Altered Expression Profile of Global Genes in U937 Cells Stably Transfected with a Leukemia Fusion Gene: MOZ-TIF2.

Abstract Chromosome translocations involving the monocytic leukemia zinc finger (MOZ) gene occurs often in patients with acute myeloid leukemia (AML) of monocytoid phenotype (FAB types M4 or M5). The resulting MOZ-related fusions include MOZ-CBP, MOZ-P300, and MOZ-TIF2. MOZ has been identified as a histone acetyltransferase of unknown function. MOZ has been demonstrated to interact directly with RUNX1 to regulate RUNX1-mediated transcription and the MOZ-CBP fusion disrupted RUNX1-mediated transcription activity. The MOZ fusion partners CBP and p300 are transcription coactivators with histone acetyltransferase activity; TIF2 is a nuclear receptor coactivator which recruits CBP/P300 in receptor activation. In an animal model, the MOZ-TIF2 fusion successfully induced the occurrence of AML. The leukemia transformation depended on a nucleosome binding motif in MOZ and two CBP binding motifs in TIF2 partner. However, the detailed molecular events relevant to leukemogenesis by MOZ-TIF2 have not been elucidated. We have constructed a MOZ-TIF2 fusion and investigated the effects of this fusion on global gene expression in U937 cells. The U937 cell line with expression of MOZ-TIF2 or wild type MOZ gene were established by stable transfection with pcDNA3-MOZ-TIF2 and pcDNA3-MOZ. RNA was isolated from the early passage of stably transfected U937 cells with TRI reagent® - RNA/DNA/protein isolation reagent according to the manufacturer's protocol. The expression of MOZ-TIF2 in the established cell line was verified by RT-PCR with specific primers for the fusion gene. The examination of global gene expression with Affymetrix gene chip was conducted on the human U95A array. Ten micrograms of total RNA was used. Synthesis of cRNA and subsequent hybridization was completed by the Core Facility at LSUHSC-S according the standard Affymetrix protocol. The human U95A array represents 12,256 oligonucleotides of known genes or expression tags. The gene chip experiment was repeated once for each established cell line with the forced expressed fusion or wild type gene. The raw data was collected and analyzed for the detection and log ratio of each gene or expression tag with the Affymetrix Microarray Suite with the scale set at 2500. The results were further analyzed with GeneSifter.Net. Compared to the expression profile of control cells stably transfected with pcDNA3 vector alone, a > 5-fold change in expression was seen with 49 genes increasing and 32 genes decreasing expression in MOZ-TIF2 expressing cells (p = 0.01). In cells overexpressing wild type MOZ, gene expression increased >5-fold in 411 genes and decreased >5-fold in 48 genes. In a comparison of MOZ expressing cells to MOZ-TIF2 expressing cells a >5-fold change of expression was seen in 296 genes with increased expression in 256 and decreased expression in 40 genes. Among the differentially expressed genes, the c-Myc oncogene expression was increased a strikingly with a 29-fold in MOZ-TIF2 expressing cells over the control cells. Our results suggest that MOZ-TIF2 fusion may interfere with the function of wild type MOZ during the development of myeloid cells by inhibiting MOZ-mediated transcription. In addition, an alternative pathway involving c-Myc may also play an important role in MOZ-TIF2 related leukemogenesis.