Gene Expression Profiling in A549 Lung Cancer Cell Line Following siRNA Mediated Knock-down of ALDH1A1 and ALDH3A1
ABSTRACT: Aldehyde dehydrogenase isozymes ALDH1A1 and ALDH3A1 are highly expressed in non small cell cell lung cancer. Neither the mechanism nor the biological significance for such over expression have been studied. We used microarrays to analyze changes in A549 lung cancer cell line in which ALDH activity was reduced using lentiviral mediated expression of siRNA against both isozymes (Lenti 1+3) Keywords: Gene Profiling after ALDH Knock Down Overall design: A549 lung cancer cell lines were transduced with lentiviral vectors containing specific siRNA sequences against ALDH1A1, ALDH3A1, both vectors (Lenti 1+3 cells), and against the green flourescent protein (GFP) gene (GFP cells, used as control).
Project description:Aldehyde dehydrogenase isozymes ALDH1A1 and ALDH3A1 are highly expressed in non small cell cell lung cancer. Neither the mechanism nor the biological significance for such over expression have been studied. We used microarrays to analyze changes in A549 lung cancer cell line in which ALDH activity was reduced using lentiviral mediated expression of siRNA against both isozymes (Lenti 1+3) Experiment Overall Design: A549 lung cancer cell lines were transduced with lentiviral vectors containing specific siRNA sequences against ALDH1A1, ALDH3A1, both vectors (Lenti 1+3 cells), and against the green flourescent protein (GFP) gene (GFP cells, used as control).
Project description:The emergence of tumor cells with certain stem-like characteristics such as high aldehyde dehydrogenase (ALDH) activity contributes to chemotherapy resistance. Here we report that inhibition of the BET protein BRD4 potentiates the tumor suppressive effects of cisplatin by targeting ALDH activity. The clinically applicable small molecule BET inhibitor JQ1 synergized with cisplatin by suppressing the growth of epithelial ovarian cancer cells both in vitro and in vivo. This correlated with the suppression of ALDH activity and ALDH1A1 gene expression. BRD4 regulates ALDH1A1 gene transcription through a super-enhancer and expression of its associated enhancer RNA. Thus, targeting the BET protein BRD4 using clinical applicable small molecule inhibitors such as JQ1 is a promising strategy to enhance cisplatin response. Overall design: DMSO and JQ1 treated cells were assayed by BRD4 CHIP-seq and RNA-seq
Project description:Transcriptional profiling of human lung cancer cell lines A549 comparing control cells transfected with an empty lentiviral expression vector pEZ-Lv201 and A549 cells transfected with a pEZ-Lv201-FENDRR. The latter forms a FENDRR stably over-expressed cell lines. Goal was to to assess the alteration of gene expression profiles caused by FENDRR upregulation. Overall design: Two-condition experiment, A549 vs. FENDRR stably over-expressed A549 cells. No replicate.
Project description:In this experiments different treatments were applied to lung cancer cell lines 1)H720 cell line suspension was treated by 1 micromole of 5-lipoxygenase activating protein inhibitor MK886 serum-free in TIS medium for 24 h. 2) A549 cell line was treated as H720 by MK886 3) A549 was treated by 100 nM of silencing cocktail (siRNA) prepared against the 4-th exon of the gene and generated by RNASEIII digestion of double stranded RNA. The cocktail was prepared by Ambion technology, see the manufacturer's protocol. The observed silencing was >90% 4) Based on Oligoengine vectors, silencing constructs were created against HRPA2B1 transcript(HNRPA2B1 is a subunit of spliceosome). Stable transfectants were raised harboring the construct. Keywords: drug treatment Overall design: 1) MK886 treatement of H720: Three chips titled MK served as "treated" part, three chips titled "control" served as controls. 2) MK886 treatment of A549: two duplicates, 1+2 and 3+4, performed on different dates 3) HNRPA2B1 silencing: pairs of controls and silenced sample 4) FLAP silencing: 1 control, 2 treatments
Project description:We evaluated the effect of NORAD (also known as LINC00657 or LOC647979) shRNA on TGF-beta induced changes in the gene expression in A549 cells by RNA-seq. Overall design: mRNA expression was determined in a lung adenocarcinoma cancer cell line A549 infected with NORAD shRNA-expressing lentiviral vector and treated with TGF-beta.
Project description:Biological effects of overexpression of miR-146b microRNAs in the A549 human lung cancer cell-line was studied. A549 cells were engineered to express the precursor RNA (pre-miR-146b) that generates the miR-146b microRNAs. Control cells were engineered using the same gene expression plasmid (pLemiR, Open Biosystems®) but without the pre-miR-146b insert. The Trans-Lentiviral GIPZ™ packaging system (Open Biosystems®) was used to generate stable transfectant populations of the engineered cells. Gene expression was compared between A549 stable transfectants of empty pLemiR plasmid (A549/vec cells) or of the pLemiR plasmid containing an insert for overexpressing human pre-miR-146b (A549/146b cells). Two different cell cultures were used for each cell-line (for biological replicates).
Project description:We sequenced mRNA from 3 biological replicates each of A549 lung adenocarcinoma cell lines expressing shRNA against GFP (control), PRMT5, or MEP50. We then determined differential gene expression. Transcriptome analysis of mRNA testing the role of PRMT5 and MEP50 by knockdown in A549 human lung adenocarcinoma cells
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:Background: α-catulin may functions as an oncoprotein, sustaining proliferation by preventing cellular senescence and promoting cancer cell migration. In this study, we investigated the mechanism of α-catulin in cancer cell migration and metastasis in lung cancer. Method: α-catulin mRNA expression was isolated from A549/AS2neo (control) and A549/AS2neo-α-catulin stable cells. The Phalanx Human OneArray microarray analysis was performed to identify α-catulin downstream genes. Results: Overexpression of α-catulin increased cancer cell migration and metastasis. By using Phalanx Human OneArray microarray we have identified panel of genes altered by α-catulin overexpression, consisting of CDC42, intergrins and genes related to cytoskeleton remodeling. Conclusion: α-catulin is an oncoprotein and promotes lung cancer cell migration and metastasis. Two-condition experiment, Vector vs. alpha-catulin overexpression cells.The cDNAs encoding full-length human alpha-catulin were amplified and subcloned into lentiviral pLKO_AS2.neo which generated full-length alpha-catulin. Vector control or alpha-catulin lentivirus were transduced into A549 cells and G418 was used to select stable cells.