Project description:Transition from a partially reprogrammed pre-iPSC state to iPSC state can be achieved by modulating levels of histone modifying enzymes or proteins that can bind to histone modifications We used microarrays to determine the gene expression profile of pre-iPSCs depleted for either 3 histone methyltransferases together or the HP1gamma protein pre-iPSCs were subjected to simultaneous siRNA mediated knockdown of Ehmt1, Ehmt2 and Setdb1- which is the 3XHMT sample that mediate H3k9me2/me3 or Cbx3( HP1gamma) or control siRNA to luciferase
Project description:We report the application of Illumina Hi-Seq sequencing technology for high-throughput profiling of Cbx3/HP1g deposition in mammalian CD8+ effector T cells. By obtaining over four billion bases of sequence from chromatin immunoprecipitated DNA, genome-wide chromatin-state maps of mouse wt CD8+ effector T cells were generated. We find that Cbx3/HP1g negatively regulates the germinal center and high-affinity antibody responses in a CD8+ T-cell-dependent manner. CD8-restricted deletion of Cbx3/HP1g confers hightened effector and persistence capacity on CD8+ effector T cells. As a result, Cbx3/HP1g-deficient CD8+ effector T cells can efficiently control solid tumor growth in vairious mouse models (melanoma, ovarian and neuroblstoma).
Project description:We examined the locations of Cbx3 by chromatin immunoprecipitation in ESCs and pre-iPSCs Examination of Cbx3 in mESC (mouse embryonic stem cells) and pre-iPSCs (fibroblast derived partially reprogrammed cells)
Project description:Cbx3 (HP1γ) that is a member of the heterochromatin protein 1 familiy enriched in gene bodies of pluripotent ESCs but promoters of differentiated pre-iPSCs. To determine whether Cbx3 becomes enriched at promoters to influence gene regulation during differentiation, we converted ESCs to NPCs in monolayer culuture and performed genomewide ChIP-seq of Cbx3. Our results shows that Cbx3 enriches at the promoters of genes upon differentiation of ESCs to NPCs and stablizes the binding of Mediator subunit Med26 to pre-initiation complex (PIC) in NPCs.
Project description:RNA-seq and RNAchIP-seq experiments were performed on MEF-derived cell lines knock-out for the expression of HP1g/CBX3 (R cell line) or recomplemented for the expression of HP1g (HPg cell line). Cells were tested under normal growth or conditions of stress (p samples) by treatment with the phorbol ester PMA. Each experiment was performed in biological triplicates.
Project description:RNA-seq and RNAchIP-seq experiments were performed on MEF-derived cell lines knock-out for the expression of HP1g/CBX3 (R cell line) or recomplemented for the expression of HP1g (HPg cell line). Cells were tested under normal growth or conditions of stress (p samples) by treatment with the phorbol ester PMA. Each experiment was performed in biological triplicates.
Project description:Gene expression analyses of Tif1b, Hp1a or Hp1g knockdown hematopoietic progenitors. Growth of hematopoietic stem cells (HSCs) are significantly impaired upon knockdown of Hp1a and Hp1g. Results provide insight into the role of these factors in hematopoiesis. HSCs (CD34- c-Kit+ Sca1+ Lineage- ) were transduced with lentivirus expressing shRNA against Tif1b, Hp1a or Hp1g, and cultured for 14 days. Then, GFP+ c-Kit+ Lineage- hematopoietic progenitors were sorted and subjected into microarray analysis
Project description:Functional crosstalk between histone modifications and chromatin remodeling has emerged as a key regulatory mode of transcriptional control during drug resistance, but the underlying mechanisms are not fully understood. Here we demonstrate that HP1g regulates chromatin dynamic and gene transcription during drug resistance in multiple myeloma. To study the cellular and molecular function of HP1g during bortezomib resistance, we used RNA sequencing (RNA-seq) to generate gene expression profiling under HP1g steady overexpression in LP-1. We identified multiple genes whose expression is significantly affected by HP1g levels.
