XI International Meeting on Paracoccidioidomycosis

XI International Meeting on Paracoccidioidomycosis

Resume:16-2

Poster (Painel)

16-2

SECRETOME OF PARACOCCIDIOIDES BRASILIENSIS PB18 ANALYZED BY LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRY

Authors:

Milene Vallejo (UNIFESP - UNIVERSIDADE FEDERAL DE SAO PAULO) ; Ernesto Nakayasu (UTEP - UNIVERSITY OF TEXAS AT EL PASO) ; Alisson Matsuo (UNIFESP - UNIVERSIDADE FEDERAL DE SAO PAULO) ; Tiago Sobreira (LNBIO - LABORATORIO NACIONAL DE BIOCIENCIAS) ; Larissa Longo (UNIFESP - UNIVERSIDADE FEDERAL DE SAO PAULO) ; Igor Almeida (UTEP - UNIVERSITY OF TEXAS AT EL PASO) ; Rosana Puccia (UNIFESP - UNIVERSIDADE FEDERAL DE SAO PAULO)

Abstract

Our group has recently described in P. brasiliensis extracellular vesicular structures of 20-200 nm, carrying highly immunogenic α-linked galactopyranosyl (α-Gal) epitopes and antigenic molecules that are recognized by total sera from PCM patients. In the present work we characterized the secretome of P. brasiliensis, Pb18 isolate, using tandem mass spectrometry. Cell-free supernatants of fungal cultures in Ham’s defined medium-glucose were concentrated and ultracentrifuged at 100,000g to precipitate vesicles. Secreted proteins from Pb18 were separated into vesicle and vesicle-free supernantant (ves-free) fractions, digested with trypsin, and analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). We found 123 proteins highly enriched or exclusively identified in vesicles, against 200 in ves-free fractions; 105 proteins were detected in both fractions at different percentages of abundance relative to total abundance in the secretome. Using prediction tools we observed an enrichment of secreted proteins containing putative signal peptides and glycosylphosphatidylinositol-anchoring in the extracellular vesicles, suggesting that these structures could be involved in the traffic and secretion of those polypeptides. Vesicle proteins showed diverse functions, such as transduction, translation, metabolism, cell division, transport, and signaling. Enrichment analysis of gene ontology biological processes, compared among Pb18 vesicle and genome proteins, showed that groups containing signaling proteins, GTPase-mediated signal transduction, cell division, and nucleosome organization were highly enriched in vesicles. Interacting network detected 45 Pb18 vesicles proteins with enriched biological process GO categories. Enzymatic activities suggested the presence of laccase and phosphatase in extracellular vesicles, although they have not been detected in the secretome. Our results reinforce that the secretome analysis can bring new insights into the host-pathogen interaction, and may lead to the discovery of novel components involved in the host-parasite relationship. Support: FAPESP, CNPq, NIH (grants # 5G12RR008124-16A1 and 5G12RR008124-16A1S1) ਀

Keyword: PARACOCCIDIOIDES BRASILIENSIS, SECRETOME, VESICLES, LC-MS/MS

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