During the last year, we continued define the molecular events that regulate hematopoietic stem cell (HSC) quiescence, survival, self-renewal and, myeloid cell lineage commitment and differentiation. We have focused our efforts on transcription factors since they are essential for stem cell growth and differentiation, and are frequently deregulated during the development of leukemias and lymphomas. We previously found that the helix loop helix (HLH) transcription factor, inhibitor of differentiation 1 (Id1), is induced during the early stages of myeloid development, and can instruct hematopoietic stem cells toward a myeloid versus erythroid and lymphoid cell fate. Furthermore, we discovered that deregulated expression of Id1 immortalizes hematopoietic progenitor cells in vitro, and leads a myeloproliferative disease (MPD) in vivo. We found that preventing Id1 expression using Id1 siRNA inhibited human leukemic cell line growth, suggesting that Id1 may represent a viable therapeutic target to treat some blood cancers. Future studies are planned to determine if we can inhibit or promote the differentiation of acute myelogenous leukemia patient samples in vitro and in vivo when transplanted into severe combined immuno-deficient (SCID) mice. We have extended these studies to Id1-/- mice to determine if Id1 is required for normal hematopoietic development. Mice that lack Id1 gene expression are viable and show no obvious defects. However, these mice show a hematopoietic phenotype, which includes increased hematopoietic stem/progenitor cell cycling and defects in lineage development (decreased B cells and increased myeloid cells). We transplanted Id1-/- bone marrow cells (BMC) into wild type (WT) recipient mice, and determined that the Id1-/- BMC developed normally in these mice. Therefore, the hematopoietic phenotype observed in the Id1-/- mice was not intrinsic to the Id1-/- HSC, but were defects in the hematopoietic micro environment. This was confirmed by transplanting WT BMC into irradiated Id1-/- recipients, where we found that the WT BMC show impaired hematopoietic development in an Id1-/- micro environment. Furthermore, we determine that stromal cells that lack Id1-/- do not support normal hematopoietic development in long term bone marrow cultures in vitro, suggesting that the differentiation or function of the stromal cell compartment is impaired by the loss of Id1 gene expression. Initial experiments to determine if loss of Id1 gene expression affects the production of critical hematopoietic growth factors, demonstrate that some the production of some HGF (CSF-1 and G-CSF) are affected, which could explain, in part, the hematopoietic phenotype in Id1-/- mice. Future experiments are planned to delete Id1 gene expression specifically in endothelial cells, osteoblasts and other stromal cell populations in vivo using conditional Id1 mice and cre-recombinase expressing transgenic mice. In addition, experiments are planned to determine if loss of Id1 gene expression in the micro environment contributes to the tumor cell niche.




In an effort to identify novel transcriptional regulator of the early stages of myeloid cell growth and differentiation, we have differentially analyzed the global gene expression profile of differentiating hematopoietic progenitor cells. We have identified a novel zinc finger transcription factor of unknown function, POGZ, which is down regulated during the early stages of myeloid development. We have constructed a POGZ retroviral vector to determine if over expression of POGZ affects hematopoietic development by transducing hematopoietic stem/progenitor cells and evaluating their growth and differentiationin vitro, andin vivo in vivo in bone marrow transplantation assays. We have generated a mouse strain with a targeted deletion of POGZ. POGZ-/- mice do not survive beyond the first few hours of life, and die at birth of unknown causes, suggesting that POGZ is essential for mouse survival. Future studies are planned to evaluate the hematopoietic defects in these mice

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Intramural Research (Z01)
Project #
1Z01BC010001-12
Application #
7592614
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
12
Fiscal Year
2007
Total Cost
$1,026,735
Indirect Cost
Name
National Cancer Institute Division of Basic Sciences
Department
Type
DUNS #
City
State
Country
United States
Zip Code
Asefa, Benyam; Dermott, Jonathan M; Kaldis, Philipp et al. (2006) p205, a potential tumor suppressor, inhibits cell proliferation via multiple pathways of cell cycle regulation. FEBS Lett 580:1205-14
Suh, Hyung Chan; Gooya, John; Renn, Katie et al. (2006) C/EBPalpha determines hematopoietic cell fate in multipotential progenitor cells by inhibiting erythroid differentiation and inducing myeloid differentiation. Blood 107:4308-16
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Berthet, Cyril; Klarmann, Kimberly D; Hilton, Mary Beth et al. (2006) Combined loss of Cdk2 and Cdk4 results in embryonic lethality and Rb hypophosphorylation. Dev Cell 10:563-73
Drutskaya, Marina S; Ortiz, Mariestela; Liepinsh, Dmitry J et al. (2005) Inhibitory effects of tumor necrosis factor on hematopoiesis seen in vitro are translated to increased numbers of both committed and multipotent progenitors in TNF-deficient mice. Exp Hematol 33:1348-56
Leeanansaksiri, Wilairat; Wang, Hui; Gooya, John M et al. (2005) IL-3 induces inhibitor of DNA-binding protein-1 in hemopoietic progenitor cells and promotes myeloid cell development. J Immunol 174:7014-21
Li, Wen Qing; Jiang, Qiong; Khaled, Annette R et al. (2004) Interleukin-7 inactivates the pro-apoptotic protein Bad promoting T cell survival. J Biol Chem 279:29160-6
Asefa, Benyam; Klarmann, Kimberly D; Copeland, Neal G et al. (2004) The interferon-inducible p200 family of proteins: a perspective on their roles in cell cycle regulation and differentiation. Blood Cells Mol Dis 32:155-67
Dermott, Jonathan M; Gooya, John M; Asefa, Benyam et al. (2004) Inhibition of growth by p205: a nuclear protein and putative tumor suppressor expressed during myeloid cell differentiation. Stem Cells 22:832-48
Heath, Victoria; Suh, Hyung Chan; Holman, Matthew et al. (2004) C/EBPalpha deficiency results in hyperproliferation of hematopoietic progenitor cells and disrupts macrophage development in vitro and in vivo. Blood 104:1639-47

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