Dataset Information

A thiol probe for measuring unfolded protein load and proteostasis in cells

ABSTRACT: When proteostasis becomes unbalanced, unfolded proteins accumulate and can aggregate. However, probes quantifying proteostasis imbalance are lacking. We report a dye, tetraphenylethene maleimide (TPE-MI) that can measure unfolded protein load.TPE-MI fluorescence is enhanced upon reaction with cellular proteomes under conditions promoting accumulation of unfolded proteins and, via proteomic analysis, can track which proteins expose more cysteine residues under stress.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Cell Culture

SUBMITTER: Nagaraj Moily

LAB HEAD: Danny Hatters

PROVIDER: PXD006527 | Pride | 2018-10-24

REPOSITORIES: Pride

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Publications

A thiol probe for measuring unfolded protein load and proteostasis in cells.

Chen Moore Z MZ Moily Nagaraj S NS Bridgford Jessica L JL Wood Rebecca J RJ Radwan Mona M Smith Trevor A TA Song Zhegang Z Tang Ben Zhong BZ Tilley Leann L Xu Xiaohong X Reid Gavin E GE Pouladi Mahmoud A MA Hong Yuning Y Hatters Danny M DM

Nature communications 20170907 1

When proteostasis becomes unbalanced, unfolded proteins can accumulate and aggregate. Here we report that the dye, tetraphenylethene maleimide (TPE-MI) can be used to measure cellular unfolded protein load. TPE-MI fluorescence is activated upon labelling free cysteine thiols, normally buried in the core of globular proteins that are exposed upon unfolding. Crucially TPE-MI does not become fluorescent when conjugated to soluble glutathione. We find that TPE-MI fluorescence is enhanced upon reacti ...[more]

PMID: 28883394

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Project description:Myocardial infarction (MI), an undesirable clinical outcome of coronary artery disease (CAD), triggers a potent inflammatory response via the release of circulatory mediators, including extracellular vesicles (EVs) by damaged cardiac cells, which is necessary for myocardial healing. However, when in excess, causes pathological tissue remodeling and eventual heart failure. Timely repression of MI-induced inflammatory response are critical to prevent and minimize cardiac tissue injuries, nonetheless, progression in this aspect remains clinically challenging. The well documentation on the ability of EVs to trigger a functional response with the delivery of bioactive cargos carried within, have made them clinically attractive as diagnostic biomarkers and drug vectors for therapeutic interventions. Using label-free quantitative proteomics approach, we compared the protein cargo of plasma EVs between patients with (MI) and from control patients with stable angina (NMI). We report, for the first time, the expression proteomics profiling on 252 plasma EV proteins that were modulated with >1.2-fold in myocardial injury. We identified a panel of six strongly up-regulated biomarkers with significant potential for clinical applications; these reflected post-infarct pathways of complement activation (Complement C1q subcomponent subunit A [C1QA], ~3.23-fold change, p = 0.012; Complement C5 [C5], ~1.27-fold change, p = 0.087), lipoprotein metabolism (Apoliporotein D [APOD], ~1.86-fold change, p = 0.033; Apolipoprotein C-III [APOCC3], ~2.63-fold change, p = 0.029) and platelet activation (Platelet glycoprotein Ib alpha chain [GP1BA], ~9.18-fold change, p < 0.0001; Platelet basic protein [PPBP], ~4.72-fold change, p = 0.027). The data have been deposited to the ProteomeXchange with identifier PXD002950. This novel biomarker panel was successfully validated in a separate cohort of 43 individual angina patients using Luminex analysis of the representative EV proteins C1QA (p = 0.005) and C5 (p = 0.0021), which act as critical regulators of complement activity in MI. We further present that all EV-derived fibrinogen components detected were paradoxically down-regulated in MI, suggesting that a compensatory mechanism may suppress post-infarct coagulation pathways, and indicating potential for therapeutic targeting of this mechanism in MI. Taken together, these data urge the further development of novel EV-based diagnostic and therapeutic strategies to benefit patients with CAD.

Dataset Information

A thiol probe for measuring unfolded protein load and proteostasis in cells

Publications

A thiol probe for measuring unfolded protein load and proteostasis in cells.

Similar Datasets

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