ABSTRACT: Transcription profiles of WT and CreER-Raptor-flox/flox common myeloid progenitors (CMP), granulocyte-macrophage progenitor (GMP), or lineage negative (Lin-) stimulated with M-CSF for 12 hours
Project description:Myelopoiesis is impaired in Raptor-deleted mice (CreER-Rptor-flox/flox). To evaluate the transcriptional changes in myeloid precursors , we isolated CMP (Lin–Sca-1–c-Kit+CD34+FcγRII/IIImid), GMP (Lin–Sca-1–c-Kit+CD34+FcγRII/IIIhigh) and Lin (B220, Ly6C, Ly6G, CD3, Ter-119) negative cells (Lin–) from bone marrow of WT or CreER-Rptor-flox/flox mice. RNA was isolated from CMP and GMP immediately after sorting and Lin- cells were cultured for 12 hours with M-CSF (10 ng/mL) in 10% FBS and 1% P/S DMEM before RNA isolation. We used microarrays to compare the global transcription profiles of CMP, GMP, or Lin– stimulated with M-CSF for 12 hours from WT and CreER-Raptor-flox/flox mice.
Project description:To identify the molecular characterisitics of parallel lineage-biased MPP populations arising from hematopoietic stem cells (HSC) we conducted genome-wide analyses of hematopoietic stem, progenitor and mature myeloid cell populations using Affymetrix Gene ST1.0 arrays. Microarray analysis of 3-5 biological replicates of the indicated hematopoietic populations, isolated by FACS sorting from C57BL/6 mouse BM. Immunophenotypic definitions: Long-term HSC (HSCLT) (Lin-/cKit+/Sca1+/Flk2-/CD48-/CD150+); Short-term HSC (HSCST) (Lin-/cKit+/Sca1+/Flk2-/CD48-/CD150-); MPP2 (Lin-/cKit+/Sca1+/Flk2-/CD48+/CD150+); MPP3 (Lin-/cKit+/Sca1+/Flk2-/CD48+/CD150-); MPP4 (Lin-/cKit+/Sca1+/Flk2+); CMP (Lin-/cKit+/Fc?R-/CD34+); GMP (Lin-/cKit+/Fc?R+/CD34+); Pre-granulocyte (PreGr) (Mac1+/Gr1int); Granulocyte (Gr) (Mac1+/Gr1hi). HSC and GMP samples listed here were also used as controls for our related microarray study GSE48893.
Project description:The common myelo-erythroid progenitor (CMP) was subdivided based on expression of CD27. To understand the relationship between the various CD27 fractions of the CMP and downstream progenitors (granulocyte-monocyte progenitor, GMP, and megakaryocyte-erythroid progenitor, MEP), genome-wide transcriptional analysis was used.
Project description:Analysis of genes induced in DC precursors and in BM cells and monocytes treated with GM-CSF For progenitor arrays, bone marrow progenitors (CMP, GMP, CDP, and pre-cDC) were harvested from WT C57Bl/6 mice. For culture arrays, BM was cultured in the presence of GM-CSF or M-CSF and adherent and non-adherent cells sorted. For monocyte cultures, sorted BM monocytes were treated with GM-CSF for 0, 24 or 48 hours.
Project description:We compared single-cell transcriptomes of fetal liver myeloid progenitor pools (CMP/GMP pool) from Gata2 -77 enhancer mutant embryos and wildtype littermates. Gene expression patterns in -77-deficient mice demonstrated loss of erythroid, megakaryocyte and granulocyte progenitors and an increase in monocyte
Project description:The study was a comparison of gene expression using RNA-seq. We analyzed the stem and progenitor cells from WT and Vav-cre+ Tet2fl/fl Flt3-ITD (T2F3) mice. We isolated stem cells LSK (lin- sca+ kit+) and granulocyte-macrophage progenitors GMP (lin- sca- kit+ fcgr+ cd34+) cells from bone marrow. Comparisons were made across genotypes WT vs. T2F3 and cell types LSK vs. GMP.
Project description:The study was a comparison of gene expression using RNA-seq. We analyzed the stem and progenitor cells from WT and Vav-cre+ Tet2fl/fl Flt3-ITD (T2F3) mice. We isolated stem cells LSK (lin- sca+ kit+) and granulocyte-macrophage progenitors GMP (lin- sca- kit+ fcgr+ cd34+) cells from bone marrow. Comparisons were made across genotypes WT vs. T2F3 and cell types LSK vs. GMP. Comparison of WT and Tet2-/-Flt3ITD bone marrow stem and progenitor cells.
Project description:The pathways by which oncogenes, such as MLL-AF9, initiate transformation and leukemia in humans and mice are incompletely defined. In a study of target cells and oncogene dosage, we found that Mll-AF9, when under endogenous regulatory control, efficiently transformed LSK (Lin- Sca1+ c-kit+) stem cells while committed granulocyte-monocyte progenitors (GMPs) were transformation-resistant and did not cause leukemia. Mll-AF9 was expressed at higher levels in hematopoietic stem (HSC) than GMP cells. Mll- AF9 gene dosage effects were directly shown in experiments where GMPs were efficiently transformed by the high dosage of Mll-AF9 resulting from retroviral transduction. Mll-AF9 up-regulated expression of 196 genes in both LSK and progenitor cells, but to higher levels in LSKs than in committed myeloid progenitors. Experiment Overall Design: Comparison of gene expression profiles among four types of hematopoietic cells (GMP, CMP, CLP and HSC), FACS sorted from wild type and Mll-AF9 knock-in mice. The goal was to identify genes differentially expressed in each Mll-AF9 cell type compared to the corresponding wild type cells.
Project description:Tamoxifen was administered to Glast-CreER;Nf1-flox/flox;Tp53-flox/flox or Glast-CreER;Ascl1-flox/flox;Nf1-flox/flox;Tp53-flox/flox at E14.5 to induce tumors in the brain
Project description:We investigated the role of mTORC1 in murine hematopoiesis by conditionally deleting the Raptor gene in murine hematopoietic stem cells. We observed mutliple alterations evoked by Raptor loss in hematopoiesis and profiled gene-expression alterations induced by raptor loss in Flt3-Lin-Sca1+cKit+ hematopoietic stem and progenitor enriched cell populations, 5 weeks post Raptor deletion. Flt3-Lin-Sca1+cKit+ cells were flow sorted from mice containing homozygous floxed alleles for exon 6 of the Raptor gene in the presence (MT group) or absence (WT group) of the MxCre transgene, which was induced with injections of mice with pIpC 5 weeks before cell isolation.
