Isolation, Characterization, and Proteomic Analysis of Crude and Purified Extracellular Vesicles Extracted from Fusarium oxysporum f. sp. cubense

文献类型： 外文期刊

第一作者： Ahmad, Mudassar

作者： Ahmad, Mudassar;Liu, Yushan;Huang, Shiyi;Huo, Yile;Yi, Ganjun;Jamil, Wajeeha;Yang, Xiaofang;Zhang, Wei;Li, Yuqing;Xiang, Dandan;Huang, Huoqing;Liu, Siwen;Li, Chunyu;Liu, Chongfei;Wang, Wei

作者机构：

关键词： extracellular vesicles (EVs); Fusarium oxysporum f. sp. cubense; banana; gradient purification; proteomics

期刊名称：PLANTS-BASEL （ 影响因子：4.1； 五年影响因子：4.5 ）

ISSN： 2223-7747

年卷期： 2024 年 13 卷 24 期

页码：

收录情况： SCI

摘要： Extracellular vesicles (EVs) produced by Fusarium oxysporum f. sp. cubense (Foc) play vital roles in plant-pathogen interactions; however, the isolation of purified Foc TR4-EVs and their pathogenicity and proteomic profiles are not well studied. This study aims to isolate and characterize purified Foc TR4-EVs and compare their pathogenic effects and protein profiles with crude TR4-EVs. Foc TR4-EVs were isolated using ultracentrifugation and purified by iodixanol gradient centrifugation. After characterization and evaluation of the pathogenicity effects on banana leaves, LC-MS/MS was performed to conduct the proteomics assay. Results indicated that Fraction 2 EVs exhibited clearer spherical structures (TEM), excessive abundance (1.70 x 109 particles/mL), greater intensity (400 a.u), mean size (154.5 nm), moderate protein content (333.16 ng/mu L), and protein profile (25-77 kDa), which were superior to Fractions 1, 3, and crude EVs. Crude EVs displayed significant background interference with EV structures (TEM), highest abundance (2.11 x 109 particles/mL), lower intensity (7.0 a.u), higher protein content (528.33 ng/mu L), and higher molecular weight proteins (55-70 kDa) compared to gradient EVs. A non-significant biocontrol effect of Foc-EVs on the growth of TR4 spores was observed. Pathogenicity assays revealed that crude EVs caused the largest (2.805 cm2), while Fraction 2 (1.386 cm2) and Fraction 3 (1.255 cm2) resulted in moderate lesions on banana leaves. Proteomic analysis identified 807 unique proteins in Fraction 2, enriched in pathways related to EV trafficking and signaling. In comparison, crude EVs contained 179 unique non-EV proteins related to metabolism and secondary metabolites, indicating that non-EV proteins of crude EVs also influence the pathogenicity observed in banana leaves. This study emphasizes the importance of EV purification, with Fraction 2 being a critical focus for future research on Foc EV pathogenicity.

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