Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The regulatory landscape of the yeast phosphoproteome – ultra-deep reference phosphoproteomic DDA data

ABSTRACT: We systematically exposed the yeast Saccharomyces cerevisiae to a wide array of 101 perturbations and generated a quantitative mass spectrometry-based atlas of early phosphoproteomic responses. Perturbations covered broad classes of environmental, chemical, and drug perturbations and systematically targeted diverse aspects of yeast cell biology. Importantly, most perturbations focused on capturing early signaling responses (5 min treatments). An ultra-deep yeast phosphoproteome was generated from a pooled sample of all 5 min perturbations by multi protease digestion, deep offline fractionations using two different peptide separation methods, and data-dependent acquisition (DDA). This submission includes 120 DDA-MS raw files that were used to generate a deep yeast phosphoproteome.

INSTRUMENT(S): Orbitrap Eclipse, Orbitrap Exploris 480, Q Exactive

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: Mario Leutert

LAB HEAD: Judit Villén

PROVIDER: PXD035029 | Pride | 2023-08-27

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Similar Datasets

R2-P2 rapid-robotic phosphoproteomics enables multidimensional cell signaling studies

Project description:Recent developments in mass spectrometry-based proteomics have enabled systems-level studies of cellular signaling, where >10,000 phosphosites can be routinely identified and quantified. Yet, current analyses are limited in throughput, reproducibility, and robustness, hampering experiments that involve multiple perturbations, such as those needed to map kinase-substrate relationships, capture pathway crosstalks, and network inference analysis. To address these challenges and fully exploit the potential of phosphoproteomics, we introduce rapid-robotic-phosphoproteomics (R2-P2), an end-to-end automated method that processes samples in a 96-well format, from a protein extract to mass spectrometry-ready phosphopeptides. R2-P2 uses magnetic particles for both protein sample cleanup and phosphopeptide enrichment. R2-P2 is more flexible, high-throughput, rapid, and robust than classical protocols. To showcase the method, we have applied it, in combination with data-independent acquisition mass spectrometry, to study signaling dynamics in the mitogen-activated protein kinase (MAPK) pathway in the yeast model Saccharomyces cerevisiae. Our results reveal broad and specific signaling events along the mating, the high-osmolarity glycerol, and the invasive growth branches of the MAPK pathway, with robust phosphorylation of downstream regulatory proteins and transcription factors. Our method greatly facilitates large-scale signaling studies involving hundreds of perturbations and will open the door to systems-level studies aiming to capture the complexity of signaling.

2019-12-12 | PXD013453 | Pride

Automated Enrichment of Phosphotyrosine Peptides for High-Throughput Proteomics

Project description:Phosphotyrosine (pY) enrichment is critical for expanding fundamental and clinical understanding of cellular signaling by mass spectrometry-based proteomics. However, current pY enrichment methods exhibit a high cost per sample and limited reproducibility due to expensive affinity reagents and manual processing. We present rapid-robotic phosphotyrosine proteomics (R2-pY), which uses a magnetic particle processor and pY superbinders or antibodies. R2-pY can handle up to 96 samples in parallel, requires 2 days to go from cell lysate to mass spectrometry injections, and results in global proteomic, phosphoproteomic and tyrosine-specific phosphoproteomic samples. We benchmark the method on HeLa cells stimulated with pervanadate and serum and report over 4000 unique pY sites from 1 mg of peptide input, strong reproducibility between replicates, and phosphopeptide enrichment efficiencies above 99%. R2-pY extends our previously reported R2-P2 proteomic and global phosphoproteomic sample preparation framework, opening the door to large-scale studies of pY signaling in concert with global proteome and phosphoproteome profiling.

