Project description:To gain the underlying insight into the functional impact of AML1-ETO expressed in HSPCs, RNA-seq was performed on purified LT-HSCs and GMPs from the bone marrow (BM) of both AML1-ETO expressed (AML1/ETO) and Wild-Type (control) C57 mice. Our work suggested that AML1-ETO resulted in impaired hematopoietic reconstitution and increased self-renewal ability.The oxidative phosphorylation and glycolysis decreased significantly in AML1/ETO LT-HSCs accompanied by increased HSC quiescence and reduced cell cycling. Furthermore, it was observed that HSCs expressing AML1-ETO exhibited an increased requirement for fatty acids when they differentiated.
Project description:In an effort to identify novel drugs targeting fusion-oncogene induced acute myeloid leukemia (AML), we performed high-resolution proteomic analysis. In AML1-ETO (AE) driven AML we uncovered a deregulation of phospholipase C (PLC) signaling. We identified PLCgamma 1 (PLCG1) as a specific target of the AE fusion protein which is induced after AE binding to intergenic regulatory DNA elements. Genetic inactivation of PLCG1 in murine and human AML inhibited AML1-ETO dependent self-renewal programs, leukemic proliferation, and leukemia maintenance in vivo. In contrast, PLCG1 was dispensable for normal hematopoietic stem- and progenitor cell function. These findings are extended to and confirmed by pharmacologic perturbation of Ca++-signaling in AML1-ETO AML cells, indicating that the PLCG1 pathway poses an important therapeutic target for AML1-ETO positive leukemic stem cells.
Project description:AE-expressing murine BM cells treated with all-trans retinoic acid (ATRA) in semi-solid methycellulose-based cultures show an increase in self-renewal capacity whilst treatment with a specific RARa agonist NRX195183 reduces their clonogenicity. Gene expression analysis was performed to further investigate the molecular mechanisms underlying these observations. Upregulated gene sets were identified in the ATRA-treated AE BM cells. 3 C57Bl/6 AML1-ETO-stop/+/Mx-Cre+ mice (#D203, 218 and 220) were treated with polyI:C to excise the stop codon allowing AML1-ETO expression. 2-4 weeks following completion of polyI:C administration, mice were euthanised and BM cells harvested. BM cells were cultured in semi-solid methylcellulose assays with cytokines for 2 weeks; then cells were harvested, pooled and replated in suspension cultures containing cytokines and the respective treatments - DMSO (control), ATRA and the specific RARa agonist NRX195183. At the 8 hour (8H) and 24 hour (24H) timepoints, cells were harvested for RNA extraction and processing.
Project description:MEIS2 collaborates with AML1-ETO in inducing acute myeloid leukemia in a murine bone marrow transplantation model We employed RNA-seq to assess similarities/differences among murine leukemic bone marrow samples transduced with either AML1-ETO/Meis2, AML1-ETO9a/Meis2, or AML1-ETO9a
Project description:MEIS2 collaborates with AML1-ETO in inducing acute myeloid leukemia in a murine bone marrow transplantation model We compared Gene expression profile (GEP) of murine bone marrow cells transduced with GFP, AML1-ETO, MEIS2, and AML1-ETO/MEIS2. Data from MEIS1 and AML1-ETO/MEIS1 is also included. Mouse bone marrow cells were kept in culture for 48hrs after retroviral transduction. GFP positive cells were then sorted and cells were kept for further 24 hours in culture before microarray analysis.
Project description:To gain the underlying insight into the functional impact of AML1-ETO expressed in hematopoietic stem cells and progenitors (HSPCs), we employed single-cell transcriptome sequencing to elucidate the characteristics of purified Lin-c-kit+ cells (HSPCs) from the bone marrow (BM) of both AML1-ETO expressed (AML1/ETO) and Wild-Type (control) C57 mice.
