Project description:This experiment uses a transgenic cell line expressing bacterial OGA BtGH84 fused to a localization peptide (NLS) and regulated by Tet-On system. OGA is a glycosidase that removes O-GlcNAc modifications. We evaluated the changes in gene expression before and after O-GlcNac removal by OGA.
Project description:This experiment uses a transgenic cell line expressing bacterial OGA BtGH84 fused to a localization peptide (NLS) and regulated by Tet-On system. OGA is a glycosidase that removes O-GlcNAc modifications. We evaluated the changes in chromatin openness before and after O-GlcNac removal by OGA.
Project description:Resveratrol (RSV) is a small compound with many reported beneficial effects, including increased expression of genes involved in inborn errors of metabolism. We wanted to investigate the effects of RSV on gene regulation on a global scale in order to detect new target genes for RSV mediated up-regulation with respect to disease treatment. Because fibroblasts are widely used diagnostically and reflect many metabolically important tissues well, we chose to investigate normal fibroblasts treated with RSV. We performed this with and without knock down of sirtuin 1 (SIRT1) since SIRT1 has been reported as a key mediator of RSV effects. We performed Next Generation Sequencing of RNA on 2 normal fibroblast cell line with and without the respective treatments.
Project description:To find target genes of MYC in GP5d human colon cancer cells, the cells were treated with siRNA against CTNNB1 and MAX. The level of MYC knockdown in human was not sufficiently efficient to be included in the experiments within the criteria list, and hence the values from CTNNB1 knockdown were used as the expression MYC dropped to below 5% of that in the control sample in RNAseq.
Project description:The objective of the present study is to investigate the role of DNA-PK inhibition in cell death induced by heat stress (44M-BM-0C, 60 min). Comparative gene expression analysis was performed with mock cells, negative control siRNA-treated cells and DNA-PK siRNA-treated cells. The expression of DNA-PK was confirmed by Western blotting. Gene expression was analyzed using GeneChip oligonucleotide microarrays and computational gene expression analysis tools. DNA-PK-knockdown human cervical carcinoma HeLa cells were treated with heat (44M-BM-0C, 60 min) and then were cultured at 37M-BM-0C for 6 h. Total RNA samples were prepared from the cells. Gene expression was analyzed by an Affymetrix GeneChipM-BM-. system with GeneChip Human Gene 1.0 ST arrays. Sample preparation for array hybridization was carried out as described in the manufacturer's instructions.
Project description:To find target genes of MYC in GP5d human colon cancer cells, the cells were treated with siRNA against CTNNB1 and MAX. The level of MYC knockdown in human was not sufficiently efficient to be included in the experiments within the criteria list, and hence the values from CTNNB1 knockdown were used as the expression MYC dropped to below 5% of that in the control sample in RNAseq.
Project description:Malignant peripheral nerve sheath tumor (MPNST) is a type of soft tissue sarcoma that occurs in carriers of mutations in the neurofibromatosis type I gene (Nf1) as well as sporadically. Plexiform neurofibromas in NF1 patients have a significant risk of developing into MPNSTs leading to increased morbidity and mortality from this syndrome. Surgery is the primary intervention but it is not always effective due to the tendency of MPNSTs to infiltrate the surrounding tissue or grow in an inoperable location. Neurofibromin, the protein coded by the Nf1 gene, functions as a GTPase activating protein (GAP) whose mutation leads to constitutive activation of RAS and mitogen-activated protein kinase (MAPK) signaling in NF1 patients’ tumors. However, therapeutic targeting of RAS and MAPK have had limited success (Kalamarides, et al., 2012). In this study, we modulated NRAS, MEK1/2 and neurofibromin levels in MPNST cell lines and determined the global gene expression changes that were associated with each experimental condition. Furthermore, gene expression changes due to neurofibromin deficiency but independent of NRAS and MEK1/2 regulation were characterized for the first time in MPNST cell lines. There are total 4 comparison scenarios. Each scenario has two different samples to compare and each sample has three replicates. Comparison 1: ST88-14 cell line versus normal human schwann cell; comparison 2: U0126 treated ST88-14 versus DMSO treat ST88-14 as control; comparison 3: siNRAS treated ST88-14 versus scrambled control treated ST88-14 cell line; comparison 4: siNf1 treated STS26T cell line versus scrambled control RNA treated STS26T cell line.
Project description:Neuropathy Target Esterase (NTE) is a protein involved in the development of a polyneuropathy caused by exposure to certain organophosphorus compounds. In vivo and in vitro studies have also associated NTE with embryonic development since NTE null mice embryos are non-viable, and silencing the NTE-codifying gene (Pnpla6) in mouse embryonic stem cells strongly alters the differentiation of vascular and nervous systems. In this paper, human embryonal carcinoma stem cells (hNT2) are used as in vitro neurodifferentiation model to determine whether PNPLA6 gene silencing is able to alter the differentiation into a neuronal phenotype. In controls, the PNPLA6 mRNA levels increased in parallel with other neuroectodermal markers during the neurodifferentiation. PNPLA6 gene silencing with specific interference RNA led to a 97% maximum decrease in gene expression by day 3 after transfection, with a maximum of 50% reduction in NTE enzymatic activity by day 4. Microarray analyses of the PNPLA6-silenced cells showed alterations in several developmental processes, mainly neurogenesis and epithelium tube morphogenesis. PNPLA6 silencing also led to a reduction in electrical activity and an altered neuronal phenotype. This work is the first proof supporting the hypothesis that NTE protein plays a critical role in human early neurodevelopment using a human cell differentiation model. PNPA6 gene silencing in undifferentiatied hNT2 cells was performed, together with a Control (non-gene silencing cells) and a Negative Control (non-specific RNA silencer). Three biological replicates.
Project description:This experiment aimed at investigating how O-GlcNac occupancy sites are impacted by RNA Polymerase II removal upon doxycycline stimulation. These human colon adenocarcinoma DLD-1 cells express OsTIR and a cassette encoding mini-AID (mAID) and fluorescent protein mClover (mAID+mClover) at the initiation site of the endogenous Rpb1 gene locus (POLR2A) (Nagashima 2019).