Project description:NIH/3T3 cells were cultured at least 2 weeks in light medium: DMEM for SILAC (Thermo scientific) supplemented with PS, dialyzed FBS (Thermo scientific), (light) L-Lysine-2HCl (0.666 mM), (light) L-Arginine-HCl (0.399 mM)44, and L-Proline (200 mg/L). Before switching from light to heavy medium, cells were treated with 0.05 µg/mL CHX or equivalent dilution of DMSO in 6-well plates. At timepoint 0 h, we replaced light medium with pre-warmed heavy medium: DMEM for SILAC (Thermo scientific) supplemented with PS, dialyzed FBS (Thermo scientific), (heavy) 13C6 15N2 L-Lysine-2HCl (0.666 mM), (heavy) 13C6 15N4 L-Arginine-HCl (0.399 mM), and L-Proline (200 mg/L) for NIH/3T3.
Project description:In order to identify how MnTE-2-PyP affects p300 association to chromatin genome-wide, we performed a p300 chromatin Immunoprecipitation assay followed by Next Generation Sequencing on PC3 cells treated with or without MnTE-2-PyP one hour post-irradiation (Figure 3A). Based on the called peaks near genes, we predicted that HIF-1βand CREB transcription factors were associating DNA less in the presence of MnTE-2-PyP. DNA was ChIP-Fixed from Pc3 cells treated with 20 Gy radiation and with and without T2E drug. There are 2 biological replicates of PC3 untreated cells and 3 biological replicates of PC3 cells treated with MnTE-2-PyP. There are two corresponding input samples for the biological replicates.
Project description:Fission yeast CSL mutants (cbf11 and cbf12 knock-outs) show altered sensitivity to hydrogen peroxide. To help elucidate the molecular basis of this phenomenon, we analyzed wild-type and mutant transcriptomes before stress and as a timecourse after treatment with hydrogen peroxide. Cells were grown to exponential phase in YES medium, untreated samples were collected (time 0), cells were treated with 0.74 mM hydrogen peroxide and harvested after 15 and 60 min post treatment. Total RNA was isolated, labelled and hybridized to custom-made two-colour microarrays. Each sample was hybridized against reference created by pooling equal aliquots from all 9 samples.
Project description:Co-IP/MS of Gli3 from NIH/3T3 cells treated with the smoothened agonist SAG fractionated into nuclear and cytoplasmic fractions. 10091, 10092, 10093, 10095, 10096 are nuclear fraction samples, 10094, 10097 are cytoplasmic fraction samples.
Project description:Ossification of the posterior longitudinal ligament (OPLL) is formed by heterogeneous ossification of posterior longitudinal ligament. The patho-mechanism of OPLL is still largely unknown. Recently, disorders of metabolism are thought to be the center of many diseases such as OPLL. Advanced glycation end product (AGE) are accumulated in many extracellular matrixes such as ligament fibers, and it can functions as cellular signal through its receptor (RAGE), contributing to various events such as atherosclerosis or oxidative stress. However, its role in OPLL formation is not yet known. Therefore, we performed high-through-put RNA sequencing on primary posterior longitudinal ligament cells treated with different doses of AGEs (1µM, 5µM and negative control), with or without BMP2 (1µM). mRNA profiles of Primary human posterior longitudinal ligament cells stimulated with various stimuli (Control, 1µM AGE-BSA, 5µM AGE-BSA, 1µM AGE-BSA with BMP2, 5µM AGE-BSA with BMP2) were generated by deep sequencing on Ion Proton