Expression profiling of HCT116 cells transfected with shCTRL or shKSR1
Ontology highlight
ABSTRACT: We engineered HCT116 cells by transfecting a non targeting control hairpin or 2 different short hairpin against KSR1 to knockdown KSR1 and microarrays were performed on non-transfected controls, non-targeting shRNA controls, shRNA #1 targeting KSR1, and shRNA #2 targeting KSR1. We used microarray for molecular profiling of colon cancer cells in presence or in absence of KSR1, an important scaffold of Raf/MEK/ERK pathway and identify genes which are regulated by KSR1 Cell lines were measured in as single biological replicates 1. HCT116 untransfected 2 HCT116 + shCTRL 3.HCT116 + shKSR1#1 4. 3.HCT116 + shKSR1#2
Project description:We engineered HCT116 cells by transfecting a non targeting control hairpin or 2 different short hairpin against KSR1 to knockdown KSR1 and microarrays were performed on non-transfected controls, non-targeting shRNA controls, shRNA #1 targeting KSR1, and shRNA #2 targeting KSR1. We used microarray for molecular profiling of colon cancer cells in presence or in absence of KSR1, an important scaffold of Raf/MEK/ERK pathway and identify genes which are regulated by KSR1
Project description:To explore the target genes of long noncoding RNA lncTCF7, we established lncTCF7-silenced HCC primary CSC cells and conducted transcriptome microarray analysis. We used microarrays to identify distinct gene expression underlying shCtrl and shlncTCF7 of hepatocellular carcinoma sample stem cells. We cultured shlncTCF7 and shCtrl cells from hepatocellular carcinoma (HCC) clinical sample, then hybridized on Affymetrix microarrays. We sought to identify distinct target genes of lncTCF7 in liver cancer stem cells (CSCs).
Project description:Pulmonary metastasis is the main cause of medical failure and death of osteosarcoma patients. Our recent study identified IRX1 as a potential metastasis-driving gene in osteosarcoma. Studies showed that IRX1 can promote the migration, invasion and anoikis resistance of osteosarcoma cells. We generated 143B stable IRX1 knockdown and control cell lines, and found that IRX1 knockdown can inhibit the pulmonary metastasis of 143B cells in orthotopic mouse osteosarcoma model. Expression microarrays are performed in143B-shCtrl and 143B-shIRX1 cells to study the mechanism of IRX1 on promoting metastasis of osteosarcoma
Project description:To further understand the cellular function of HOTAIR, we have profiled RNA expression in breast cancer cell lines harboring tetracycline-inducible shRNA of human HOTAIR (Tet-shHOTAIR) or the control shRNA of scrambled sequence (Tet-shCtrl).
Project description:We introduced into subcutaneous fully-differentiated human adipocytes from the same subject small hairpin RNAs (shRNAs) against two genes, thus enabling the study of functional consequences of sustained OLFM2 (shOLFM2; n=3 biological replicates) and PKP2 (shPKP2; n=3) knockdown in these cells. The lentiviral-based transfection was used to provide a long-lasting, convenient, reliable and efficient knockdown into in vitro cultured and fully differentiated adipocytes maintained for 6 d after reaching fully differentiation. A non-targeting shRNA was used as control (shCTRL; n=4) and reference for both modifications.
Project description:To investigate the regulatory role of orphan nuclear receptor ERRα in the progression of ccRCC, we conducted the stable knockdown of ERRα in Caki-1 with shRNA lentivirus. The total protein samples of Caki-1 with shCtrl and shERRα were collected and subjected to label free quantitative proteomics analysis based on LC-MS, the differentially expressed proteins were selected based on the p value and fold-changes between two groups. Then, bioinformatics analysis was performed to reveal the pathways and key factors related to ERRα in ccRCC
Project description:The epithelial-mesenchymal transition (EMT), considered essential for metastatic cancer, has been a focus of much research, but important questions remain. Here, we show that silencing or removing H2A.X, a histone H2A variant involved in cellular DNA repair and robust growth, induced mesenchymal-like characteristics including activation of EMT transcription factors, Slug and ZEB1, in HCT116 human colon cancer cells. Ectopic H2A.X re-expression partially reversed these changes; as did silencing Slug and ZEB1. In an experimental metastasis model, the HCT116 parental and H2A.X-null cells exhibited similar metastases levels, but the cells with re-expressed H2A.X exhibited substantially elevated levels. We surmise that H2A.X re-expression led to partial EMT reversal and increased robustness in the HCT116 cells, permitting them to both form tumors and to metastasize. In a human adenocarcinoma panel, H2A.X levels correlated inversely with Slug and ZEB1 levels. Together, these results point to H2A.X as a novel regulator of EMT. 9 samples in total including 4 replicates of control shRNA and 5 replicates of shH2A.X.
Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to analysis the differiational genes and pathways in MYCWT+shCtrl, MYCWT+shTRIB3, MYCT58A+shCtrl and MYCT58A+shTRIB3 lymphoma cells by using NGS-derived lymphoma transcriptome profiling (RNA-seq). Methods: MYCWT+shCtrl, MYCWT+shTRIB3, MYCT58A+shCtrl and MYCT58A+shTRIB3 cells' mRNA profiles were generated by deep sequencing, in triplicate, using Illumina HiSeq 4000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with following methods: Alignment by using HISAT2 v2.1, IGV was used to to view the mapping result by the Heatmap, histogram, scatter plot or other stytle, FPKM was then calculated to estimate the expression level of genes in each sample, DEGseq v1.18.0 was used for differential gene expression analysis between two samples with non biological replicates and Function Enrichment Analysis including GO enrichment analysis and KEGG . Conclusions: Our study represents the first detailed analysis of MYCWT+shCtrl, MYCWT+shTRIB3, MYCT58A+shCtrl and MYCT58A+shTRIB3 cells' transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.
Project description:KSR1 signaling alters the translational landscape of human CRC cells to support their survival. To determine the effect of KSR1 on translatomes in colon cancer cells, we performed genome-wide polysome profiling. We stably expressed short hairpin RNA (shRNA) constructs targeting KSR1 or a non-targeting control in two K-Ras mutant CRC cell lines, HCT116 and HCT15. We isolated and quantified both total mRNA and efficiently translated mRNAs (associated with ≥ 3 ribosomes) using RNA sequencing. We used Anota2seq to calculate translation efficiency (TE) by comparing the differences in efficiently translated mRNAs to the total transcript of each mRNA.