Project description:MLL-fusions represent a large group of leukemia drivers, whose diversity originates from the vast molecular heterogeneity of C-terminal fusion partners of MLL protein. While studies of selected MLL-fusions have revealed critical molecular pathways, unifying mechanisms across all MLL-fusions remain poorly understood. We present the first comprehensive survey of protein-protein interactions of seven distantly related MLL-fusion proteins: MLL-AF1p, MLL-AF4, MLL-AF9, MLL-CBP, MLL-EEN, MLL-ENL and MLL-GAS7.
Project description:RNA-Seq of 1) human AML samples; 2) sorted, uncultured distinct population from human cord blood (CB); 3) short-term (ST) cultured sorted CB cells transduced with MLL-ENL, MLL-AF6 or untransduced; and 4) cultured (LT) sorted CB cells transformed with MLL-ENL or MLL-AF6. Cells from MLL-fusion AML patients are bulk. Several cords were used for the sorting (CB1, CB2, CB3, 135, 141...) and these represent biological replicates. Several samples were sequenced several times in different lanes and results were merged together for the analysis (rep1,rep2...). Samples were used to determine the different effect of MLL-fusions in different celltypes just after the transduction, and after a longer time period when cells were transformed. Sorted CB samples, uncultured as well as transformed by MLL-fusions, were used in machine learning approach to predict which of the patients originated from which cell-type of origin.
Project description:Chromosomal translocations encoding the MLL-AF9 and MLL-ENL fusion transcription factors are prevalent in infant acute leukaemia and therapy-related leukaemia. In order to conditionally express the MLL-fusion oncogene in primary haematopoietic progenitor cells (HPC), retroviral delivery of the Tet-off expression system was used (Horton et al., Cancer Res, 2005). Treatment of the conditional cells with Doxycycline caused a decrease in MLL-AF9/ENL mRNA and protein expression, and resulted in terminal differentiation of the cells. By analysing global changes in gene expression after treatment of cells with Doxycycline we were able to identify a number of potential transcriptional target genes of the MLL-AF9 and MLL-ENL fusion oncogenes.
Project description:To investigate the contribution of ENL YEATS domain and downstream sequences in MLL-ENL leukemogenesis program, we generated MLL-ENL and MLL-ENL ∆YEATS (ENL aa372-559) cell lines by retrovirally introducing these constructs into lineage negative hematopoietic stem and progenitor cells (HSPCs).
Project description:To identify the target genes of Runx1 in MLL fusion leukemia, we performed microarray analysis using control and Runx1-deficient MLL-ENL leukemia cells. Runx1 intact and excised bone marrow cells were transduced with MLL-ENL and transplanted into congenic mice. Leukemic cells were harvested from moribund mice, and gene expression was compared using 3 independent leukemia cells for each genotype.
Project description:MLL encodes a histone methyltransferase that is critical in maintaining gene expression during embryonic development and hematopoiesis. 11q23 translocations encode chimeric MLL fusions that act as potent drivers of acute leukemia. However, it remains unclear what portion of the leukemic genome is under the direct control of the MLL fusion protein. By comparing patient-derived leukemic cell lines, we find that MLL fusion-bound genes are a small subset of that recognized by wild-type MLL. In an inducible MLL-ENL cellular model, binding of the MLL fusion protein and changes in H3K79 methylation are limited to a specific portion of the genome, whereas wild-type MLL distributes to a much larger set of gene loci. Surprisingly, among 223 MLL fusion-bound genes, only 12 demonstrate a significant increase in mRNA expression upon induction of the fusion protein. In addition to Hoxa9 and Meis1, this includes Eya1 and Six1 which comprise a heterodimeric transcription factor important in several developmental pathways. We show that Eya1 has the capacity to immortalize hematopoietic progenitor cells in vitro and collaborates with Six1 in hematopoietic transformation assays. Altogether, our data suggest that MLL fusions contribute to the development of acute leukemia through direct activation of a small set of target genes. We explored an inducible MLL-ENL cellular model, which was obtained from Dr. Robert Slany (University Erlangen, Germany). We wished to examine the differential expressed genes that are bound by MLL wild type (No 4-OHT) and fusion (4-OHT) proteins, combinding the ChIP-chip data to explore the potential MLL fusion-regulated genes.
Project description:To identify the target genes of Runx1 in MLL fusion leukemia, we performed microarray analysis using control and Runx1-deficient MLL-ENL leukemia cells.
Project description:MLL-fusions may induce leukemogenic gene expression programs by recruiting the histone H3K79 methyltransferase to MLL-target promoters. We evaluated gene expression changes after cre-mediated loss of Dot1l in leukemia cells obtained from mice injected with MLL-9 transformed lineage negative bone marrow cells.
