Transcription can be quite disruptive for chromatin so cells have evolved mechanisms to preserve chromatin integrity during transcription, hence preventing the emergence of cryptic transcript from spurious promoter sequences. How these transcripts are regulated and processed by cells remains poorly characterized. Notably, very little is known about the termination of cryptic transcription. Here we used RNA-Seq to identify and characterize cryptic transcripts in Spt6 mutant cells (spt6-1004) in Saccharomyces cerevisiae. We found polyadenylated cryptic transcripts running both sense and anti-sense relative to genes in this mutant. Cryptic promoters were enriched for TATA boxes, suggesting that the underlying DNA sequence defines the location of cryptic promoters. While intragenic sense cryptic transcripts terminate at the terminator of the genes that host them, we found that anti-sense cryptic transcripts preferentially terminate at the 3’-end of upstream genes. These findings led us to demonstrate that most terminators in yeast are bidirectional, leading to termination and polyadenylation of transcripts coming from either direction. We propose that S. cerevisiae has evolved this mechanism in order to prevent spurious transcription from invading neighbouring genes, a feature particularly critical for organisms with small compact genomes. Overall design: Cells from spt16-1004 and its respective WT strain were grown to an OD600 of 0.5 at 30°C and shifted to 37°C for 80 min before RNA extraction. Total RNA was extracted using the hot phenol method. Prior to library preparation, total RNA was either depleted for ribosomal RNA using the Ribo-zero Gold yeast kit (Epicentre-Illumina) or enriched for polyadenylated RNA using the NEBnext Poly(A) kit (New England Biolabs). Strand specific RNA-seq libraries were prepared using the KAPA stranded RNA-Seq library preparation kit prior to paired-end sequencing on an Illumina Hi-Seq2000. Reads were mapped to the sacCer3 assembly of the S. cerevisiae genome using Tophat2 (23). Intron length range was set at 50 to 1000 bp and a reference annotation file was provided to guide the assembly.
Bidirectional terminators in Saccharomyces cerevisiae prevent cryptic transcription from invading neighboring genes.
Cell line, SubjectView Samples
Most of the breast cancer samples used in clinical research contain multiple cell types other than epithelial cells alone. The non-epithelial cell types have have a substantial effect on the gene expression-profile, which is used to define molecular subtypes of the tumours. The purpose of this data set is to retrieve gene-expression profile within tumour epithelial cells. We collected 9 breast cancer epithelial cell lines and 5 tumour sampes from which epithelial cells were sorted and enriched using BerEp4 antibody coated beads. We profiled the mRNA expression level of these samples and classified probe sets into epithelial genes which were those genes with present calls in at least 50% of the samples. Then we derived an 23-gene signature based on only the epithelial genes to stratify breast cancer.
Minimising immunohistochemical false negative ER classification using a complementary 23 gene expression signature of ER status.
Specimen partView Samples
The clonal CD3- CD4+ Th2 cell population characterizing some hypereosinophilic syndrome patients stably endures for years provoking a chronic inflammatory skin disease, with a subgroup of patients ultimately progressing to T-cell lymphoma. The aim of this study is the identification of the molecular changes (1) associated with the persistence of the pre-malignant clone (2) associated with the activation of co-stimulatory receptors and (3) associated with the emergence of malignant T-cell subclones.
Molecular profiling of CD3-CD4+ T cells from patients with the lymphocytic variant of hypereosinophilic syndrome reveals targeting of growth control pathways.
Specimen part, Disease, Disease stage, TimeView Samples
Triple-Negative breast cancer (TNBC) is an aggressive subtype of breast cancer that is associated with poor prognosis due to its propensity to form metastases. Unfortunately, the current treatment options are limited to chemotherapy such that identification of actionable targets are needed. The receptor tyrosine kinase AXL plays a role in the tumor cell dissemination and its expression in TNBC correlates with poor patients? survival. Here, we explored whether exploiting an AXL knockdown gene signature in TNBC cells may offer an opportunity for drug repurposing. To this end, we queried the PharmacoGx pharmacogenomics platform with an AXL gene signature which revealed Phenothiazines, a class of Dopamine Receptors antagonists (Thioridazine, Fluphenazine and Trifluoperazine) typically used as anti-psychotics. We next tested if drugs may be active to limit growth and metastatic progression of TNBC cells, similarly to AXL depletion. We found that the Phenothiazines were able to reduce cel l invasion, proliferation and viability, and also increased apoptosis of TNBC cells in vitro. Mechanistically, these drugs did not affect AXL activity but instead reduced PI3K/AKT/mTOR and ERK signaling. When administered to mice bearing TNBC xenografts, these drugs showed were able to reduce tumor growth and metastatic burden. Collectively, these results suggest that these antipsychotics are novel anti-tumor and anti-metastatic agents that could potentially be repurposed, in combination with standard chemotherapy, for use in TNBC. Overall design: RNA-seq of the Triple Negative Breast Cancer cell line MDA-MB-231 treated with siCt or siAXL Differential gene expression profile between MDA-MB-231 siCt and siAXL by RNA sequencing (Illumina HiSEq 2000)
AXL knockdown gene signature reveals a drug repurposing opportunity for a class of antipsychotics to reduce growth and metastasis of triple-negative breast cancer.
