Project description:Human Burkitt's lymphoma ST486 cells were transduced with non-target control shRNA lentiviral vectors, FOXM1 shRNA, and MYB shRNA lentiviral vectors. Total RNA was isolated 24h later. cRNA was produced with the standard one-step IVT protocol (Affymetix) and hybridized in U95Av2 gene chips (Affymetrix). Experiment consists in 3 independent samples: Expression profiling of Burkitt's lymphoma cells 24h after non-target control shRNA lentiviral mediated transduction. Expression profiling of Burkitt's lymphoma cells 24h after FOXM1 shRNA lentiviral mediated transduction. Expression profiling of Burkitt's lymphoma cells 24h after MYB shRNA lentiviral mediated transduction. Data processing performed using MAS5 or GCRMA.
Project description:Human Burkitt's lymphoma ST486 cells were transduced with non-target control shRNA lentiviral vectors, FOXM1 shRNA, and MYB shRNA lentiviral vectors. Total RNA was isolated 24h later. cRNA was produced with the standard one-step IVT protocol (Affymetix) and hybridized in U95Av2 gene chips (Affymetrix).
Project description:Interferon regulatory factor 4 (IRF4) is a master transcription factor required for the maturation of germinal center B cells that eventually develop into antibody secreting plasma cells and memory B cells. IRF4-deficient mice exhibit a profound reduction in serum immunoglobulin levels. In spite of wealth of the information relating to IRF4 and B cell biology, little is known about the intricate molecular details of the role of this transcription factor during B cell development. We therefore examined the genome-wide targets of IRF4 by ChIP-chip analysis in GC derived BL2 Burkittâs lymphoma cells. ChIP studies were further supplemented by whole genome expression analysis after shRNA-mediated knockdown of IRF4. Our study revealed that IRF4 regulates expression of genes important for a) BCR signaling b) antigen processing and presentation by MHC. In addition we found that IRF4 possibly in some way involved to regulate LTA, LTB and CXCR5 those involved in immune system development, particularly light zone development related genes such as FDC clustering regulating and IL21R and IL10 who are involved in B cell development.. On the other hand, IRF4 suppressesd genes in the oxidative phosphorylation pathway. Our findings illuminate hitherto unexplored roles of IRF4 in GC B cell development. BL2 Burkitt's lymphoma-derived B cells were infected with lentivirus expressing shRNA for IRF4 or control, and total RNA was subjected to Illumina BeadsExpression Arrays analysis.
Project description:RNAseq analysis of USP7 shRNA KD in T-ALL cell lines. USP7 shRNAs lentivirus was generated by co-transfecting 293T cells with shRNA vectors (OriGene or Sigma). T-ALL cells were transduced with USP7 shRNA lentivirus and sorted for GFP positive cells five days after transduction or selected by puromycin.
Project description:We aimed to detect the mRNA expression levels in HGC-27 cells after transduction with lentivirus harboring DDX5 shRNA or non-targeting shRNA.
Project description:Eos expression in Treg cells have been documented by several microarrays including ours. We hypothesized that Eos facilitates Foxp3- dependent gene repression in Regulatory T cells. In order to investigate the role of Eos in mediating the Foxp3-dependent gene silencing program, we utilized lentiviral shRNA knockdown of Eos in natural Tregs isolated from the periphery of Balb/C mice. A renilla luciferase (RL) gene specific shRNA lentivirus was used as a control for the transduction of cells. The transcriptional profile of naive T cells, natural Tregs, Eos knockdown Tregs, and control shRNA knockdown Tregs was compared using Agilent 4x 44K whole mouse genome array. The goal of this microarray is to document the global effect of the loss of Eos expression on the transcriptional profile of natrual Treg cells. Experiment Overall Design: Eos knock-down (si-Eos) was mediated by Lentivirus expressing GFP as a reporter of transduction and sorting marker. Renilla luciferase specific shRNA lentiviral transduction was carried out in parallel as a control. Naïve (CD4+CD25-CD62Lhi) T cells and nTreg cells were freshly isolated from Balb/C mice.
Project description:IRF2, IRF6, and MYB are candidate regulators of human erythropoiesis. We here examine primary CD34+ hematopoietic stem/progenitor cells (HSPCs)-derived erythroid progenitors with control, IRF2, IRF6, or MYB shRNA lentiviral transduction prior to differentiation. Gene expression microarray profiling datasets for MYB shRNA and control shRNA were obtained from Gene Expression Omnibus (GEO) under accession number GSE25678. The data were analyzed together with the datasets obtained in this study.
Project description:MYB plays a critical role as a regulator of erythropoieisis. We have shown that MYB silences epsilon and gamma-globin expression in erythroid progenitors. We here examine erythroid cells at the basophilic erythroblast stage of differentiation with MYB shRNA or control lentiviral transduction prior to differentiation.
Project description:Acute basophilic leukemia is a rare subtype of acute myeloblastic leukemia. We recently described a recurrent t(X;6)(p11;q23) translocation occurring in male infants with acute basophilic leukemia, generating a MYB-GATA1 fusion gene. To better understand the role of MYB-GATA1 in the leukemogenesis of acute basophilic leukemias, we expressed this chimeric transcription factor in the UT-7 cell line under normal or differentiating conditions. A gene expression analysis revealed 9 genes deregulated by both MYB-GATA1 and the basophilic differentiation. Three of these genes (CCL23, IL1RL1 and NTRK1) were also increased by MYB-GATA1 in CD34-positive cells, as indicated by quantitative RT-PCR. Two independant experiments (A and S) were performed. For each experiment, UT7 cells were transduced with two lentivirus: a lentivirus carrying the MYB-GATA1 (or a control) coding sequence, and a lentivirus carrying a GATA1 (or control) shRNA. Cells transduced with both lentiviruses were selected, and kept in culture under standard or differentiating conditions for 7 days before RNA extraction.
Project description:DDX5 (DEAD-Box Helicase 5) plays a role as an adaptor molecule, is involved in a variety of cellular processes. Here, we performed ChIP-seq with antibody specific for RNA Polymerase II in HGC-27 cells after transduction with lentivirus harboring DDX5 shRNA or non-targeting shRNA.
