Project description:ING1b and GADD45a are nuclear proteins involved in the regulation of cell growth, apoptosis and DNA repair. We found that ING1b and GADD45a physically and functionally interact in the epigenetic regulation of specific target genes. In order to study this interaction further, we analysed the transcriptional changes in MEF cells from single and double Ing1/Gadd45 knockout mice via microarray profiling. Mouse embryonic fibroblasts (MEF cells) were isolated from embryonic day E15.5 male embryos, either wild-type (WT) or knockout for Ing1 (Ing1-/-), Gadd45a (Gadd45a-/-) or Ing1/Gadd45a (double knockout, DKO), and cultured for 3 passages. Samples were then collected in duplicates per MEF line for expression array profiling.
Project description:Immunoprecipitation of GNAQ protein by immunoprecipitation with two different antibodies from Santa Cruz Biotechnology (C-19 and E-17) in isolated mitochondria from different cell lines. The cell lines used include: Mouse embryonic fibroblasts (MEF) wild-type (WT), GNAQ knockout (KO) and knockout expressing GNAQ WT (KO_Gq). NIH3T3 cells were also used.
Project description:Purpose: To determine transcriptome profile of the facial processes of Wnt9b, Rspo2 and double gene knockout embryos by NG RNA sequencing Method: Embryonic facial process RNA samples isolated from wild type, Rspo2, Wnt9b and Rspo2;Wnt9b double mutants in triplicate (except wild type (n=2)) at embryonic day 10.5 Results: single-end 50bp reads, 19.8 million reads - 30.7 million reads.
Project description:Fibrillins, which are extensively O-glucosylated by POGLUT2 and 3, are crucial for development and cardiopulmonary function. Using primary dermal fibroblasts derived from our POGLUT 2 and 3 double knockout mouse model combined with proteomics analyses, we show that loss of O-glucose modifications on fibrillin causes compositional changes in the extracellular environment compared to controls. Notably, a 30-50% reduction in the abundance of FBN1 and 2 was found in media samples whereas an 85% reduction was found in the extracellular matrix. Other proteins in the extracellular matrix also exhibited changes in abundance in the double knockout samples compared to wild type, but it is still unclear if these changes are simply secondary to the significant reduction in fibrillins.
Project description:To study the role of GADD45 proteins in early mouse development, RNA-seq expression profiling was performed with Gadd45a/Gadd45b double knockout (DKO) and wild-type (WT) mouse embryos at 2-cell stage (2C).
Project description:The daily organisation of most mammalian cellular functions is attributed to circadian regulation of clock-controlled protein expression, driven by daily cycles of CRYPTOCHROME-dependent transcriptional feedback repression. To test this, we used quantitative mass spectrometry to compare wild type and CRY-deficient fibroblasts under constant conditions. In CRY-deficient cells, we found that temporal variation in protein, phosphopeptide, and K+ abundance was at least as great as wild type controls. Most strikingly. the extent of temporal variation within either genotype was much smaller than overall differences in proteome composition between WT and CRY-deficient cells. This proteome imbalance in CRY-deficient cells and tissues was associated with increased susceptibility to proteotoxic stress, which impairs circadian robustness, and may contribute to the wide-ranging phenotypes of CRY-deficient mice. Rather than generating largescale daily variation in proteome composition, we suggest it is plausible that the various transcriptional and post-translational functions of CRY proteins ultimately act to promote protein and osmotic homeostasis against daily perturbation.
Project description:ATAC-Seq of wild-type and nm1-knockout mouse embryonic fibroblasts to investigate the impact of nm1 loss on chromatin accessibility
