Project description:CK1 enzymes are conserved, acidophilic serine/threonine kinases with a variety of critical cellular functions; their misregulation contributes to cancer, neurodegenerative diseases, and sleep phase disorders. Here, we describe a new mechanism of CK1 regulation conserved from yeast to human – autophosphorylation of a threonine in the mobile L-EF loop proximal to the active site – that inhibits kinase activity. Consequently, yeast and human CK1 enzymes with phosphoablating mutations at this site are hyperactive in vitro. We used quantitative phosphoproteomics to show that disruption of this regulatory mechanism rewires CK1 signaling in Schizosaccharomyces pombe. In accord, we found that a known CK1 pathway, a mitotic checkpoint, is downregulated in these mutants, while new pathways that confer heat shock resistance and suppress meiotic transcripts are upregulated. Molecular dynamics simulations demonstrated that phosphorylation on the L-EF loop alters the conformation of the substrate docking site, and we propose that this affects which CK1 substrates can be phosphorylated. Due to the functional importance of the L-EF loop, which is unique to the CK1 family of kinases, this mechanism is likely to regulate the majority of CK1 enzymes in vivo.
Project description:One of the main hallmarks of aging is aging-associated inflammation, also known as inflammaging. In this study, by comparing young and old mice using LC-MS profiling, we show that immunoglobulins are the proteins that are most increased with age in plasma. This observation appears to have been previously overlooked as many of the aging proteome profiling experiments to date use targeted antibody/aptamer-based arrays, which typically do not include high abundance proteins or use depletion prior to LC-MS to remove the high abundant proteins. Through LC-MS profiling of young and old mouse kidneys, we also show that immunoglobulins are among the top proteins changing with age. Immunofluorescence staining of kidney sections shows that the main increases in immunoglobulins with age are localized in the glomeruli of the kidney. Using laser capture microdissection coupled with LC-MS, we show an increase in multiple immune related proteins in glomeruli from aged mice. Increased deposition of immunoglobulins and complement proteins in the kidney glomeruli may be a factor leading to reduced filtering capacity of the kidney with age.
Project description:Comparison at two different timepoints of expression profiles of wild_type Bacillus and a deletion mutant of the PlcR regulator. Additional processed data can be found in the FTP directory for this experiment <a href="ftp://ftp.ebi.ac.uk/pub/databases/microarray/data/experiment/MEXP/E-MEXP-1472/">ftp://ftp.ebi.ac.uk/pub/databases/microarray/data/experiment/MEXP/E-MEXP-1472/</a>
Project description:The mammalian neocortex comprises an enormous diversity regarding cell types, morphology, and connectivity. In this work, we discover a post-transcriptional mechanism of gene expression regulation, protein translation, as a determinant of cortical neuron identity. We find specific upregulation of protein synthesis in the progenitors of later-born neurons and show that translation rates and concomitantly protein half-lives are inherent features of cortical neuron subtypes. In a small molecule screening, we identify Ire1a as a regulator of Satb2 expression and neuronal polarity. In the developing brain, Ire1a regulates global translation rates, coordinates ribosome traffic, and the expression of eIF4A1. Furthermore, we demonstrate that the Satb2 mRNA translation requires eIF4A1 helicase activity towards its 5’-untranslated region. Altogether, we show that cortical neuron diversity is generated by mechanisms operating beyond gene transcription, with Ire1a-safeguarded proteostasis serving as an essential regulator of brain development.
Project description:rs08-03_glutathion - glutathion - How does glutathione content or reduction state affect H2O2-induced changes in the transcriptome? - Three single Arabidopsis mutants were used: cat2, knockout for catalase2 and so enriched in H2O2; cad2, defective in glutathione content; cytGR, knockout for cytosolic glutathione reductase. Cat2 was crossed with cad2 and cyt GR, and col0, 3 single mutants, and 2 double mutants were sampled in controlled growth conditions either in 8h or 16h photoperiod. Keywords: gene knock out 20 dye-swap pairs - CATMA arrays: 40 arrays
Project description:GABAergic transmission controls cortical sleep, however the molecular determinants of GABAergic control of this phenomenon are unknown. We used immunoprecipitation of the inhibitory postsynaptic protein gephyrin to identify gephyrin interactors at ZT6 (sleep) and ZT6 with sleep deprivation (ZT6SD).
Project description:rs09-04_plc_2 - edelfosine and wortmannin effect at 22°c and 4°c - Is there an overlap between PI4-kinase and phospholipase C signaling in cold response ? - Experiments were carried out with Columbia suspension cells. Cells were treated or not with edelfosine (PLC inhibitor) or wortmannin (PI4-kinase inhibitor). These agents were added 15 min before cold stress. ARN were extracted 4h after stress. Controls were made at 22°C. Keywords: treated vs untreated comparison 11 dye-swap - CATMA arrays
Project description:Plants are subjected to perpetual fluctuations of light intensity and spectral composition in their natural growth environment, particularly due to movement of clouds and upper canopy leaves. Sudden exposure to intense light is accompanied by absorption of excess light energy, which results in an overload of photosynthetic electron transport chain and generation of reactive oxygen species in and around thylakoid membranes. To cope with this photooxidative stress and to prevent chronic photoinhibition under dynamically changing light intensities, plants have evolved various short- and long-term photoprotective mechanisms. We used quantitative mass spectrometry to investigate long-term acclimation of Arabidopsis thaliana leaf proteome to fluctuating light (FL) which induces photooxidative stress. After three days of FL exposure the proteomes of young and mature leaves were analyzed separately in the morning and at the end of day to examine possible interaction between FL acclimation and leaf development or time of day.