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HEK293 cells LC-MSMS - XBP1 mitigates aminoglycoside-induced endoplasmic reticulum stress and neuronal cell death

ABSTRACT: The iTRAQ 4-plex experiment was carried out with 2 biological replicates of untreated HEK cells (114 and 116 label) and 2 biological replicates of HEK cells treated with geneticin (16 µM) for 32 h (115 and 117 label).

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Stefan Duscha

LAB HEAD: Prof. Dr. Erik Christian Böttger

PROVIDER: PXD000933 | Pride | 2015-08-10

REPOSITORIES: Pride

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XBP1 mitigates aminoglycoside-induced endoplasmic reticulum stress and neuronal cell death.

Oishi N N Duscha S S Boukari H H Meyer M M Xie J J Wei G G Schrepfer T T Roschitzki B B Boettger E C EC Schacht J J

Cell death & disease 20150514

Here we study links between aminoglycoside-induced mistranslation, protein misfolding and neuropathy. We demonstrate that aminoglycosides induce misreading in mammalian cells and assess endoplasmic reticulum (ER) stress and unfolded protein response (UPR) pathways. Genome-wide transcriptome and proteome analyses revealed upregulation of genes related to protein folding and degradation. Quantitative PCR confirmed induction of UPR markers including C/EBP homologous protein, glucose-regulated prote ...[more]

PMID: 25973683

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Analysis of the Escherichia coli membrane vesicle proteome identifies markers of purity and culture conditions

Project description:Bacteria release nano-sized membrane vesicles (MVs) into the extracellular milieu. Bacterial MVs contain molecular cargo originating from the parent bacterium and have important roles in bacterial survival and pathogenesis. Using 8-plex iTRAQ approaches, we profiled the MV proteome of two Escherichia coli strains - uropathogenic E. coli (UPEC) 536 and probiotic Nissle 1917. For these strains, we compared the difference in the MV proteome between a crude input MV prepared by ultracentrifugation alone with that from MVs that were further purified by either density gradient centrifugation (DGC) or size exclusion chromatography (SEC); and also compared MVs from bacterial cultures that were grown in iron-restricted (R) and ironsupplemented (RF) conditions. We found that overall, outer membrane components were highly enriched, and bacterial inner membrane components were significantly depleted in both UPEC and Nissle MVs, in keeping with an outer membrane origin. In addition, we found enrichment of ribosome-related Gene Ontology terms in UPEC MV, and proteins involved in glycolytic process and ligase activity in Nissle MV. We have identified that three proteins (RbsB of UPEC in R; YoeA of UPEC in RF; BamA of Nissle in R) were consistently enriched in the DGC- and SEC-purified MV samples in comparison to their crude input MV, whereas conversely the 60 kDa chaperonin GroEL was enriched in the crude input MVs for both UPEC and Nissle in R condition. Such proteins may have a potential utility as technical markers for assessing the purity from contaminating non-vesicular protein after MV purification. Several proteins were changed in their abundance depending on the iron availability in the media.

2019-07-15 | PXD011345 | Pride

The RNA-binding protein RBP42 regulates cellular energy metabolism in mammalian-infective Trypanosoma brucei

Project description:RNA-binding proteins are key players in coordinated post-transcriptional regulation of functionally related genes, defined as RNA regulons. RNA regulons play particularly critical roles in parasitic trypanosomes, which exhibit unregulated co-transcription of long unrelated gene arrays. In this report, we present a systematic analysis of an essential RNA-binding protein, RBP42, in the mammalian-infective bloodstream form of African trypanosome, and show that RBP42 is a key regulator of parasite’s central carbon and energy metabolism. Using individual-nucleotide resolution UV cross-linking and immunoprecipitation (iCLIP) to identify genome-wide RBP42-RNA interactions, we show that RBP42 preferentially binds within the coding region of mRNAs encoding core metabolic enzymes. Global quantitative transcriptomic and proteomic analyses reveal that loss of RBP42 reduces the abundance of target mRNA-encoded proteins, but not target mRNA, suggesting a positive translational regulatory role of RBP42. Significant changes in central carbon metabolic intermediates, following loss of RBP42, further support its critical role in cellular energy metabolism.

2021-06-11 | PXD026622 | Pride

Tetraploid wheat (Triticum turgidum L.),LC-MSMS

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2018-10-19 | PXD008910 | Pride

Spaceflight adaptation requires organ specific alterations in the proteomes of Arabidopsis

Project description:This experiment was launched on STS-129 in 2009 and was supported by NASA grant NNX07AH27G - Transgenic Plant Biomonitors of Space Flight Exposure to R.J. Ferl and A-L. Paul.<br><br>Life in spaceflight demonstrates remarkable adaptive processes within the specialized environments of space vehicles which are subject to the myriad of attending and unique environmental issues associated with orbital trajectories. To examine the adaptive processes that occur in plants in space, leaves and roots from Arabidopsis seedlings that were grown from seed for 12 days on the International Space Station and preserved on orbit in RNAlater were returned to earth and analyzed using iTRAQ broad scale proteomics procedures.

