Project description:Dystroglycan has recently been characterised in blood tissue cells, as part of the dystrophin glycoprotein complex involved in the differentiation process of neutrophils.In the present study we have investigated the role of dystroglycan in the human promyelocytic leukemic cell line Kasumi-1 differentiated to macrophage-like cells.We characterised the pattern expression and subcellular distribution of dystroglycans in non-differentiated and differentiated Kasumi-1 cells.Our results demonstrated by WB and flow cytometer assays that during the differentiation process to macrophages, dystroglycans were down-regulated; these results were confirmed with qRT-PCR assays. Additionally, depletion of dystroglycan by RNAi resulted in altered morphology and reduced properties of differentiated Kasumi-1 cells, including morphology, migration and phagocytic activities although secretion of IL-1? and expression of markers of differentiation are not altered.Our findings strongly implicate dystroglycan as a key membrane adhesion protein involved in actin-based structures during the differentiation process in Kasumi-1 cells.
Project description:Acute lymphoblastic leukemia harboring the fusion genes involving the MEF2D transcription factor (MEF2D-ALL) is associated with poor clinical outcomes. To explore binding sites in the genome in MEF2D-ALL, we genome-edited a MEF2D-ALL cell line Kasumi-7 so that the fusion is tagged with HA at the carboxyl-terminal and co-expressed with GFP. We used this cell line for ChIP-seq using anti-HA antibody. Pair-end reads for Input and HA ChIP DNA are provided.
Project description:Kasumi-1 has played an important role in an experimental model with t(8;21) translocation, which is a representative example of leukemia cell lines. However, previous studies using Kasumi-1 show discrepancies in the genome profile. The wide use of leukemia cell lines is limited to lines that are well-characterized. The use of additional cell lines extends research to various types of leukemia, and to further explore leukemia pathogenesis, which can be achieved by uncovering the fundamental features of each cell line with accurate data. In this study, ten Kasumi cell lines established in Japan, including five that were previously unknown, have been characterized by SNP microarray and targeted sequencing. SNP genotyping suggested that the genetic ancestry in four of the ten Kasumi cell lines was not classified as Japanese but covered several different east-Asian ethnicities, suggesting that patients in Japan are genetically diverse. TP53 mutations were detected in two cell lines with complex array profiles, indicating chromosomal instability (CIN). A quantitative assessment of tumor genomes at the chromosomal level was newly introduced to reveal total DNA sizes and Scales of Genomic Alterations (SGA) for each cell line. Kasumi-1 and 6 derived from relapsed phases demonstrated high levels of SGA, implying that the level of SGA would reflect on the tumor progression and could serve as an index of CIN. Our results extend the leukemia cellular resources with an additional five cell lines and provide reference genome data with ethnic identities for the ten Kasumi cell lines.
Project description:The oncogenic fusion protein RUNX1-ETO is a product of the t(8;21) translocation and consists of the hematopoietic transcriptional master regulator RUNX1 and the repressor ETO. RUNX1-ETO is found in 10-15% of acute myeloid leukemia and interferes with the expression of genes that are essential for myeloid differentiation. The neutrophil serine protease Cathepsin G is one of the genes suppressed by RUNX1-ETO, but little is known about its impact on the regulation of other lysosomal proteases. By lentiviral transduction of the t(8;21) positive cell line Kasumi-1 with an RUNX1-ETO specific shRNA, we analyzed long-term effects of stable RUNX1-ETO silencing on cellular phenotypes and target gene expression. Stable anti RUNX1-ETO RNAi reduces both proliferation and apoptosis in Kasumi-1 cells. In addition, long-term knockdown of RUNX1-ETO leads to an upregulation of proteolytic activity in Kasumi-1 cells, which may be released in vitro upon cell lysis leading to massive degradation of cellular proteins. We therefore propose that protein expression data of RUNX1-ETO-silenced Kasumi-1 cells must be analyzed with caution, as cell lysis conditions can heavily influence the results of studies on protein expression. Next, a mass spectrometry-based approach was used to identify protease cleavage patterns in RUNX1-ETO-depleted Kasumi-1 cells and Neutrophil Elastase has been identified as a RUNX1-ETO candidate target. Finally, proteolytic activity of Neutrophil Elastase and Cathepsin G was functionally confirmed by si/shRNA-mediated knockdown in Kasumi-1 cells.
Project description:Gain-of-function mutations in KIT, a member of the receptor type tyrosine kinases, are observed in certain neoplasms, including mast cell tumors (MCTs) and acute myelogenous leukemias (AMLs). A MCT line HMC1.2 harboring the KIT mutation was reported to express CD72, which could suppress the cell proliferation. Here, we examined the ability of CD72 to modify the growth of AMLs harboring gain-of-function KIT mutations. CD72 was expressed on the surface of the AML cell line, Kasumi-1. CD72 ligation by an agonistic antibody BU40 or by a natural ligand CD100, suppressed the proliferation of the Kasumi-1 cells and enhanced cell death, as monitored by caspase-3 cleavage. These responses were associated with the phosphorylation of CD72, the formation of the CD72 - SHP-1 complex and dephosphorylation of src family kinases and JNK. Thus, these results seemed to suggest that CD72 was the therapeutic potential for AML, as is the case of MCTs.
Project description:miRNA expression profiling of KASUMI-1 ctrl vs KASUMI-1 treated Saha 5 µM 6h Overall design: Two-condition experiment, KA NT (KASUMI-1 ctrl) vs KA 6h s (KASUMI-1 treated Saha 5 µM 6h) Biological replicates: 3 control, 3 treated, independently grown and harvested at 6 hours. One replicate per array. LC sciences service