2023-05-05 | PXD041271 | Pride

The regulatory landscape of the yeast phosphoproteome – quantitative proteomics DIA data

Project description:We systematically exposed the yeast Saccharomyces cerevisiae to a wide array of 101 perturbations and generated a quantitative mass spectrometry-based atlas of early phosphoproteomic responses. Perturbations covered broad classes of environmental, chemical, and drug perturbations and systematically targeted diverse aspects of yeast cell biology. Importantly, most perturbations focused on capturing early signaling responses (5 min treatments), with the reasoning that functional phosphorylation of specific substrates occurs rapidly following stimulation, but promiscuous phosphorylation and confounding changes in transcript and protein abundances takes place more slowly. Large-scale proteomics and phosphoproteomics was enabled by our recently developed R2-P2 (Rapid-Robotic Phosphoproteomics) sample preparation workflow in combination with DIA measurements. In parallel to phosphoproteomic measurements we performed total proteome measurements on the same samples for 30 selected perturbations. This submission includes 126 quantitative DIA-MS raw files that were used for the total proteome analysis.

2023-08-27 | PXD034997 | Pride

The regulatory landscape of the yeast phosphoproteome – quantitative phosphoproteomic DIA data

Project description:We systematically exposed the yeast Saccharomyces cerevisiae to a wide array of 101 perturbations and generated a quantitative mass spectrometry-based atlas of early phosphoproteomic responses. Perturbations covered broad classes of environmental, chemical, and drug perturbations and systematically targeted diverse aspects of yeast cell biology. Importantly, most perturbations focused on capturing early signaling responses (5 min treatments), with the reasoning that functional phosphorylation of specific substrates occurs rapidly following stimulation, but promiscuous phosphorylation and confounding changes in transcript and protein abundances takes place more slowly. Treatments were done in 6 biological replicates and large-scale phosphoproteomics was enabled by our recently developed R2-P2 (Rapid-Robotic Phosphoproteomics) sample preparation workflow in combination with DIA measurements. This submission includes 559 quantitative phospho DIA-MS raw files that went into the analysis of the systems-level phosphorylation stress response.

2023-08-27 | PXD035050 | Pride

Investigation of Proteomic and Phosphoproteomic Responses to Signaling Network Perturbations Reveals Functional Pathway Organizations in Yeast

Project description:Governance of protein phosphorylation by kinases and phosphatases constitutes an essential regulatory network in eukaryotic cells. Network dysregulation leads to severe consequences and is often a key factor in disease pathogenesis. Previous studies revealed multiple roles for protein phosphorylation and pathway structures in cellular functions from different perspectives. We sought to understand the roles of kinases and phosphatases from a protein homeostasis point of view. Using a streamlined tandem mass tag (SL-TMT) strategy, we systematically measured proteomic and phosphoproteomic responses to perturbations of phosphorylation signaling networks in yeast deletion strains. Ours results emphasize the requirement for protein normalization for more complete interpretation of phosphorylation data. Functional relationships between kinases and phosphatases were characterized at both proteome and phosphoproteome levels in three ways: 1) gene ontology enrichment analysis, 2) ∆gene-∆gene correlation networks and 3) molecule covariance networks. This resource illuminates kinase and phosphatase functions and pathway organizations.

2019-11-15 | PXD015575 | Pride

Cdk1 controls global epigenetic landscape in embryonic stem cells

Project description:The cyclin-dependent kinase 1 (Cdk1) represents an ancient cell cycle kinase that has been conserved from yeast to humans. Cdk1 is the only essential mammalian Cdk, which drives cytokinesis by phosphorylating a large number of cellular proteins. To uncover additional functions of Cdk1, we generated a knock-in strain of mice expressing an analog-sensitive version of Cdk1 in place of wild-type Cdk1. These mice and cells derived from them allow us to investigate Cdk1 function in essentially any compartment and at any stage of development. In our study, we focused on embryonic stem (ES) cells, as this cell type expresses particularly high levels of Cdk1. Very unexpectedly, we found that in ES cells the majority of Cdk1 substrates are localized on chromatin. Cdk1 phosphorylates a large number of proteins involved in epigenetic regulation, including writers and erasers of all major histone marks. Consistent with this finding, inhibition of Cdk1 altered histone modification status of ES cells. High levels of Cdk1 in ES cells (and in induced pluripotent stem cells) phosphorylate and partially inactivate Dot1l, the histone H3 lysine 79 methyltransferase responsible for placing activating H3K79 marks on gene bodies. Decrease of Cdk1 activity during ES cell differentiation de-represses Dot1l, thereby allowing coordinated expression of differentiation genes. These analyses indicate that Cdk1 functions to maintain the epigenetic identity of ES cells.

