Genome Wide Location Analysis Reveals Deregulated MicroRNA Genes In MLL Rearragned Leukemic Genome

Abstract 2507

The MLL (mixed-lineage leukemia) gene encodes a histone methyltransferase that is critical in maintaining gene expression during hematopoiesis. Chromosomal translocations disrupting MLL often leads to the creation of MLL fusion genes that act as potent drivers of acute leukemia. MLL fusion proteins are oncogenic transcription factors that activate the expression of downstream target genes. Expression profiling on patient primary samples and established mouse models has revealed hundreds of protein coding genes which are either up-regulated or down-regulated in MLL associated leukemias. Persistent coexpression of two of those genes, HoxA9 and Meis1, is essential for the initiation and maintenance of MLL leukemia. Our studies have also shown strong association of a microRNA (miRNA) expression signature with MLL- rearranged leukemia, and the expression of several miRNAs were under the control of MLL wild type and fusion proteins. Although profiling of miRNA expression has been reported, the mechanisms underlying deregulated miRNA expression in MLL associated leukemia are poorly understood. Given the role of miRNA as a global suppressor of mRNA gene expression, we hypothesized that the expression of miRNAs could be directly activated by MLL fusion and/or wild type proteins upon MLL gene rearrangement and subsequently down-regulate pertinent target mRNAs to contribute to leukemogenesis. To test our hypothesis in a systematic way, we examined an inducible MLL-ENL-ER transformed mouse cell line, which grow as myeloblastic cells in the presence of MLL-ENL, and differentiate into neutrophils upon inactivation of the fusion protein. Using chromatin immunoprecipitation assay followed by next generation sequencing (ChIP-Seq), we determined whole genome MLL binding pattern in this cellular model. Upon activation of MLL-ENL, 24 miRNAs showed a significant increase in the level of MLL binding (FDR<0.25), suggesting that those genes are directly bound by the MLL-ENL fusion protein. To explore the impact of MLL fusion protein on miRNA and mRNA gene regulation, we performed whole genome expression analysis using Affymetrix mouse microarray in the presence or absence of MLL-ENL. Upon induction of MLL-ENL, the expression levels of 38 miRNAs (out of 609 tiled on the array) were increased, and 57 of 7858 expressed protein-coding genes were down-regulated. An integrative analysis of MLL binding and mRNA/miRNA expression profiling data showed that transcription of three miRNAs were activated upon binding of MLL-ENL, and ten protein coding genes are potential targets of these miRNAs. We are currently exploring the role of these three miRNAs and their respective mRNA target genes in leukemogenesis using in vitro and in vivo models. Taken together, our data suggest that MLL fusion protein may play an important role in leukemogenesis by promoting miRNA transcription, which subsequently inhibit the expression of critical mRNA target genes. Our study provides a basis to further explore the regulatory network involving MLL fusion protein and its key miRNA target genes in the leukemic genome.