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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV100062	1/1	Saccharomyces cerevisiae	Yeast	(d)(U)C Filtration UF	Oliveira DL	2010	29%
Study summary Full title Characterization of yeast extracellular vesicles: evidence for the participation of different pathways of cellular traffic in vesicle biogenesis All authors Oliveira DL, Nakayasu ES, Joffe LS, Guimarães AJ, Sobreira TJ, Nosanchuk JD, Cordero RJ, Frases S, Casadevall A, Almeida IC, Nimrichter L, Rodrigues ML Journal PLoS One Abstract BACKGROUND: Extracellular vesicles in yeast cells are involved in the molecular traffic across the c (show more...)BACKGROUND: Extracellular vesicles in yeast cells are involved in the molecular traffic across the cell wall. In yeast pathogens, these vesicles have been implicated in the transport of proteins, lipids, polysaccharide and pigments to the extracellular space. Cellular pathways required for the biogenesis of yeast extracellular vesicles are largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: We characterized extracellular vesicle production in wild type (WT) and mutant strains of the model yeast Saccharomyces cerevisiae using transmission electron microscopy in combination with light scattering analysis, lipid extraction and proteomics. WT cells and mutants with defective expression of Sec4p, a secretory vesicle-associated Rab GTPase essential for Golgi-derived exocytosis, or Snf7p, which is involved in multivesicular body (MVB) formation, were analyzed in parallel. Bilayered vesicles with diameters at the 100-300 nm range were found in extracellular fractions from yeast cultures. Proteomic analysis of vesicular fractions from the cells aforementioned and additional mutants with defects in conventional secretion pathways (sec1-1, fusion of Golgi-derived exocytic vesicles with the plasma membrane; bos1-1, vesicle targeting to the Golgi complex) or MVB functionality (vps23, late endosomal trafficking) revealed a complex and interrelated protein collection. Semi-quantitative analysis of protein abundance revealed that mutations in both MVB- and Golgi-derived pathways affected the composition of yeast extracellular vesicles, but none abrogated vesicle production. Lipid analysis revealed that mutants with defects in Golgi-related components of the secretory pathway had slower vesicle release kinetics, as inferred from intracellular accumulation of sterols and reduced detection of these lipids in vesicle fractions in comparison with WT cells. CONCLUSIONS/SIGNIFICANCE: Our results suggest that both conventional and unconventional pathways of secretion are required for biogenesis of extracellular vesicles, which demonstrate the complexity of this process in the biology of yeast cells. (hide) EV-METRIC 29% (58th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Yeast Sample origin NAY Focus vesicles extracellular vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Filtration UF Protein markers EV: non-EV: Proteomics yes Show all info Study aim Biogenesis/Sorting Sample Species Saccharomyces cerevisiae Sample Type Yeast Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 60 Filtration steps > 0.45 µm, Characterization: Protein analysis Characterization: Particle analysis DLS EM EM-type transmission EM Image type Close-up

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EV-TRACK ID	EV100062
species	Saccharomyces cerevisiae
sample type	Yeast
condition	NAY
separation protocol	(d)(U)C Filtration UF
Exp. nr.	1
EV-METRIC %	29