title: Secretome analysis identifies potential virulence factors of Diplodia corticola, a fungal pathogen involved in cork oak (Quercus suber) decline.
abstract: The characterisation of the secretome of phytopathogenic fungi may contribute to elucidate the molecular mechanisms of pathogenesis. This is particularly relevant for Diplodia corticola, a fungal plant pathogen belonging to the family Botryosphaeriaceae, whose genome remains unsequenced. This phytopathogenic fungus is recognised as one of the most important pathogens of cork oak, being related to the decline of cork oak forests in the Iberian Peninsula. Unfortunately, secretome analysis of filamentous fungi is limited by the low protein concentration and by the presence of many interfering substances, such as polysaccharides, which affect the separation and analysis by 1D and 2D gel electrophoresis. We compared six protein extraction protocols concerning their suitability for further application with proteomic workflows. The protocols involving protein precipitation were the most efficient, with emphasis on TCA-acetone protocol, allowing us to identify the most abundant proteins on the secretome of this plant pathogen. Approximately 60% of the spots detected were identified, all corresponding to extracellular proteins. Most proteins identified were carbohydrate degrading enzymes and proteases that may be related to D. corticola pathogenicity. Although the secretome was assessed in a noninfection environment, potential virulence factors such as the putative glucan-β-glucosidase, neuraminidase, and the putative ferulic acid esterase were identified. The data obtained forms a useful basis for a deeper understanding of the pathogenicity and infection biology of D. corticola. Moreover, it will contribute to the development of proteomics studies on other members of the Botryosphaeriaceae.
authors.affiliation: Department of Biology, CESAM, Aveiro University, Campus Universitário de Santiago, 3810-193 Aveiro, Portugal; Department of Biochemistry, Physiology and Microbiology, Laboratory of Protein Biochemistry and Protein Engineering, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium.
authors.forename: Isabel
authors.initials: I
authors.lastname: Fernandes
authors.affiliation: Department of Biology, CESAM, Aveiro University, Campus Universitário de Santiago, 3810-193 Aveiro, Portugal.
authors.forename: Artur
authors.initials: A
authors.lastname: Alves
authors.affiliation: Department of Biology, CESAM, Aveiro University, Campus Universitário de Santiago, 3810-193 Aveiro, Portugal.
authors.forename: António
authors.initials: A
authors.lastname: Correia
authors.affiliation: Department of Biochemistry, Physiology and Microbiology, Laboratory of Protein Biochemistry and Protein Engineering, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent, Belgium.
authors.forename: Bart
authors.initials: B
authors.lastname: Devreese
authors.affiliation: Department of Biology, CESAM, Aveiro University, Campus Universitário de Santiago, 3810-193 Aveiro, Portugal. Electronic address: acesteves@ua.pt.
authors.forename: Ana Cristina
authors.initials: AC
authors.lastname: Esteves
mesh_terms: D001203:Ascomycota / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000382:microbiology*; D021381:Protein Transport; D040901:Proteomics; D029963:Quercus / Q000382:microbiology*; D037521:Virulence Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D037521:Virulence Factors
keywords: 2D-electrophoresis; Botryosphaeriaceae; Filamentous fungi; Plant pathogen; Quercus suber