SiRNAi knockdown of ZBTB4 in human SH-EP cells reveals Zbtb4 represses transcription of P21CIP1 and controls the cellular response to p53 activation
ABSTRACT: SH-EP cells were depleted for ZBTB4; Transfection of the cells were performed by oligofectamin RNAimax (Invitrogen) with siRNA smart pool (Dharmacon RNA Technologies); total RNA was isolated using RNAeasy Mini Kit (QIAGEN), preamplified, labeled with Cy3 and Cy5, respectively, and hybridized to cDNA-microarrays
Project description:Induction of apoptosis by the tumor suppressor p53 is known to protect from Myc-driven lymphomagenesis. The p53 family member p73 is also a pro-apoptotic protein, which is activated in response to oncogenes like Myc. We therefore investigated whether p73 provides a similar protection from Myc-driven lymphomas as p53. To generate B-cell lymphomas with defined genetic alterations in p53 or p73, we crossed the Eµ-Myc transgenic to mice heterozygous for germ-line deletions in p53 (p53+/) or p73 (p73+/-). Lymphomas which have spontaneously developed in Eµ-Myc transgenic animals with the genotypes p53+/+, p53+/-, p73+/+, p73+/- or p73-/- were isolated when the animals were moribund and further processed for gene expression profiling with 22.5K cDNA microarrays.
Project description:Total RNA was isolated from fresh-frozen tissue of 5 benign and 5 malignant pheochromocytomas. The reference consisted of laser microdissected (LMD) tissue from normal adrenal medullae from 10 different patients. After generating Cy3- and Cy5-fluorescently labeled cDNAs, F-chips containing 11,540 spots were hybridized. Data were analyzed with the IMAGENE 3.0 software.
Project description:Aim: identification of differentially expressed genes after LIN-9 depletion.<br> <br> hTERT immortalized human BJ fibroblasts were infected with pMSCV-Blasticidin based retroviruses encoding a shRNA which targets human LIN-9 and the empty vector respectively.<br> After 4 days of selection total RNA was isolated. Cy3 and Cy5 labelled cDNA probes were generated using the CyScribe Post-labelling Kit (Amersham Biosiences).
Project description:The p300 protein is one of more than 15 known mammalian HATs. We have used mice that carry a mutant allele of the p300 gene, which encodes a full-length protein that, however, lacks detectable acetyltransferase (AT)-activity and that acts in a dominant-negative manner. In addition, the allele is conditional, so that it can be expressed in a tissue-specific manner with the help of mice that express Cre-recombinase. Expression of the mutant allele specifically in B-cells, using CD19-cre mice, results in premature death with full penetrance from an autoimmune disease that closely mimics systemic lupus erythematosus (SLE) in its pathological manifestations.
Project description:To investigate the role of the dimerization interface for p53 function, we introduced modest charge-neutralizing (R181→L "EL") and more severe charge-inverting (R181→E "EE" and E180→R, R181→E "RE") mutations into the H1 helix of the full-length p53 molecule. To characterize the nuclear transcriptional function of p53 mutants in an unbiased manner we employed gene expression profiling using cDNA microarrays. Saos-2 cells were infected with adenoviruses expressing the p53 proteins EE, EL, WT and RE which span the entire spectrum of apoptotic activity. Total cellular RNA was isolated 18 hours after infection when apoptosis had not yet occurred.
Project description:Signaling pathways specifically induced by the oncogenic receptor Xmrk can be elucidated by comparing gene expression in cells with the activated receptor to gene expression in cells with the non-activated receptor. Mouse melanocytes transfected with an inducible version of the oncogenic receptor Xmrk, were starved for 72 h and afterwards stimulated with EGF. We compared cells stimulated for 15 minutes, 1 h, 2 h, 4 h, 8 h or 24 h, respectively, to unstimulated cells.
Project description:Analysis of Retinoblastoma protein (Rb) (Hs.408528) dependent transcriptional changes following siRNA mediated ablation of the RET finger protein/ Tripartite protein (RFP/TRIM27) (Hs.440382). Common reference design, two biological replicates with two technical replicates each.
Project description:Epithelial cells are the first cell type Salmonella Typhimurium will encounter when infecting a host. Using DNA array techology we identified the Salmonella enterica Typhimurium genes regulated inside human epithelial cells and their variation through time.