MEIS2 is a novel oncogenic partner in AML1-ETO positive AML [RNA-Seq mouse]
ABSTRACT: 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:The AML1/ETO fusion protein is essential to the development of acute myeloid leukemia (AML), and is well recognized for its dominant-negative effect on the co-existing wild-type protein AML1. However, the involvement of wild-type AML1 in AML1/ETO-driven leukemogenesis remains elusive. Through chromatin immunoprecipitation sequencing, computational analysis plus a series of experimental validations, we report here that AML1 is able to orchestrate the expression of AML1/ETO targets regardless of being activated or repressed, via forming a complex with AML1/ETO and via recruiting the cofactor. 4 ChIP-seq assays were used to identify the high confidence binding regions of AML1-ETO and AML1 in t(8;21) AML Kasumi-1 cell lines. The anti-AML1 (N20) antibody targets the N-terminus of AML1 and recognizes both AML1 and AML1/ETO; the anti-AML1 (C19) antibody targets the C-terminus of AML1 and recognizes AML1 but not AML1/ETO; the anti-ETO (C20) antibody targets the C-terminus of ETO and specifically recognizes AML1/ETO. 2 ChIP-seq assays were used to identify the binding regions of AML1 in human macrophage U937 cell lines. And the total input was used as control.
Project description:The MYB oncogene is widely expressed in acute leukemias and is important for the continued proliferation of leukemia cells, raising the possibility that MYB may be a therapeutic target. However realization of this potential requires (i) a significant therapeutic window for MYB inhibition, given its essential role in normal hematopoiesis; and (ii) an approach for developing an effective therapeutic. We previously showed that the interaction of Myb with the coactivator CBP/p300 is essential for its transforming activity. Here we use hematopoietic cells from the Booreana mouse strain, which carries a mutation in Myb that prevents interaction with CBP/p300, to examine the requirement for this interaction in myeloid transformation and leukemogenesis. Using this strain and a strain (plt6) carrying a “complementary” mutation in p300, we show that the Myb-p300 interaction is essential for in vitro transformation by the myeloid leukemia oncogenes AML1-ETO, AML1-ETO9a, MLL-ENL, and MLL-AF9. We further show that unlike cells from wild-type (WT) mice, Booreana cells fail to induce leukemia upon transplantation into irradiated recipients following transduction with an AML1-ETO9a retrovirus. These data highlight disruption of the Myb-p300 interaction as a potential therapeutic strategy for AML and suggest that such a strategy would have a useable therapeutic index since Booreana mice, unlike Myb null mice, are viable. Finally we have begun to explore the molecular basis of the these observations by gene expression profiling; this highlighted several genes previously implicated in myeloid leukemogenesis as being differentially expressed between WT and Booreana cells transduced with AML1-ETO9a. Total RNA was obtained from FACS sorted GFP+;c-Kit+ primary bone marrow cells from WT and Booreana mouse strains which had been cultured for 48 hours post-transduction with Control or AML1-ETO9a retroviruses. RNA was extracted from each of 4 samples per group and used to probe Illumina mouse Beadchips array.
Project description:MEIS2 collaborates with AML1-ETO in inducing acute myeloid leukemia in a murine bone marrow transplantation model RNA-seq to evaluate consequence of MEIS2 Knock-down on the transcriptional profle of the t(8;21) positive Kasumi-1 cell line
Project description:Microarray gene profilling indentified snoRNAs are downstream target of Amino Enhancer of Split (AES) and are essential for AML1-ETO9a induced leukemia. Amino Enhancer of Split (Aes) is strongly induced by leukemia oncogenes AML1-ETO, PML-RARα and PLZF-RARα. With a conditional AES knockout mouse model we showed that AES is essential for AML1-ETO9a indeced leukemia. We performed gene expression microarray using mouse primary AML1-ETO9a transformed AES wildtype and knockout and showed that snoRNAs were downregulated in AES knockout cells. We found that SnoRNA induction is a common mechanism shared by distinct oncogenes including AML1-ETO, MYC and MLL-AF9. Suppression of C/D box snoRNA complexes or deletion of several single C/D box snoRNAs inhibit clonogenic growth of leukemia cells. These findings suggest that enhancement of snoRNA levels is a critical mechanism of leukemic transformation. Overall design: AES wildtype or knockout AML1-ETO9a transformed cells in triplicates
Project description:MEIS2 collaborates with AML1-ETO in inducing acute myeloid leukemia in a murine bone marrow transplantation model Overall design: We employed ChIP-seq to assess the impact of MEIS2 knockdown on the DNA binding profile of AML1-ETO (Kasumi-1 cell line)
Project description:Compare the gene expression profile among human CD34+ cord blood cells infected with MIGR1, MIGR1-AML1-ETO or MIGR1-AML1-ETO∆NHR1 AML1-ETO promotes the self-renewal of human hematopoietic stem/progenitor cells (HSPCs). We found deletion of NHR1 domain abrogates AML1-ETO induced expasion of HSPCs. GFP+CD34+ human cord blood cells were sorted by FACS 72 hours after the infection for RNA extraction and hybridyzation for Affymetrix microarrays.
Project description:Kasumi-1 AML cells that were transfected in triplicate with AML1-ETO or luciferase siRNA constructs by either Amaxa nucleofection or Biorad siLentFect and incubated for 96 hours. Microarrays used to discover an AML1-ETO signature for a GE-HTS screen to identify AML1-ETO modulators. Experiment Overall Design: Kasumi-1 AML cells incubated for 96 hours after they were transfected in triplicate with AML1-ETO or luciferase siRNA constructs by either Amaxa nucleofection or Biorad siLentFect along with three control samples not transfected with a construct.
Project description:U937 AML cells that express an inducible AML1-ETO construct under the control of the tetracycline promoter. Microarrays used to discover an AML1-ETO signature for a GE-HTS screen to identify AML1-ETO modulators. Experiment Overall Design: U937 AML cells that express an inducible AML1-ETO construct under the control of the tetracycline promoter at 0, 12, 24, 48, and 72 hours and corresponding controls (2 replicates each).
Project description:This SuperSeries is composed of the following subset Series:; GSE15646: Kasumi-1 AML1-ETO knockdown samples; GSE15647: U937 AML1-ETO inducible samples Experiment Overall Design: Refer to individual Series