Project description:We used microarrays to detail the global gene transcription underlying T cells activation during the first 24 hours after stimulation. CD4+CD45RA+ T cells were sorted and cultured with different stimulatory conditions (Non-stimulated, anti-CD3 and anti-CD3 plus anti-CD28) for different times (0 hours, 4 hours and 24 hours). 3 replicates for each condition were analyzed.
Project description:We used microarrays to detail the global gene transcription effect of Dec1 underlying T cells activation during the first 24 hours after stimulation. CD4+CD62LhiCD25- T cells were sorted and cultured with different stimulatory conditions (anti-CD3 and anti-CD3 plus anti-CD28) for 24 hours. 4 replicates for each condition were analyzed.
Project description:Natural isolates of Burkholderia pseudomallei (Bp), the causative agent of melioidosis, are known to exhibit diverse phenotypic traits, suggesting significant intraspecies genetic heterogeneity. Using whole-genome Bp microarrays, we experimentally mapped patterns of large-scale genomic variation in 93 South East Asian clinical, environmental, and animal Bp isolates. 14% of the reference Bp K96243 genome was variably present across the strain panel, more than double previous estimates, and both hypothetical proteins and paralogous gene pairs (PGPs) were significantly over-represented in the set of strain-variable genes. Examining patterns of PGP retention and loss, we successfully sub-categorized the PGPs into non-redundant, functionally biased, and completely redundant classes. We then identified 20 novel regions (âislandsâ) variably present between strains previously missed by computational analysis. Three of these novel islands contained lipopolysaccharide (LPS) biosynthesis genes, and strains lacking one such LPS island demonstrated reduced virulence in mouse infection assays. Clinical isolates associated with human melioidosis were strongly associated with the presence of specific genomic islands, but a common set of virulence-related genes was present in all strains. Our results suggest that most Bp strains possess a core virulence machinery capable of causing disease, but accessory functions provided by mobile elements may predispose distinct host species and ecological niches to specific individual strains. This hierarchical model of Bp virulence reconciles previous conflicting studies comparing Bp environmental and clinical isolates, and suggests novel molecular strategies for disease surveillance and outbreak detection efforts in melioidosis. Keywords: aCGH of 93 Bp strains genomic DNA of 93 Bp strains were assessed on Bp_array_v2
Project description:We investigated the influence of genome position on propensity to amplify. First, we integrated a mutant form of DHFR into different positions in the human genome, challenged cells with methotrexate and then studied the genomic alterations arising in drug resistant cells. We observed site specific differences in methotrexate sensitivity, organization of amplicons and amplification frequency. One site was uniquely associated with a significantly enhanced propensity to amplify and recurrent amplicon boundaries, possibly implicating a rare folate sensitive fragile site in initiating amplification. Hierarchical clustering of gene expression patterns and subsequent gene enrichment analysis revealed two clusters differing significantly in expression of MYC target genes independent of integration site. We introduced a mutant form of DHFR (L22F), which confers greater resistance to methotrexate than the wild type (endogenous) gene into HCT116+chr3 cells and isolated independent clones containing DHFR* at different positions in the genome and identified genome sequences flanking the integration site of DHFR* using inverse PCR. For further analysis, we selected only clones, which were considered to have a single insertion of DHFR* by inverse PCR (13 independent insertion sites). The individual insertion site clones were further characterized with respect to genome copy number profiles. To select methotrexate resistant colonies, we exposed cells to a concentration of methotrexate that was three to four times the IC-50 for each integration site. Genomic copy number profiles were obtained for isolated resistant colonies (GSE6262) by using UCSF HumArray platform (GPL4421). Twelve methotrexate resistant colonies (four different integration sites) were selected for microarray analysis of gene expression at the mRNA level. The untreated integration site clone was used as the reference (Cy5 labeled cDNA) for each of the hybridizations with its respective resistant colonies (Cy3 labeled cDNA). Hybridizations were carried out on arrays of printed long oligonucleotides (70mers) containing 21,000 elements (Operon V2.0, printed in J. David Gladstone Institutes, Genomics Core Laboratory).
Project description:This project was aimed to identify the proteomic of pri-miR-31 interacting in naïve and activated cd4+ T cells. Immunoprecipitates pulled down by pri-miR-31 were subjected to LC-MS. Our results reveal the interacting protein in different state of CD4+ T cells.
Project description:A small number of tumor-derived cell lines have formed the mainstay of cancer therapeutic development, yielding drugs with impact typically measured as months to disease progression. To develop more effective breast cancer therapeutics, and more readily understand their potential clinical impact, we constructed a functional metabolic portrait of 46 independently-derived breast tumorigenic cell lines, contrasted with purified normal breast epithelial subsets, freshly isolated pleural effusion breast tumor samples and culture-adapted, non-tumorigenic mammary epithelial cell derivatives. We report our analysis of glutamine uptake, dependence, and identification of a significant subset of triple negative samples that are glutamine auxotrophs. This NCBI GEO submission comprises a small datasest generated to compare the expression profiles of the above nontumorigenic, purified normal and purified pleural effusion samples with 10 established breast cancer-derived cell lines. This dataset was subsequently merged with a previously published expression dataset derived from 45 independent breast cancer derived cell lines (Neve, et al 2006), and analyses contrasting various subsets of the merged dataset were published. Expression data from 26 samples, no replicates: purified normal human mammary epithelial breast cellular subsets CD10+, BerEP4+, and remaining stromal cell samples from 3 independent anonymous donors; 3 anonymous purified human breast cancer pleural effusion samples; 4 HMEC-derived culture adapted but not transformed samples (184A1, 184B5, HMLE, HMLE-PR); and 10 established human breast cancer cell lines.
Project description:We aim to find the gene-specific effects of Rnmt KO in mouse CD4 T cells. TMT proteomics datasets were generated to find out which proteins are dependent on RNMT for their expression.
Project description:The ability to detect and isolate bGL1-22/LGL1âspecific human type II NKT cells allowed us to compare the global gene expression profiles of these cells with type I NKT cells using microarray analysis. Principal component analysis revealed that the gene expression profile signature for bGL1-22 and LGL1-specific T cells both before and after activation with anti-CD3/CD28 beads is distinct from that of type I NKT cells. RNA from CD1d tetramer-sorted b-GL122/LGL1âspecific T cells or iNKT cells was amplified, labeled, and hybridized on the Affymetrix Human Genome U133 Plus 2.0 microarray chips. The data were analyzed with GeneSpring GX 12.5 (Agilent Technologies)