Cell line, Treatment, SubjectView Samples
Integrated DNA and expression array analysis in primary human breast tumors identified chromosome 8q22 copy number gain and a suite of over-expressed genes in this region highly relevant to subsequent recurrence.
Amplification of LAPTM4B and YWHAZ contributes to chemotherapy resistance and recurrence of breast cancer.
Age, Specimen part, SubjectView Samples
Decreased mitochondrial mass and function in muscle of diabetic patients is associated with low PGC-1alpha, a transcriptional coactivator of the mitochondrial gene program. To investigate whether reduced PGC-1alpha and oxidative capacity in muscle directly contributes to age-related glucose intolerance, we compared the genetic signatures and metabolic profiles of aging mice lacking muscle PGC-1alpha. Microarray analysis revealed that a significant proportion of PGC-1alpha-dependent changes in gene expression overlapped with age-associated effects, and aging muscle and muscle lacking PGC-1alpha shared gene signatures of impaired electron transport chain activity and TGFbeta signalling.
Loss of Pgc-1α expression in aging mouse muscle potentiates glucose intolerance and systemic inflammation.
Specimen partView Samples
The loss of E-cadherin causes dysfunction of the cell-cell junction machinery, which is an initial step in epithelial-to-mesenchymal transition (EMT), facilitating cancer cell invasion and the formation of metastases. A set of transcriptional repressors of E-cadherin (CDH1) gene expression, including Snail1, Snail2 and Zeb2 mediate E-cadherin down-regulation in breast cancer. However, the molecular mechanisms underlying the control of E-cadherin expression in breast cancer progression remain largely unknown. Here, by using global gene expression approaches, we uncover a novel function for Cdc42 GTPase-activating protein (CdGAP) in the regulation of expression of genes involved in EMT. We found that CdGAP used its proline-rich domain to form a functional complex with Zeb2 to mediate the repression of E-cadherin expression in ErbB2-transformed breast cancer cells. Conversely, knockdown of CdGAP expression led to a decrease of the transcriptional repressors Snail1 and Zeb2, and this correlated with an increase in E-cadherin levels, restoration of cell-cell junctions, and epithelial-like morphological changes. In vivo, loss of CdGAP in ErbB2-transformed breast cancer cells impaired tumor growth and suppressed metastasis to lungs. Finally, CdGAP was highly expressed in basal-type breast cancer cells, and its strong expression correlated with poor prognosis in breast cancer patients. Together, these data support a previously unknown nuclear function for CdGAP where it cooperates in a GAP-independent manner with transcriptional repressors to function as a critical modulator of breast cancer through repression of E-cadherin transcription. Targeting Zeb2-CdGAP interactions may represent novel therapeutic opportunities for breast cancer treatment. Overall design: Total RNA profiles of ErbB2-expressing control mammary tumor explants cells (shCON) and CdGAP-depleted cells (shCdGAP) were generated by deep sequencing, in triplicate, using Illumina HiSEq2000.
The Cdc42/Rac1 regulator CdGAP is a novel E-cadherin transcriptional co-repressor with Zeb2 in breast cancer.
Specimen part, SubjectView Samples
Comparison of Chronic Lymphocytic Leukemia patients expressing high or low levels of ZAP70 mRNA: prognostic factors and interaction with the microenvironment.
Gene expression profiling reveals differences in microenvironment interaction between patients with chronic lymphocytic leukemia expressing high versus low ZAP70 mRNA.
Sex, AgeView Samples
Background: We have previously shown that the Gene expression Grade Index (GGI) was able to identify two subtypes of estrogen receptor (ER)-positive tumors that were associated with statistically distinct clinical outcomes in both untreated and tamoxifen-treated patients. Here, we aim to investigate the ability of the GGI to predict relapses in postmenopausal women who were treated with tamoxifen (T) or letrozole (L) within the BIG 1-98 trial.
The Gene expression Grade Index: a potential predictor of relapse for endocrine-treated breast cancer patients in the BIG 1-98 trial.
Age, Specimen part, Disease stage, TreatmentView Samples
Microarrays have revolutionized breast cancer (BC) research by enabling studies of gene expression on a transcriptome-wide scale. Recently, RNA-Sequencing (RNA-Seq) has emerged as an alternative for precise readouts of the transcriptome. To date, no study has compared the ability of the two technologies to quantify clinically relevant individual genes and microarray-derived gene expression signatures (GES) in a set of BC samples encompassing the known molecular BC's subtypes. To accomplish this, the RNA from 57 BCs representing the four main molecular subtypes (triple negative, HER2 positive, luminal A, luminal B), was profiled with Affymetrix HG-U133 Plus 2.0 chips and sequenced using the Illumina HiSeq 2000 platform. The correlations of three clinically relevant BC genes, six molecular subtype classifiers, and a selection of 21 GES were evaluated.
Transfer of clinically relevant gene expression signatures in breast cancer: from Affymetrix microarray to Illumina RNA-Sequencing technology.
Specimen part, Disease stageView Samples