Browse
Submit Data
Databases
API
Help

Dataset Information

311 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Identifying RasGAP SH2 binding partners in Drosophila using mass spectrometry

ABSTRACT: Drosophila S2 cells were transfected with RasGAP(WT), RasGAP(SH2*32*) and GFP control. 4 mg of S2 cell lysate was used to pulldown LAP and GFP tagged constructs using GFP-Trap. The pulled down fraction was loaded on SDS-PAGE gels and coomassie stained before several bands (a-u) from two independent but parallel experiments were excised and subjected to MS analysis.

INSTRUMENT(S): LTQ Orbitrap, instrument model

ORGANISM(S): Drosophila Melanogaster (fruit Fly)

SUBMITTER: Behzad Rowshanravan

PROVIDER: PXD000176 | Pride | 2014-05-20

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Publications

RasGAP mediates neuronal survival in Drosophila through direct regulation of Rab5-dependent endocytosis.

Rowshanravan Behzad B Woodcock Simon A SA Botella José A JA Kiermayer Claudia C Schneuwly Stephan S Hughes David A DA

Journal of cell science 20140509 Pt 13

The GTPase Ras can either promote or inhibit cell survival. Inactivating mutations in Drosophila RasGAP (encoded by vap), a Ras GTPase-activating protein, lead to age-related brain degeneration. Genetic interactions implicate the epidermal growth factor receptor (EGFR)-Ras pathway in promoting neurodegeneration but the mechanism is not known. Here, we show that the Src homology 2 (SH2) domains of RasGAP are essential for its neuroprotective function. By using affinity purification and mass spect ...[more]

PMID: 24816559

Similar Datasets

N-terminomics Proteogenomics - N-terminal proteomics assisted profiling of the unexplored translation initiation landscape in Arabidopsis thaliana

Project description:Proteogenomics is a research field where proteome data is used to improve gene annotation. To achieve this, customized protein databases are constructed to match proteomic data. We perform a proteogenomic analysis using N-terminal COFRADIC data in order to identify novel translational initiation start sites. We use a multistage search strategy where spectra that remained unidentified after searching the Arabidopsis proteome are used for our proteogenomic analysis. Here, the unidentified spectra were searched against a customized N-terminal peptide library derived from a six-frame translation of the Arabidopsis (Arabidopsis thaliana) genome as well as Augustus predicted gene models.

2017-04-24 | PXD004896 | Pride

Structural analysis of the regulatory GAF domains of cGMP phosphodiesterase elucidates the allosteric communication pathway

Project description:Regulation of photoreceptor phosphodiesterase (PDE6) activity is responsible for the speed, sensitivity, and recovery of the photoresponse during visual signaling in vertebrate photoreceptor cells. It is hypothesized that the physiological differences in the light responsiveness of rods and cones may result in part from differences in the structure and regulation of the distinct isoforms of rod and cone PDE6. Although rod and cone PDE6 catalytic subunits share a similar domain organization consisting of tandem GAF domains (GAFa and GAFb) and a catalytic domain, cone PDE6 is a homodimer whereas rod PDE6 consists of two homologous catalytic subunits. Here we provide the x-ray crystal structure of cone GAFab regulatory domain solved at 3.3 Å resolution, in conjunction with chemical cross-linking and mass spectrometric analysis of conformational changes to GAFab induced upon binding of cGMP and the PDE6 inhibitory γ-subunit (Pγ). Ligand-induced changes in cross-linked residues implicate multiple conformational changes in the GAFa and GAFb domains in forming an allosteric communication network that communicates with the PDE6 catalytic domains. Molecular dynamics (MD) simulations of cone GAFab revealed asymmetry in the two GAFab subunits forming the homodimer and allosteric perturbations on cGMP binding. Cross-linking of Pγ to GAFab in conjunction with solution NMR spectroscopy of isotopically labeled Pγ identified the central polycationic region of Pγ interacting with the GAFb domain. These results provide a mechanistic basis for developing allosteric activators of PDE6 with therapeutic implications for halting the progression of several retinal degenerative diseases.

2022-02-15 | PXD020817 | Pride

Identification of O-glycosylation sites in Clostridium botulinum and Geobacillus kaustophilus flagellins

Project description:In this study, we identified the O-glycosylation sites in recombinant Clostridium botulinum flagellin (CbFla) and Geobacillus kaustophilus flagellins, GkFlaA1 and GkFlaA2, expressed from the E. coli strains, EV136 and EV240 (K1:K12 strains engineered by Prof. Eric Vimr’s group to accumulate CMP-sialic acid in the cytosol), EV36 (wildtype K1:K12 strain), and BL21(DE3) (laboratory strain used for protein expression using T7 RNA polymerase). The recombinant flagellins were expressed with or without co-expression of motility associated factor (Maf) (flagellin nonulosonic acid glycosyltransferase) in these strains, purified and subjected to MS/MS analysis at Taplin Biological Mass Spectrometry facility.

