FLT3 activation cooperates with MLL-AF4 fusion protein to abrogate the hematopoietic specification of human ESCs

Blood. 2013 May 9;121(19):3867-78, S1-3. doi: 10.1182/blood-2012-11-470146. Epub 2013 Mar 11.


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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Differentiation / genetics*
  • Cell Lineage / genetics
  • Cells, Cultured
  • Embryonic Stem Cells / metabolism
  • Embryonic Stem Cells / physiology*
  • Enzyme Activation / physiology
  • Gene Expression Profiling
  • Hematopoiesis / genetics*
  • Hematopoietic Stem Cells / metabolism
  • Hematopoietic Stem Cells / physiology
  • Humans
  • Mice
  • Mice, SCID
  • Microarray Analysis
  • Myeloid-Lymphoid Leukemia Protein / genetics
  • Myeloid-Lymphoid Leukemia Protein / metabolism
  • Myeloid-Lymphoid Leukemia Protein / physiology*
  • Oncogene Proteins, Fusion / genetics
  • Oncogene Proteins, Fusion / metabolism
  • Oncogene Proteins, Fusion / physiology*
  • fms-Like Tyrosine Kinase 3 / genetics
  • fms-Like Tyrosine Kinase 3 / metabolism
  • fms-Like Tyrosine Kinase 3 / physiology*


  • MLL-AF4 fusion protein, human
  • Oncogene Proteins, Fusion
  • Myeloid-Lymphoid Leukemia Protein
  • FLT3 protein, human
  • fms-Like Tyrosine Kinase 3