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Project description:After long-term aluminum treatment, thereafter, we investigated Citrus root protein profiles using isobaric tags for relative and absolute quantification (iTRAQ).

2017-10-26 | PXD002301 | Pride

Identify Novel Iron–Transporting Proteins in Streptococcus pneumoniae

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2018-10-26 | PXD003313 | Pride

Differential regulation of tobacco host factors upon TMV infection

Project description:Tobacco mosaic virus (TMV) became one of the model virus for fundamental biological research and understanding of the host-pathogen interaction. It utilizes host machinery smartly to complete the life cycle, though many parts of life cycle remained a mystery, such as host factors involved in virus replication processes. We utilized the 8-Plex iTRAQ study to understand host-pathogen interaction between TMV and host tobacco.

2018-11-07 | PXD008837 | Pride

iTRAQ MS of mouse LbetaT2 cells transfected with Galphas siRNA vs. scrambled siRNA

Project description:Twenty million LbetaT2 cells were transfected with either control or Galphas siRNA, then were seeded in 100-mm cell culture plates in DMEM + 10% FBS. Two days later, cells were washed twice with pre-warmed PBS. Conditioned media was harvested another 24 h later, and centrifuged at 20,000 g for 10 min at 4°C to remove cell debris. To enrich secreted proteins in the conditioned media, conditioned media samples were centrifuged using Amicon centrifugal filters with a 3kDa cutoff (Millipore, Billerica, MA). A total of 8 concentrated conditioned media samples were independently prepared: 4 samples from control siRNA-treated cells, and 4 samples from Galphas siRNA-treated cells. Samples were stored at –70°C until they were sent to the Mount Sinai Proteomics Core Facility.HPLC-isobaric tags for relative and absolute quantitation mass-spectrometry (iTRAQ MS) - Data analysis ProteinPilot 3.0 (AB Sciex) was used to search the MS/MS spectra for protein identification and quantitation with its searching algorithm Paragon 3.0.0.0 (*Reference). The protein database used for searching was Uniprot mouse fasta file (release-2010_11). The search parameters include quantitation for iTRAQ 8-plex (peptide-labeled), MMTS for cysteine alkylation, trypsin for enzyme digestion, biological modifications for ID focus, and taxonomy set for Mus musculus. The detected protein threshold was set to 1.3 (95% confidence). Additionally, we converted our AB Sciex mass spectral data (TOF/TOF data) into an mzML format, using the AB Sciex MS Data Converter (beta version 1.3) tool. Finally, we used the command line tool group2xml, which is included with ProteinPilot Software, to convert the .group search engine result file to an XML file.

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Ctenopharyngodon idella iTRAQ - Understanding the Immune Mechanisms of Vibrio mimicus targeted epitopes gene vaccine by iTRAQ Technology

Project description:Understanding the Immune Mechanisms of Vibrio mimicus targeted epitopes gene vaccine by iTRAQ Technology

2018-12-18 | PXD012011 | Pride

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Project description:Cadmium is one of several heavy metals present in contaminated soils. Apparently, it has no biological role but can produce DNA damage, overexpression of stress response proteins and misfolded proteins, amongst other deleterial effects. Acidithiobacillus ferrooxidans is an acidophilic bacterium capable of resisting very high concentrations of heavy metals such as cadmium. This is important for industrial bioleaching processes where Cd+2 concentrations can be in the range of 5-100 mM. Cadmium resistance mechanisms in these microorganisms have not been fully characterized. A. ferrooxidans ATCC 53993 contains genes coding for possible metal resistance determinants such as efflux systems belonging to three families: P-type ATPases, RND transporters and cation diffusion facilitators (CDF). In addition, it has some extra copies of these genes in its exclusive genomic island (GI). Several of these putative genes were characterized in the present report by determining their transcriptional expression profiles and functionality. Moreover, a global quantitative proteomic analysis was carried out to further explore new cadmium resistance determinants in this biomining acidophile. Changes in iron oxidation pathways, upregulation of transport proteins (P-type ATPases and CDFs) and changes in ribosomal protein levels were seen. Finally, increased concentrations of exclusive putative cadmium ATPases present in strain ATCC 53993 GI and other non-identified proteins such as Lferr_0210, which forms part of a possible operon, could explain its greater resistance to cadmium compared to other acidophiles such as A. ferrooxidans ATCC 23270.

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