title: FLT3 activation cooperates with MLL-AF4 fusion protein to abrogate the hematopoietic specification of human ESCs.
abstract: Mixed-lineage leukemia (MLL)-AF4 fusion arises prenatally in high-risk infant acute pro-B-lymphoblastic leukemia (pro-B-ALL). In human embryonic stem cells (hESCs), MLL-AF4 skewed hematoendothelial specification but was insufficient for transformation, suggesting that additional oncogenic insults seem required for MLL-AF4-mediated transformation. MLL-AF4+ pro-B-ALL expresses enormous levels of FLT3, occasionally because of activating mutations, thus representing a candidate cooperating event in MLL-AF4+ pro-B-ALL. Here, we explored the developmental impact of FLT3 activation alone, or together with MLL-AF4, in the hematopoietic fate of hESCs. FLT3 activation does not affect specification of hemogenic precursors but significantly enhances the formation of CD45(+) blood cells, and CD45(+)CD34(+) blood progenitors with clonogenic potential. However, overexpression of FLT3 mutations or wild-type FLT3 (FLT3-WT) completely abrogates hematopoietic differentiation from MLL-AF4-expressing hESCs, indicating that FLT3 activation cooperates with MLL-AF4 to inhibit human embryonic hematopoiesis. Cell cycle/apoptosis analyses suggest that FLT3 activation directly affects hESC specification rather than proliferation or survival of hESC-emerging hematopoietic derivatives. Transcriptional profiling of hESC-derived CD45(+) cells supports the FLT3-mediated inhibition of hematopoiesis in MLL-AF4-expressing hESCs, which is associated with large transcriptional changes and downregulation of genes involved in hematopoietic system development and function. Importantly, FLT3 activation does not cooperate with MLL-AF4 to immortalize/transform hESC-derived hematopoietic cells, suggesting the need of alternative (epi)-genetic cooperating hits.
authors.affiliation: Centre for Genomics and Oncological Research, Pfizer-University of Granada-Andalusian Regional Government, Granada, Spain. clara.bueno@genyo.es
authors.forename: Clara
authors.initials: C
authors.lastname: Bueno
authors.forename: Verónica
authors.initials: V
authors.lastname: Ayllón
authors.forename: Rosa
authors.initials: R
authors.lastname: Montes
authors.forename: Oscar
authors.initials: O
authors.lastname: Navarro-Montero
authors.forename: Verónica
authors.initials: V
authors.lastname: Ramos-Mejia
authors.forename: Pedro J
authors.initials: PJ
authors.lastname: Real
authors.forename: Damià
authors.initials: D
authors.lastname: Romero-Moya
authors.forename: Marcos J
authors.initials: MJ
authors.lastname: Araúzo-Bravo
authors.forename: Pablo
authors.initials: P
authors.lastname: Menendez
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000235:genetics*; D019070:Cell Lineage / Q000235:genetics; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000378:metabolism / Q000502:physiology*; D004789:Enzyme Activation / Q000502:physiology; D020869:Gene Expression Profiling; D006410:Hematopoiesis / Q000235:genetics*; D006412:Hematopoietic Stem Cells / Q000378:metabolism / Q000502:physiology; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D046228:Microarray Analysis; D051788:Myeloid-Lymphoid Leukemia Protein / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D051941:fms-Like Tyrosine Kinase 3 / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: C113855:MLL-AF4 fusion protein, human; D015514:Oncogene Proteins, Fusion; D051788:Myeloid-Lymphoid Leukemia Protein; C497970:FLT3 protein, human; D051941:fms-Like Tyrosine Kinase 3