Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression.
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs. One-condition experment, gene expression of 3A6
Project description:The tumor suppressor p53 can induce various biological responses. Yet it is not clear whether it is p53 in vivo promoter selectivity that triggers different transcription programs leading to different outcomes. Our analysis of genome-wide chromatin occupancy by p53 using ChIP-seq (deposited in Sequence Read Archive database as SRP007261) revealed “p53 default program”, i.e. the pattern of major p53-bound sites that is similar upon p53 activation by nutlin3a, RITA or 5-FU in breast cancer cells, despite different biological outcomes triggered by these compounds. Parallel analysis of gene expression allowed identification of 280 previously unknown p53 target genes, including p53-repressed AURKA. The consensus p53 binding motif was present more frequently in p53-induced, than in repressed targets, indicating different mechanisms of gene activation versus repression. We identified several possible cofactors of p53, and found that STAT3 antagonised p53-mediated repression of a subset of genes, including AURKA. Finally, we showed that the expression of the novel p53 targets correlates with p53 status and survival in breast cancer patients. We used microarrays to detail the global programme of gene expression changed upon nutlin3a treatment and knocking-down of STAT3 transcription program MCF7 cell with or without knock-down of STAT3 were treated with nutlin3a and collected for RNA extraction and hybridization on Affymetrix microarrays.
Project description:Gene methylation profiling of immortalized human mesenchymal stem cells comparing HPV E6/E7-transfected MSCs cells with human telomerase reverse transcriptase (hTERT)- and HPV E6/E7-transfected MSCs. hTERT may increase gene methylation in MSCs. Goal was to determine the effects of different transfected genes on global gene methylation in MSCs.
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression. Two-condition experiment, Normoxic MSCs vs. Hypoxic MSCs.
Project description:Kynureninase is a member of a large family of catalytically diverse but structurally homologous pyridoxal 5'-phosphate (PLP) dependent enzymes known as the aspartate aminotransferase superfamily or alpha-family. The Homo sapiens and other eukaryotic constitutive kynureninases preferentially catalyze the hydrolytic cleavage of 3-hydroxy-l-kynurenine to produce 3-hydroxyanthranilate and l-alanine, while l-kynurenine is the substrate of many prokaryotic inducible kynureninases. The human enzyme was cloned with an N-terminal hexahistidine tag, expressed, and purified from a bacterial expression system using Ni metal ion affinity chromatography. Kinetic characterization of the recombinant enzyme reveals classic Michaelis-Menten behavior, with a Km of 28.3 +/- 1.9 microM and a specific activity of 1.75 micromol min-1 mg-1 for 3-hydroxy-dl-kynurenine. Crystals of recombinant kynureninase that diffracted to 2.0 A were obtained, and the atomic structure of the PLP-bound holoenzyme was determined by molecular replacement using the Pseudomonas fluorescens kynureninase structure (PDB entry 1qz9) as the phasing model. A structural superposition with the P. fluorescens kynureninase revealed that these two structures resemble the "open" and "closed" conformations of aspartate aminotransferase. The comparison illustrates the dynamic nature of these proteins' small domains and reveals a role for Arg-434 similar to its role in other AAT alpha-family members. Docking of 3-hydroxy-l-kynurenine into the human kynureninase active site suggests that Asn-333 and His-102 are involved in substrate binding and molecular discrimination between inducible and constitutive kynureninase substrates.