Transcription profiling of mouse aorta (whole or regions) from mice of different embryonic ages
ABSTRACT: In order to identify novel regulators of haematopoietic stem cell emergence in the mouse embryonic AGM region, 3 different types of gene expression comparisons were performed. In the first one, whole dorsal aortas from embryonic day (E) 11 embryos and E9 embryos were dissected, tissues from each time point were pooled and their expression profile compared (whole aorta (WA) E9 vs E11). 3 different pools/biological replicas were obtained for each time point, a dye swap was included in each experiment, a technical replicate was performed for each pool (including the dye swap) and 2 self-self hybridisation controls were included. In the second comparison, GFP+ cells were isolated by fluorescence-activated cell sorting from pooled E9 and E11 aortas from transgenic embryos that express GFP under the regulatory elements of the Ly-6A gene. We obtained one pool for each time point and compared their transcription profile (GFP E9 vs E11). This comparison also contained a dye swap and a technical replicate (also of the dye swap). In the third comparison, the E11 aorta was cut into 3 roughly equal parts. Pools were obtained of the middle region ("m") and the two outer regions, rostral and caudal, together ("r+c") and their expression profile compared (11AO m vs r+c). There was one pool of each population, and the experiment included a dye swap and a technical replicate (also of the dye swap) and one self-self hybridisation control. Isolated RNA was amplified by T7-mediated in vitro transcription, with one round for the first and the third comparison and two rounds for the second comparison (sorted GFP+ cells). Probes were labelled during the reverse transcription step. Within array normalisation was achieved with Lowess Smooth and between arrays with Z scores. ANOVA analysis identified differentially expressed genes and K means clustering was used to segregate them into upregulated and downregulated genes.
The first adult-repopulating hematopoietic stem cells (HSCs) are detected starting at day 10.5 of gestation in the aorta-gonads-mesonephros (AGM) region of the mouse embryo. Despite the importance of the AGM in initiating HSC production, very little is currently known about the regulators that control HSC emergence in this region. We have therefore further defined the location of HSCs in the AGM and incorporated this information into a spatial and temporal comparative gene expression analysis of ...[more]
Project description:To understand how metabolic and nutritional factors governing adaptation to the host niche contribute to the virulence of Aspergillus fumigatus we compared transcriptomes of developmentally matched A.fumigatus isolates following laboratory culture or initiation of infection in the neutropenic murine lung.
Project description:The presence of Set2-mediated methylation of H3K36 (K36me) correlates with transcription frequency throughout the yeast genome. K36me targets the Rpd3S complex to deacetylate transcribed regions and suppress cryptic transcription initiation at certain genes. Here, using a genome-wide approach, we report that the Set2-Rpd3S pathway is generally required for controlling acetylation at coding regions. When using acetylation as a functional read-out for this pathway, we discovered that longer genes and, surprisingly, genes transcribed at lower frequency exhibit a stronger dependency. Moreover, a systematic screen using high resolution tiling microarrays allowed us to identify a group of genes that rely on Set2-Rpd3S to suppress spurious transcripts. Interestingly, most of these genes are within the group that depend the same pathway to maintain a hypo-acetylated state at coding regions. These data highlight the importance of using the functional readout of histone codes to define the roles of specific pathways.
Project description:The Goal of this study is to explore the transcriptional differences between angiogenic and non-angiogenic tumors generated in Dr. Folkman's Lab, Harvard Medical School.<br>Referneces:<br>Almog N, Henke V, Flores L, Hlatky L, Kung AL, Wright RD, Berger R, Hutchinson L, Naumov GN, Bender E, Akslen LA, Achilles EG, Folkman J.<br>Prolonged dormancy of human liposarcoma is associated with impaired tumor angiogenesis.<br>FASEB J. 2006 May;20(7):947-9.<br><br>Naumov GN, Bender E, Zurakowski D, Kang SY, Sampson D, Flynn E, Watnick RS, Straume O, Akslen LA, Folkman J, Almog N.<br>A model of human tumor dormancy: an angiogenic switch from the nonangiogenic phenotype.<br>J Natl Cancer Inst. 2006 Mar 1;98(5):316-25.<br>
Project description:The aim of the experiment was to identify novel genes with relevance to cellular cholesterol metabolism. HeLa cells were cultivated while sterol was depleted from the medium by applying 2-Hydroxypropyl-cyclodextrin. RNA for expression profiling was isolated 3h, 4.5h and 6h after depletion.
Project description:One major effect of PI3-kinase activation downstream of the serine/threonine kinase Akt is the phosphorylation of the transcription factor FOXO1 and its neutralization. FOXO1 has several ubiquitous targets genes in many cell types that control cell quiescence, oxydative stress or apoptosis. However, it has been demonstrated that FOXO1 also has specific targets depending of the cellular context. The role of FOXO1 to regulate specific genes in T lymphocytes has not been investigated yet. To examine this point, we used the CD4+ leukemia Jurkat T-cell line, in which the PI3K pathway is constitutively turned-on and FOXO1 transcriptional activity strongly repressed. These cells were transduced with lentiviruses coding for a constitutively active form of FOXO1 fused to GFP and having the three AKT phosphorylation sites mutated to alanine (FOXO1-AAA-GFP) to restore its transcriptional activity. GFP-transduced cells were used as a control and the gene activation levels in the two cell populations analyzed 48 hours post-infection.
Project description:Normal human dermal fibroblasts (NHDF) and human lung microvascular endothelial cells (HMVEC-L) were irradiated with iron ions (0, 0.2, 0.4 and 1 Gy, 1GEv/n) at Brookhaven National Labs (BNL). Aim of the study is to find differentially transcribed genes in dependance of radiation dose/source and cell type.
Project description:Normal human dermal fibroblasts (NHDF) and human lung microvascular endothelial cells (HMVEC-L) were irradiated with protons (0, 0.5, 1 and 2 Gy, 1GEv/n) at Brookhaven National Labs (BNL). Aim of the study is to find differentially transcribed genes in dependence of radiation dose/source and cell type.
Project description:To establish the effect of CPT in the regulation of global transcription in S. cerevisiae, we have used a yeast TOP1 null strain, JEL1Δtop1, bearing a low-copy number plasmid that expresses, under the control of the yeast TOP1 promoter, either a wild-type (wt) yeast Top1p (pCC10) or an inactive Y727F mutant enzyme (pAR7).We determined global transcript levels upon CPT treatments of JEL1Δtop1 cells expressing either a yeast wt or Y727F mutant Top1p.
Project description:Perineural invasion (PNI) is considered to be a specific path for the spread of pancreatic cancer cells and PNI is thought to be one of the important factors that determine local recurrence after resection. In addition PNI has been implicated in the pain syndrome that affects the majority of pancreatic ductal adenocarcinoma (PDAC) patients. Here we aimed to investigate the transcriptional differences between neuro-invasive tumors engineered in-vitro compared to less invasive parental cells. Two colour differential hybridisations were performed with human universal reference total RNA (Clontech, #636538).