Dataset Information

Unravelling the proteomic landscape of extracellular vesicles in prostate cancer by density-based fractionation of urine

ABSTRACT: Mass spectrometry-based proteomic analysis of urinary EV (uEV) in men with benign and malignant prostate disease, profiling the proteome of EV separated from prostate tumor interstitial fluid and matched uEV, and a comparative proteomic analysis with uEV from patients with bladder and renal cancer.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Urine, Extracellular Vesicle

DISEASE(S): Prostate Cancer,Urinary Bladder Cancer,Kidney Cancer

SUBMITTER: Pekka Rappu

LAB HEAD: Olivier de Wever

PROVIDER: PXD015289 | Pride | 2020-04-09

REPOSITORIES: Pride

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Preparation of Multi-omics Grade Extracellular Vesicles by Density-Based Fractionation of Urine.

Dhondt Bert B Lumen Nicolaas N De Wever Olivier O Hendrix An A

STAR protocols 20200727 2

The multidimensional cargo of extracellular vesicles (EV) released in urine is a reflection of the pathophysiological processes occurring within their cells and tissues of origin in the urogenital system. Here, we describe a step-by-step protocol for density-based separation of urinary EV with high specificity and repeatability. The implementation of integrative omics allows the study of the molecular complexity of highly purified urinary EV, supporting the identification of EV-specific function ...[more]

PMID: 33111109

Publication: 1/2

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Project description:Extracellular vesicles (EVs) are membranous particles released by different organisms. EVs carry several kinds of macromolecules implicated in cell communication. EVs have become an important topic in the study of pathogenic fungi, due to their relation with the fungal-host interactions. One of the essential research areas in this field is the characterization protein profile of EV cargo, because of plant fungal pathogens rely heavily on secreted proteins to invade their hosts. However, little is known about EVs of Botrytis cinerea, which is one of the most devastating phytopathogenic fungi. The present study has two main objectives: the first isolation and characterization of B. cinerea EVs; and the description of their potential role during the infective process. All the experimental process was conducted in B. cinerea growing in mimimal salt medium supplemented with glucose as a constitutive stage, and deproteinized tomato cell walls (TCW) as virulence inductor. The isolation of EV was performed by differential centrifugation, filtration, ultrafiltration, and sucrose cushion ultracentrifugation. EV fractions were visualised by TEM using negative staining. Proteomic analysis of EV cargo was addressed by LC-MS/MS. The used methodology had allowed, for the first time, the correct isolation of B. cinerea EVs, and the identification of a high number of EV proteins, including potential EV markers. The isolated EVs displayed differences in the morphology under both assayed conditions. GO analysis of EV proteins showed an enrichment in cell wall metabolism and proteolysis under TCW. KEGG analysis also showed difference in EV function under both conditions, highlighting the presence of potential virulence/pathogenic factors implicated in cell wall metabolism, between others. This is the first evidence and characterization of B. cinerea EVs production, showing that EVs are essential in the infection process of B. cinerea and share crucial functions with the conventional secretion pathway.

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