute to leukemogenesis. To test these possibilities, I will generate transgenic animals in which wnt16, ror1, and the oncogene E2A-PBX1, which has previously been associated with WNT16-directed disease progression in pre-B-ALL, are expressed in B-cells at a variety of maturation stages. Since no initiating lesions are known for CLL, these models have the potential to be extremely informative. Finally, I will use the B-cell transgenic animals as a platform for unbiased discovery of additional mutations that are involved in B-cell leukemia by forward genetics.
Funding Period: 2011-09-01 - 2015-05-31
more information: NIH RePORT

Top Publications

  1. pmc Signalling pathways that control vertebrate haematopoietic stem cell specification
    Wilson K Clements
    Department of Hematology, Division of Experimental Hematology, St Jude Children s Research Hospital, 262 Danny Thomas Pl, Memphis, Tennessee 38105, USA
    Nat Rev Immunol 13:336-48. 2013

Detail Information

Publications1

  1. pmc Signalling pathways that control vertebrate haematopoietic stem cell specification
    Wilson K Clements
    Department of Hematology, Division of Experimental Hematology, St Jude Children s Research Hospital, 262 Danny Thomas Pl, Memphis, Tennessee 38105, USA
    Nat Rev Immunol 13:336-48. 2013
    ..Here, we review current knowledge of the intraembryonic signals required for the specification of HSCs in vertebrates...

Research Grants30

  1. HEMANGIOBLAST DEVELOPMENT AND REGULATION
    Kyunghee Choi; Fiscal Year: 2013
    ..Thus, positive outcome from the current proposed studies would be directly applicable to regenerative medicine utilizing pluripotent stem cells in the future. ..
  2. Defining Estrogenic Regulation of Hematopoietic Stem Cell Formation and Function
    Trista E North; Fiscal Year: 2013
    ..Using murine assays, they will examine how these compounds effect HSC function across vertebrate species. ..
  3. Integrated signaling of eicosanoids in HSC formation and regeneration
    Trista E North; Fiscal Year: 2013
    ..These results are expected to reveal deeper insight into interactive roles of inflammatory mediators in marrow regeneration and have relevance for combinatorial targeting to direct clinical outcome in transfusion biology. ..
  4. Molecular and functional dissection of the zebrafish hematopoietic stem cell nich
    David Traver; Fiscal Year: 2013
    ..With the elucidation of this signaling axis, and the cellular behaviors it controls, our work will ultimately enable ex vivo approaches to direct patient-specific iPS cells towards the HSC fate for cellular replacement therapies. ..
  5. Herpesviral, Oncogenesis, Latency and Reactivation
    NANCY JOAN RAAB-TRAUB; Fiscal Year: 2013
    ..abstract_text> ..
  6. UNMC EPPLEY CANCER CENTER SUPPORT GRANT
    Kenneth H Cowan; Fiscal Year: 2013
    ....
  7. Center for Neuroplasticity at the University of Puerto Rico
    Steven N Treistman; Fiscal Year: 2013
    ..This UPR COBRE Center should define pathways and benchmarks for basic and translational research across the UPR system for the next decades. ..
  8. Cadiorenal and Metabolic Diseases Research Center
    John E Hall; Fiscal Year: 2013
    ..abstract_text> ..
  9. MLL in Hematopoiesis and Leukemia in the Zebrafish Model
    Carolyn A Felix; Fiscal Year: 2013
    ..The exploitation of zebrafish to model the functions of human MLL in the regulation of normal and malignant blood cell development will streamline future inroads to new molecularly targeted therapies for patients with MLL leukemia. ..
  10. Role of the zinc finger transcription factor ZBP89 in blood &vascular developmen
    M Amin Arnaout; Fiscal Year: 2013
    ..We have discovered a transcription factor that lies at the cross road of blood and blood vessel development. We are proposing a series of studies to pinpoint the role of this factor, and define the developmental networks it coordinates. ..
  11. HORMONAL REGULATION OF BLOOD PRESSURE
    Michal Laniado Schwartzman; Fiscal Year: 2013
    ..ular tone, in the pathophysiology of hypertension and cardiovascular disease. ..
  12. Arterial Dysfunction: Basic and Clinical Mechanisms
    Thomas Michel; Fiscal Year: 2013
    ..Gladyshev. P. Libby directs the Redox Biomarkers Core;metabolic characterizations of mouse models studied in this Program will take place at the Yale Mouse Metabolic Phenotyping Center, led by G. Shulman. ..
  13. Molecular Pathways to Thynmic Lymphoma and Leukemia
    A Thomas Look; Fiscal Year: 2013
    ..Such interactions are expected to accelerate the pace at which important discoveries are generated in these projects and in the program as a whole. ..
  14. HSC Diversity: Regulation by Clonal Selection vs Epigenetic Induction
    Irving L Weissman; Fiscal Year: 2013
    ....