Transcriptome drift of FADU after expression of HIF-1a, Twist1 or Bmi1
ABSTRACT: Expression of HIF-1a or Twist1 or Bmi1 in human hypopharyngeal cancer cell line FADU results in the drift of transcriptome profile from an epithelial cell-like signature to a mesenchymal stem cell-like signature. Overall design: Stable transfection of pHA-HIF1a(dODD), pFLAG-Twist1 or pcDNA3-Bmi1 in FADU cell and analyzed the transcriptome by cDNA microarray. FADU transfected with pcDNA3.1 empty vector was used as a control of experiment.
INSTRUMENT(S): [HG-U133_Plus_2] Affymetrix Human Genome U133 Plus 2.0 Array
Project description:Expression of HIF-1a or Twist1 or Bmi1 in human hypopharyngeal cancer cell line FADU results in the drift of transcriptome profile from an epithelial cell-like signature to a mesenchymal stem cell-like signature. Stable transfection of pHA-HIF1a(dODD), pFLAG-Twist1 or pcDNA3-Bmi1 in FADU cell and analyzed the transcriptome by cDNA microarray. FADU transfected with pcDNA3.1 empty vector was used as a control of experiment.
Project description:The critical role of Bmi1 in promoting stem cell properties has been shown in different type of human cancers. Here, we established four stable clones to study Bmi-regulated miRNA expression patterns in head and neck caners. Bmi1-overexpressing cell lines (FaDu- Bmi1vs. FaDu-pcDNA3 cell line), and knock-down of Bmi1 cell lines (OECM1-sh-Bmi vs. OECM1-sh-Bmi1 cell lines) were established and used for analyzing miRNA expression patterms in Bmi-regulatory mechanism.
Project description:To screen the microRNA regulated by Twist1 and Bmi1 Establish stable transfectants of pSUPER-sh-Twist1 or pSUPER-sh-Bmi1 in OECM1 cells and analyze the miRNA expression level of by microRNA microarray. OECM1 transfected with pSUPER-sh-scr was used as a control experiment.
Project description:Through measuring expression levels of lncRNAs/mRNAs in RS-FaDu vs. FaDu cells at 0, 24 and 48 h after 4 Gy radiation, we identified a number of lncRNAs/mRNAs with dysregulation. And the microarray data were verified by qRT-PCR assays. By informatics analyses, we predicted pathways, which were potentially associated with radioresistance of hypopharyngeal carcinoma. Overall design: The radioresistant hypopharyngeal carcinoma cell line (RS-FaDu) was estabished from its parental human hypopharyngeal carcinoma cell line (FaDu) by multi-fractioned radiation. Time-course differential lncRNA and mRNA expression profiles in RS-FaDu vs. FaDu cells were detected by microarray approach, at 0, 2, 48 h after 4 Gy radiation.
Project description:To identify differentially expressed genes by anti cancer treatments (microRNAs or siRNAs) in human cancer, several cell lines (pancreatic cancer, hypopharyngeal squamous cell carcinoma and prostate cancer) were subjected to Agilent whole genome microarrays. Human cell lines (Panc-1, FaDu and PC3) were treated with miRNAs (miR99a-5p, miR-99a-3p, miR-100-3p, miR-150-5p and miR-150-3p), siRNAs (si-FOXQ1).
Project description:Microarray analyses for the identification of differences in gene expression patterns have increased our understanding of the molecular mechanism of ARID3B in HNSCC.We used gene expression analysis data from FaDu-ARID3B and FaDu-pCDH to identify differentially expressed probes. The expression of the embryonic stem cell (ESC) signature in cancer cells indicates the coordinated regulation of the stemness genes in cancer stem cells, which are responsible for cancer initiation and dissemination. let-7 family microRNAs are crucial regulators for stem cell differentiation. In cancer cells, let-7 suppresses cancer stemness through targeting different oncogenes such as c-Myc, RAS, and HMGA2. However, most let-7 target genes are oncogenes rather than stemness factors, and the mechanism of let-7-repressed stemness is unclear. Here we demonstrate that let-7 supresses the formation of AT-rich interacting domain 3B (ARID3B) complex through targeting the expression of ARID3B, the interacting partner ARID3A, and importin 9. ARID3B complex recruits histone demethylase 4C (KDM4C) to the regulatory region of stemness genes for reducing histone 3 lysine 9 trimethylation, resulting in an open configuration of the chromatin of stemness genes. In cancer tissues, ARID3B expression correlates with the nuclear ARID3A expression and a worse prognosis. This result highlights the role of let-7 in regulating stemness through histone modifications. Overall design: The plasmids pCMVΔR8.9, pDVsVg (from the National RNAi Core Facility, Taiwan) and expression lentivectors (pCDH-GFP, pCDH-ARID3B) were co-transfected into 293T cells for virus packaging. The virus-containing supernatant was collected at 48 and 72 hours after transfection. While virus transduction, FaDu cells were infected with 10-fold virus concentrates supplemented with 8 μg/ml polybrene. pCDH-GFP as vector control.
Project description:miRNAs profiling of HepG2 cells comparing vector-control treated HepG2 cells with HepG2 cells transfected with Twist1, Bcl-2, and Twist1/Bcl-2 plasmids. Microarray analysis revealed a panel of miRNAs with significant differential expression among these four HCC cell lines. Overall design: In the study presented here, four HCC cell lines ( HepG2-Twist1, HepG2-Bcl-2, HepG2-Twist1/Bcl-2 HCC cell lines, and HepG2-vector cell lines ) was used to analyze miRNA expression profiles. HepG2-Twist1, HepG2-Bcl-2, HepG2-Twist1/Bcl-2 HCC cell lines can stably express Twist1, Bcl-2, and Twist1/Bcl-2, respectively.
Project description:Our analysis reveals an extensive methylomic drift between normal squamous esophagus and BE tissues in nonprogressed BE patients, with differential drift affecting 4024 (24%) of 16,984 normally hypomethylated cytosine-guanine dinucleotides (CpGs) occurring in CpG islands. The majority (63%) of islands that include drift CpGs are associated with gene promoter regions. Island CpGs that drift have stronger pairwise correlations than static islands, reflecting collective drift consistent with processive DNA methylation maintenance. Individual BE tissues are extremely heterogeneous in their distribution of methylomic drift and encompass unimodal low-drift to bimodal high-drift patterns, reflective of differences in BE tissue age. Further analysis of longitudinally collected biopsy samples from 20 BE patients confirm the time-dependent evolution of these drift patterns. Drift patterns in EAC are similar to those in BE, but frequently exhibit enhanced bimodality and advanced mode drift. To better understand the observed drift patterns, we developed a multicellular stochastic model at the CpG island level. Importantly, we find that nonlinear feedback in the model between mean island methylation and CpG methylation rates is able to explain the widely heterogeneous collective drift patterns. Using matched gene expression and DNA methylation data in EAC from TCGA and other publically available data, we also find that advanced methylomic drift is correlated with significant transcriptional repression of ~ 200 genes in important regulatory and developmental pathways, including several checkpoint and tumor suppressor-like genes. Taken together, our findings suggest that epigenetic drift evolution acts to significantly reduce the expression of developmental genes that may alter tissue characteristics and improve functional adaptation during BE and EAC progression. Overall design: Here we analyze the evolution of epigenetic drift patterns during progression from normal squamous esophagus tissue to Barrett’s esophagus (BE) to esophageal adenocarcinoma (EAC) using 173 tissue samples from 100 (nonfamilial) BE patients, along with publically available datasets including The Cancer Genome Atlas (TCGA).