Abstract
Aberrant activation of the TALE (three-amino-acid loop extension) homeobox gene MEIS1 is frequently observed in acute leukemia. A high level of Meis1 expression shortens the latency, as well as accelerates the onset and progression of acute myeloid leukemia in mice. Although available data support the notion that MEIS1 is oncogenic by means of overexpression rather than mutation, the molecular mechanism underlying persistent activation of this gene in leukemia remains poorly understood. We have identified a distal enhancer (designated EX) at the MEIS1 locus and found that MEIS1 itself and HOXA9 synergistically activate the regulatory activity of EX through a conserved DNA motif. Using comparative genomics-based strategies, we computationally predicted 14 putative enhancer DNA elements in the 1300 Kb MEIS1 locus. We confirmed that the human sequences of 6 of these 14 elements showed enhancer function as they were able to direct GFP expression in a spatiotemporal manner during embryonic hematopoiesis in an in vivo transgenic zebrafish assay. To explore whether these 6 elements play a role in regulating MEIS1 expression in human leukemia, we examined the presence of histone modifications that are associated with gene activation in a panel of 8 leukemia cell lines. In MEIS1-expressing cells, but not MEIS1 transcript-negative cell lines, the genomic region corresponding to enhancer EX showed extensive H3K4 mono-methylation (me1H3K4), a hallmark of active distal enhancers. When endogenous MEIS1 expression decreases during cellular differentiation, the active histone mark me1H3K4 was replaced by repressive H3K27 methylation and the associated EZH2 polycomb protein at the EX region. In murine leukemia models, retroviral integration at the Meis1 locus has been shown to result in overexpression of Meis1 leading to development of AML. Strikingly, we found that viral integration frequently occurred within the genomic sequence corresponding to enhancer EX. Collectively, these data suggest that enhancer EX plays an important role in maintaining high level expression of endogenous MEIS1 in transformed leukemic cells. In an effort to identify the molecular basis underlying the observed regulatory function of EX, we found MEIS1 and HOXA9 synergistically stimulate the enhancer activity of EX in reporter gene assays, suggesting that production of MEIS1 protein can reinforce its own expression. Mutation of a conserved site within the enhancer abolished the ability of these two factors to activate reporter gene expression. Using ChIP (Chromatin Immunoprecipitation), we further demonstrated that MEIS1 and HOXA9 specifically bind to the genomic region containing the conserved site in vivo, and the presence of these two factors in the region is associated with active MEIS1 expression. These studies provide insight on the molecular mechanisms controlling transcription regulation of MEIS1, suggesting that overexpression of MEIS1 in acute leukemia is sustained by an autoregulatory loop mediated through a distal enhancer element.
Evidence for Redundancy But Not trans Factor-cis Element Coevolution in the Regulation of Drosophila Yp Genes
