Showing papers by "Dorothea Fiedler published in 2015"
TL;DR: In this paper, a large-scale genetic interaction analysis in Saccharomyces cerevisiae involving approximately 49 × 1,200 double mutants in the presence of five different stress conditions, including osmotic, oxidative and cell wall-altering stresses.
Abstract: Genetic interaction screens have been applied with great success in several organisms to study gene function and the genetic architecture of the cell. However, most studies have been performed under optimal growth conditions even though many functional interactions are known to occur under specific cellular conditions. In this study, we have performed a large-scale genetic interaction analysis in Saccharomyces cerevisiae involving approximately 49 × 1,200 double mutants in the presence of five different stress conditions, including osmotic, oxidative and cell wall-altering stresses. This resulted in the generation of a differential E-MAP (or dE-MAP) comprising over 250,000 measurements of conditional interactions. We found an extensive number of conditional genetic interactions that recapitulate known stress-specific functional associations. Furthermore, we have also uncovered previously unrecognized roles involving the phosphatase regulator Bud14, the histone methylation complex COMPASS and membrane trafficking complexes in modulating the cell wall integrity pathway. Finally, the osmotic stress differential genetic interactions showed enrichment for genes coding for proteins with conditional changes in phosphorylation but not for genes with conditional changes in gene expression. This suggests that conditional genetic interactions are a powerful tool to dissect the functional importance of the different response mechanisms of the cell.
45 citations
01 Apr 2015
TL;DR: The large‐scale genetic interaction analysis in Saccharomyces cerevisiae involving approximately 49 × 1,200 double mutants in the presence of five different stress conditions resulted in the generation of a differential E‐MAP (or dE‐MAP), which suggests that conditional genetic interactions are a powerful tool to dissect the functional importance of the different response mechanisms of the cell.
Abstract: © 2015 The Authors.Abstract Genetic interaction screens have been applied with great success in several organisms to study gene function and the genetic architecture of the cell. However, most studies have been performed under optimal growth conditions eve
42 citations
TL;DR: A resin-bound dinuclear zinc(II) complex for the selective capture of pyrophosphopeptide in the presence of cell lysate is reported, and the high affinity and specificity of the reagent makes it an attractive tool for the study of in vivo pyroph phosphorylation.
Abstract: A resin-bound dinuclear zinc(II) complex for the selective capture of pyrophosphopeptides is reported. The metal complex binds diphosphate esters over other anionic groups, such as monophosphate esters, sulfate esters, and carboxylic acids, with high specificity. Immobilization of the compound provided a reagent capable of binding and retaining nanomolar quantities of pyrophosphopeptide in the presence of cell lysate. The high affinity and specificity of the reagent makes it an attractive tool for the study of in vivo pyrophosphorylation.
28 citations
TL;DR: The design and synthesis of a fluorescent sensor of diphosphate esters along with its application for in-gel detection and competent stain for visualization of pyrophosphorylated proteins in polyacrylamide gels are described.
Abstract: The design and synthesis of a fluorescent sensor of diphosphate esters along with its application for in-gel detection is described. Dinuclear zinc complex 1 selectively binds diphosphate esters in the presence of various other functional groups, including monophosphate esters. Complex 1 also constitutes a competent stain for visualization of pyrophosphorylated proteins in polyacrylamide gels. This reagent will facilitate the validation and exploration of protein pyrophosphorylation.
23 citations
TL;DR: The findings provide evidence for the reversibility of pyrophosphorylation and thereby highlight the potential impact of this modification on cellular signal transduction pathways.
Abstract: Protein pyrophosphorylation is emerging as a new post-translational modification, yet its role in cellular signaling remains poorly characterized. Important factors in determining the biological relevance of pyrophosphorylation include understanding the chemical and biochemical stability of the pyrophosphoryl group and elucidating the reversibility of modification in a cellular context. Towards this end, we prepared a series of synthetic pyrophosphopeptides, which were utilized to demonstrate that the modification is quite inert over a wide pH range but can be removed biochemically by alkaline phosphatases. Importantly, we observed enzyme-dependent removal of the pyrophosphate in mammalian and yeast cell lysates using the synthetic pyrophosphopeptides. The findings provide evidence for the reversibility of pyrophosphorylation and thereby highlight the potential impact of this modification on cellular signal transduction pathways.
20 citations
TL;DR: In this article, a robust method to incorporate pyrophosphate residues into peptide sequences was developed to explore the signaling role of protein pyrophophosphorylation, a recently discovered modification mediated by the inositol pyrophosate messengers.
Abstract: The detection and annotation of posttranslationally modified proteins in complex samples has benefited greatly from the ability to chemically synthesize and characterize model compounds harboring the modification of interest. To explore the signaling role of protein pyrophosphorylation, a recently discovered modification mediated by the inositol pyrophosphate messengers, our group has developed a robust method to incorporate pyrophosphate residues into peptide sequences. We highlight several examples from the literature that inspired our chemical approach and discuss future applications of our orthogonal pyrophosphorylation strategy.