Project description:Analysis of genes and biological processes influenced by Neuronal calcium sensor 1 (NCS1) based on whole transcriptome analyiss of Ncs1+/+ (wildtype) and Ncs1-/- (knockout) mouse brain tissues, i.d. the frontal cortex and the hippocampus.
Project description:This project investigates the conformational dynamics of the ASK1(269-658) protein fragment and the structural changes induced by its interaction with the small molecule H43. Using hydrogen-deuterium exchange mass spectrometry (HDX-MS), we compared the deuterium uptake profiles of apo ASK1(269-658) and its complex with H43. Samples were prepared with different exchange times (30, 90, 600, and 1800 seconds) in D₂O buffer, quenched, enzymatically digested with immobilized pepsin, and analyzed by LC-MS on a Thermo Orbitrap QE mass spectrometer. Peptide identification was performed using PEAKS Online, and HDX data analysis was conducted with HDExaminer software. The results provide insights into dynamic regions within ASK1(269-658) and reveal ligand-induced conformational changes.
Project description:Metastases in the bone marrow (BM) are grim prognostic factors in patients with neuroblastoma (NB). In spite of extensive analysis of primary tumor cells from high- and low-risk NB patients, a characterization of freshly isolated BM-infiltrating metastatic NB cells is still lacking. Our aim was to identify proteins specifically expressed by metastatic NB cells, that may be relevant for prognostic and therapeutic purposes. Metastatic NB cells were freshly isolated from patients’ BM by positive immunomagnetic bead manipulation using anti-GD2 monoclonal antibody. Unselected BM samples from patients with metastatic NB were also included. Gene expression profiles were compared with those obtained from archived NB primary tumors from patients with 5y-follow-up. After validation by RT-qPCR, expression/secretion of the proteins encoded by the up-regulated genes in the BM-infiltrating NB cells was evaluated by flow cytometry and ELISA. Compared to primary tumor cells, BM-infiltrating NB cells down-modulated the expression of CX3CL1, AGT, ATP1A2 mRNAs, whereas they up-regulated several genes commonly expressed by various lineages of BM resident cells. BM-infiltrating NB cells expressed indeed the proteins encoded by the top-ranked genes, S100A8 and A9 (calprotectin), CD177 and CD3, and secreted the CXCL7 chemokine. BM-infiltrating NB cells also expressed CD271 and HLA-G. We have identified proteins specifically expressed by BM-infiltrating NB cells. Among them, calprotectin, a potent inflammatory protein, and HLA-G, endowed with tolerogenic properties facilitating tumor escape from host immune response, may represent novel biomarkers and/or targets for therapeutic intervention in high-risk NB patients.