Project description:Methods for direct capturing global substrates of protein-modifying enzymes in living cells are with many challenges, and yet largely unexplored. Here, we report a strategy to directly capture substrates of protein-modifying enzymes via PTM-acceptor residue crosslinking in living cells, enabling global profiling of substrates of PTM-enzymes and validation of PTM-sites in a straightforward manner.
Project description:A precise understanding of DNA methylation dynamics on a genome wide scale is of great importance for the comprehensive investigation of a variety of biological processes such as reprogramming of somatic cells to iPSCs, cell differentiation and also cancer development. To date, a complex integration of multiple and distinct genome wide data sets is required to derive the global activity of DNA modifying enzymes. We present GwEEP - Genome-wide Epigenetic Efficiency Profiling as a versatile approach to infer dynamic efficiency changes of DNA modifying enzymes at base pair resolution on a genome wide scale. GwEEP relies on genome wide oxidative Hairpin Bisulfite sequencing (HPoxBS) data sets, which are translated by a sophisticated hidden Markov model into quantitative enzyme efficiencies with reported confidence around the estimates. GwEEP in its present form predicts de novo and maintenance methylation efficiencies of Dnmts, as well as the hydroxylation efficiency of Tets but its purposefully flexible design allows to capture further oxidation processes such as formylation and carboxylation given available data in the future. Applied to a well characterized ES cell model, GwEEP precisely predicts the complex epigenetic changes following a Serum-to-2i shift i.e., (i) instant reduction in maintenance efficiency (ii) gradually decreasing de novo methylation efficiency and (iii) increasing Tet efficiencies. In addition, a complementary analysis of Tet triple knock-out ES cells confirms the previous hypothesized mutual interference of Dnmts and Tets. GwEEP is applicable to a wide range of biological samples including cell lines, but also tissues and primary cell types.
Project description:A hyperthermophilic archaeon Thermococcus kodakarensis can grow in a wide range of temperatures from 60°C to 93°C, optimally at 85°C. To screen the genes that being specifically expressed at lowest growth temperature, 60°C, we investigated the transcriptional profilings of T. kodakarensis grown at 60˚C by comparing with those grown at 85°C.
Project description:Heterotypic ubiquitin (Ub) chains have emerged as fundamental components in a wide range of cellular processes. The integrative identification of Ub-interacting proteins (readers) and Ub-modifying enzymes (writers and erasers) that selectively recognize and regulate heterotypic ubiquitination may provide crucial insights into these processes. In this study, we employed the bifunctional molecule-assisted (CAET) strategy to develop a new type of disulfide-bond-activated heterotypic Ub reagents, which allowed to enrich heterotypic Ub-interacting proteins and modifying enzymes simultaneously. The sequential release of readers which are non-covalently bound and writers or erasers which are covalently conjugated by using urea and reductant, respectively, combined with label-free quantitative (LFQ) MS indicated that these innovative heterotypic Ub tools would facilitate future investigations into functional roles played by heterotypic Ub chains.
Project description:This SuperSeries is composed of the following subset Series: GSE28549: Anaerobic Oxidation of Benzene by the Hyperthermophilic Archaeon Ferroglobus placidus (Phenol vs. Benzoate) GSE30798: Anaerobic Oxidation of Benzene by the Hyperthermophilic Archaeon Ferroglobus placidus (Benzene vs. Acetate) GSE30799: Anaerobic Oxidation of Benzene by the Hyperthermophilic Archaeon Ferroglobus placidus (Benzene vs. Phenol) GSE30801: Anaerobic Oxidation of Benzene by the Hyperthermophilic Archaeon Ferroglobus placidus (Benzene vs. Benzoate) Refer to individual Series