2020-10-05 | PXD015173 | Pride

Histone deacetylation at coding sequences adjusts transcription kinetics during Candida albicans morphogenesis [ChIP-seq]

Project description:The dataset contains ChIP-Seq data of the Set3 and Hos2 proteins in Candida albicans, assayed in two morphological phases (yeast and hypha). The Set3 and Hos2 proteins in the respective strains carry 9myc epitopes and ChIP was performed with an anti-myc antibody. Included samples are the following: 1 input and 1 ChIP sample of an untagged wild type strain as negative control assayed in the yeast phase, 1 input and 3 ChIP biological replicates of the Set3-9myc strain in the yeast phase, 1 input and 2 ChIP biological replicates of the Set3-9myc strain in the hypha phase, 1 input and 2 ChIP biological replicates of the Hos2-9myc strain in the yeast phase, 1 input and 2 ChIP biological replicates of the Hos2-9myc strain in the hypha phase, 1 input and 3 ChIP biological replicates of Set3-9myc in a set1delta/delta background in the yeast phase. ChIP-Seq was performed of Candida albicans strains in two morphological phases (yeast and hypha). Yeast-phase cells were grown to the exponential phase in YPD at 30C. Hyphal differentiation was induced by resuspending the cells in YPD+20% Fetal Calf Serum and a shift of the growth temperature to 37C. Induction was performed for 30 minutes. Cells were crosslinked with 1% formaldehyde for 15 minutes at room temperature.

2013-01-01 | E-GEOD-38425 | biostudies-arrayexpress

The Inhibitory Receptor Siglec-8 Interacts with FcεRI and Globally Inhibits Intracellular Signaling in Primary Mast Cells Upon Activation

Project description:Immunomodulation of mast cell (MC) activity is warranted in allergic and inflammatory diseases where MCs have a central role in pathogenesis. Targeting Siglec-8, an inhibitory receptor on MCs and eosinophils, has shown promising activity in preclinical and clinical studies. While the intracellular pathways that regulate Siglec-8 activity in eosinophils have been well studied, the signaling mechanisms that lead to MC inhibition have not been fully elucidated. Here, we evaluate the intracellular signaling pathways of Siglec‐8‐mediated inhibition in primary MCs using an anti‐Siglec‐8 monoclonal antibody (mAb). Phospho‐proteomic profiling of FcεRI‐activated MCs revealed Siglec-8 mAb-treatment globally inhibited proximal and downstream kinases, leading to attenuated MC activation and degranulation. In fact, Siglec‐8 was found to directly interact with FcεRI signaling molecules. Siglec‐8 inhibition was dependent on both cytoplasmic immunoreceptor tyrosine-based inhibitory motifs (ITIMs) that interact with the SH2 containing protein phosphatase Shp-2 upon Siglec-8 phosphorylation. Taken together, these data support a model in which Siglec-8 regulates proximal FcεRI‐induced phosphorylation events through phosphatase recruitment and interaction with FcεRI, resulting in global inhibition of MCs upon Siglec-8 mAb engagement.

2022-01-21 | PXD030820 | Pride

Clontech RNA samples

Project description:A series of experiments used to compare Clontech spleen plyA, Clontech liver plyA and Clontech universal RNA for microarrays on our available human array printed on Amersham type 7 * slides. Keywords = human, clontech, automated slide processor, spleen, liver, universal RNA

2010-05-27 | E-GEOD-40 | biostudies-arrayexpress

Phosphoproteome of Hepatitis C Virus infected Huh7.5.1 cells

Project description:This project enriched and identified phosphoproteins in human hepatocarcinoma 7.5.1 cell line (Huh7.5.1) upon Hepatitis C virus (HCV) infection.

2016-07-08 | PXD000988 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data