ABSTRACT
Cytokine signaling pathways are frequent targets of oncogenic mutations in acute myeloid leukemia (AML), promoting proliferation and survival. We have previously shown that the transcription factor PLAGL2 promotes proliferation and cooperates with the leukemia fusion protein Cbfß-SMMHC in AML development. Here, we show that PLAGL2 upregulates expression of the thrombopoietin receptor Mpl, using two consensus sites in its proximal promoter. We also show that Mpl overexpression efficiently cooperates with Cbfß-SMMHC in development of leukemia in mice. Finally, we demonstrate that PlagL2-expressing leukemic cells show hyper-activation of Jak2 and downstream STAT5, Akt and Erk1/2 pathways in response to Thpo ligand. These results show that PlagL2 expression activates expression of Mpl in hematopoietic progenitors, and that upregulation of wild-type Mpl provides an oncogenic signal in cooperation with CBFß-SMMHC in mice.
Subject(s)
DNA-Binding Proteins/physiology , Hematopoietic Stem Cells/metabolism , Leukemia, Myeloid, Acute/metabolism , RNA-Binding Proteins/physiology , Receptors, Thrombopoietin/genetics , Signal Transduction , Transcription Factors/physiology , Transcription, Genetic , Animals , Base Sequence , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Bone Marrow Transplantation , Cells, Cultured , Electrophoretic Mobility Shift Assay , Extracellular Signal-Regulated MAP Kinases/genetics , Extracellular Signal-Regulated MAP Kinases/metabolism , Female , Flow Cytometry , Gene Expression Profiling , Humans , Immunoblotting , Janus Kinase 2/genetics , Janus Kinase 2/metabolism , Leukemia, Myeloid, Acute/genetics , Luciferases/metabolism , Male , Mice , Mice, Transgenic , Molecular Sequence Data , Oligonucleotide Array Sequence Analysis , Oncogene Proteins, Fusion/physiology , Promoter Regions, Genetic/genetics , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , RNA, Messenger/genetics , Receptors, Thrombopoietin/metabolism , Reverse Transcriptase Polymerase Chain Reaction , STAT5 Transcription Factor/genetics , STAT5 Transcription Factor/metabolism , Sequence Homology, Nucleic Acid , TransfectionABSTRACT
Acute myeloid leukemia subtype M4 with eosinophilia is associated with a chromosome 16 inversion that creates a fusion gene CBFB-MYH11. We have previously shown that CBFB-MYH11 is necessary but not sufficient for leukemogenesis. Here, we report the identification of genes that specifically cooperate with CBFB-MYH11 in leukemogenesis. Neonatal injection of Cbfb-MYH11 knock-in chimeric mice with retrovirus 4070A led to the development of acute myeloid leukemia in 2-5 months. Each leukemia sample contained one or a few viral insertions, suggesting that alteration of one gene could be sufficient to synergize with Cbfb-MYH11. The chromosomal position of 67 independent retroviral insertion sites (RISs) was determined, and 90% of the RISs mapped within 10 kb of a flanking gene. In total, 54 candidate genes were identified; six of them were common insertion sites (CISs). CIS genes included members of a zinc finger transcription factors family, Plag1 and Plagl2, with eight and two independent insertions, respectively. CIS genes also included Runx2, Myb, H2T24, and D6Mm5e. Comparison of the remaining 48 genes with single insertion sites with known leukemia-associated RISs indicated that 18 coincide with known RISs. To our knowledge, this retroviral genetic screen is the first to identify genes that cooperate with a fusion gene important for human myeloid leukemia.