2020-09-07 | PXD018167 | Pride

Venom gland transcriptome and venom proteome of the Tibellus oblongus spider unveil molecular diversity of polypeptide toxins

Project description:The Tibellus genus spider is an active hunter that does not spin webs and remains highly underinvestigated in terms of the venom composition. Here, we present a combination of venom glands transcriptome cDNA analysis, venom proteome analysis for unveiling of the Tibellus genus spider venom composition.

2023-10-23 | PXD040219 | Pride

Human neuronal extracellular vesicles contain a transcriptional network and tetraspanin-defined subpopulations

Project description:Healthy brain function is mediated by several complementary signalling pathways, many of which are driven by extracellular vesicles (EVs). EVs are heterogeneous in both size and cargo and are constitutively released from cells into the extracellular milieu. They are subsequently trafficked to recipient cells, whereupon their entry can modify the cellular phenotype. Here, in order to further understand the functional role of EVs in neurons, we isolated EVs by size exclusion chromatography from human induced pluripotent stem cell (iPSC)-derived neurons. Electron microscopy and dynamic light scattering revealed that the isolated EVs had a diameter of 30-100 nm. Transcriptomic and proteomics analyses of the EVs and neurons identified key molecules enriched in the EVs involved in cell surface interaction (integrins and collagens), internalisation pathways (clathrin- and caveolin-dependent), downstream signalling pathways (phospholipases, integrin-linked kinase and MAPKs), and long-term impacts on cellular development and maintenance. Overall, we show that key signalling networks and mechanisms are enriched in EVs isolated from human iPSC-derived neurons, and identify subpopulations of EVs defined by differential tetraspanin expression.

2021-09-08 | PXD015255 | Pride

A probiotics strain attenuating the effects of aspirin in the gut

Project description:Transcriptome analysis of partially degraded and fragmented RNA samples from mus musculus gut Global gene expression profiling has shown the gut transcripts changes through administration of aspirin while probiotics strain administration beforehand attenuates the effect more than teprenone. We analyzed five groups of mice gut using the Affymetrix Mouse Gene 1.0 ST platform. Array data was processed by Affymetrix Exon Array Computational Tool. No techinical replicates were performed.

2014-01-07 | E-GEOD-53851 | biostudies-arrayexpress

Endogenous, in vivo, and site-specific characterization of the SUMO architecture across species and organs

Project description:Small ubiquitin-like modifiers (SUMOs) are post-translational modifications that play crucial roles in most cellular processes. While methods exist to study exogenous SUMOylation, large-scale characterization of endogenous SUMO has remained technically daunting. Here, we describe a proteomics approach facilitating system-wide and in vivo identification of lysines modified by endogenous and native SUMO2/3. We identified 14,869 endogenous SUMO sites in human cells during heat stress and proteasomal inhibition, and mapped 1,963 SUMO sites across eight mouse tissues; brain, heart, kidney, lung, liver, muscle, spleen, and testis. Quantification of the SUMO equilibrium highlighted striking differences in SUMO metabolism, between cells and tissues. Targeting preferences of SUMO varied across different organ types, coinciding with markedly differential SUMOylation states of all enzymes involved in the SUMO conjugation cascade. Collectively, our systemic investigation details the SUMOylation architecture across species and organs and provides a resource of endogenous SUMOylation sites on factors important in organ-specific functions and disease.

2018-06-26 | PXD008003 | Pride

Talin1-GFP pulldown in U2OS cells

Project description:In this study, we pull-down human Talin1-GFP and GFP from U2OS cells plated on fibronectin. Binding partners were then analysed using mass-spectrometry.

2021-05-12 | PXD024634 | Pride

An Interaction between RRP6 and SU(VAR)3-9 Targets RRP6 to Heterochromatin and Contributes to Heterochromatin Maintenance in Drosophila melanogaster [RNA-seq]

Project description:We have investigated the effect of RRP6 depletion on the transcriptome of S2 cells using Illumina deep RNA sequencing. We have also carried out Illumina ChIP-seq analysis of RRP6 genome occupancy in control S2 cells (GFP-KD) and in cells depleted of SU(VAR)3-9.

2015-08-20 | GSE70625 | GEO

Cupiennius salei Hyal LC-MSMS - Isolation, N-glycosylations and Function of a Hyaluronidase-Like Enzyme from the Venom of the Spider Cupiennius salei

Project description:Hyaluronidases are important venom components by acting as spreading factor of neurotoxins. In several studies this spreading effect was tested on vertebrate tissue. However, data about the spreading activity on invertebrates, which are the main prey organisms of spiders, are lacking. In this study, a hyaluronidase-like enzyme was isolated from the venom of the spider Cupiennius salei. The amino acid sequence of the enzyme was solved by cDNA analysis of the venom gland transcriptome and confirmed by protein analytics. Two complex N-linked glycans, which akin honey bee hyaluronidase glycosylations, were identified by tandem mass spectrometry.

2015-12-10 | PXD002489 | 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