We exposed a panel of 32 breast cancer cell lines or normal human mammary epithelial cells to 20% or 1% O2 concentration for 24h. Total RNA was extracted from cells using TRIzol (Invitrogen) and treated with DNase I (Ambion). All samples had a RIN value of >9.0 when measured on an Agilent Bioanalyzer. Libraries for RNA-Seq were prepared with KAPA Stranded RNA-Seq Kit. The workflow consisted of mRNA enrichment, cDNA generation, end repair to generate blunt ends, A-tailing, adaptor ligation and 12 cycles of PCR amplification. Unique adaptors were used for each sample in order to multiplex samples into several lanes. Sequencing was performed on Illumina Hiseq 3000/4000 with a 150bp pair-end run. A data quality check was done on Illumina SAV. Demultiplexing was performed with Illumina Bcl2fastq2 v 2.17 program. Overall design: 32 breast cancer cell lines exposedto standard tissue culture conditions normoxic (20% O2) or hypoxic (1% O2) conditions.
Fate-mapping post-hypoxic tumor cells reveals a ROS-resistant phenotype that promotes metastasis.
Cell line, Subject
View SamplesWe designed a novel approach to fate-map hypoxic cells in order to determine their cellular response to physiological O2 gradients. Our system causes a change in the expressing fluorescent protein upon hypoxic exposure (DsRed -> GFP). We generated hypoxia fate-mapping tumors using MDA-MB-231 cells expressing our system. Tumors were resected 2 weeks post-implantation, mechanically disrupted and digested with collagenase to obtain a cell suspension. The cell suspension was enriched using magnetic-activated cell sorting (MACS) and DsRed+ cells were isolated from GFP+ cells by fluorescence-activated cell sorting (FACS) directly into Tris Reagent (Zymo Research). Total RNA was extracted from cells using TRIzol (Invitrogen) and purified using Direct-zol RNA mini kit (Zymo Research) with DNase I treatment. After RNA purification, samples were confirmed to have a RIN value > 9.0 when measured on an Agilent Bioanalyzer. Libraries for RNA-Seq were prepared with KAPA Stranded RNA-Seq Kit. The workflow consisted of mRNA enrichment, cDNA generation, end repair to generate blunt ends, A-tailing, adaptor ligation and 12 cycles of PCR amplification. Unique adaptors were used for each sample in order to multiplex samples into several lanes. Sequencing was performed on Illumina Hiseq 4000 with a 150bp pair-end run. A data quality check was done on Illumina SAV. Demultiplexing was performed with Illumina Bcl2fastq2 v 2.17 program. Overall design: In order to permanently mark hypoxic cells upon exposure to hypoxia, we generated a dual-vector hypoxia fate-mapping system delivered by a lentiviral approcah. Vector 1 expresses a red fluorescent reporter protein (DsRed) with a stop codon flanked by tandem loxP sites (“floxed”) and located in front of a gene encoding a green fluorescent protein (GFP). Vector 2 contains an altered Cre gene modified by the addition of an oxygen-dependent degradation domain (ODD) that is under the transcriptional control of a synthetic HIF-DNA binding sequence (HRE). HIF stabilization causes the activation of vector 2 by binding to hypoxia-dependent DNA response elements (HREs). Vector 2 activation causes the production of a genetically modified Cre protein that is only stable under hypoxia, leading to the cleavage of DsRed and permanent GFP expression. DsRed+ and GFP+ cells were sorted from MDA-MB-231 hypoxia fate-mapping tumors.
Fate-mapping post-hypoxic tumor cells reveals a ROS-resistant phenotype that promotes metastasis.
Cell line, Subject
View SamplesThe study analyzes analyzes gene expression changes in the ankle joint in mouse TNFa overexpression models with or without sphingosine kinase 1 activity.
Genetic sphingosine kinase 1 deficiency significantly decreases synovial inflammation and joint erosions in murine TNF-alpha-induced arthritis.
No sample metadata fields
View SamplesIn order to identify genes dysregulated by the aberrant transcriptional activity of RUNX1-RUNX1T1, we used microarrays to determine the effect of this mutation on gene expression during myeloid and erythroid development of normal human progenitor cells.
Transcriptional dysregulation mediated by RUNX1-RUNX1T1 in normal human progenitor cells and in acute myeloid leukaemia.
Specimen part
View SamplesThis study supports an active role for PLZF and RAR-PLZF in leukemogenesis, identifies upregulation of CRABPI as a novel mechanism contributing to retinoid resistance and reveals the ability of the reciprocal fusion gene products to mediate distinct
RARalpha-PLZF overcomes PLZF-mediated repression of CRABPI, contributing to retinoid resistance in t(11;17) acute promyelocytic leukemia.
No sample metadata fields
View SamplesArabidopsis fc2-1 mutants fail to properly de-etiolate after a prolonged period in the dark. Our goal was to monitor whole genome expression during the first 2 hours of de-etiolation to determine the cuase of this growth arrest.
Ubiquitin facilitates a quality-control pathway that removes damaged chloroplasts.
Specimen part
View SamplesActivation of the Ras/Erk pathway upregulates expression of the Kruppel-like Factor 5 (KLF5) transcription factor, and KLF5 is a downstream mediator of Ras oncogenic signaling. Specifically, in bladder and colon cancer cell lines KLF5 upregulates the Ras-pathway target gene cyclin D1, and facilitates entry into the S phase of the cell cycle. Ras mutations are common in lung cancer, but a role for KLF5 in lung tumorigenesis has not been defined. To this end, we manipulated KLF5 expression in four Ras-mutant human lung adenocarcinoma cell lines to find that KLF5 significantly modulates anchorage-independent growth, a mutant Ras phenotype. However, in a mouse model of human lung adenocarcinoma, K-RasG12D does not critically require Klf5 to mediate oncogenesis or induce cyclin D1 expression.
Kruppel-like factor 5 is not required for K-RasG12D lung tumorigenesis, but represses ABCG2 expression and is associated with better disease-specific survival.
No sample metadata fields
View SamplesCXXC5 inhibits the canonical Wnt signaling pathway
Downregulation of the Wnt inhibitor CXXC5 predicts a better prognosis in acute myeloid leukemia.
Specimen part, Cell line
View SamplesThis SuperSeries is composed of the SubSeries listed below.
An international standardization programme towards the application of gene expression profiling in routine leukaemia diagnostics: the Microarray Innovations in LEukemia study prephase.
No sample metadata fields
View SamplesAn International Multi-Center Study to Define the Clinical Utility of MicroarrayBased Gene Expression Profiling in the Diagnosis and Sub-classification of Leukemia (MILE Study)
An international standardization programme towards the application of gene expression profiling in routine leukaemia diagnostics: the Microarray Innovations in LEukemia study prephase.
Disease
View Samples