title: Interactions between the Powdery Mildew Effector BEC1054 and Barley Proteins Identify Candidate Host Targets.
abstract: There are over 500 candidate secreted effector proteins (CSEPs) or Blumeria effector candidates (BECs) specific to the barley powdery mildew pathogen Blumeria graminis f.sp. hordei. The CSEP/BEC proteins are expressed and predicted to be secreted by biotrophic feeding structures called haustoria. Eight BECs are required for the formation of functional haustoria. These include the RNase-like effector BEC1054 (synonym CSEP0064). In order to identify host proteins targeted by BEC1054, recombinant BEC1054 was expressed in E. coli, solubilized, and used in pull-down assays from barley protein extracts. Many putative interactors were identified by LC-MS/MS after subtraction of unspecific binders in negative controls. Therefore, a directed yeast-2-hybrid assay, developed to measure the effectiveness of the interactions in yeast, was used to validate putative interactors. We conclude that BEC1054 may target several host proteins, including a glutathione-S-transferase, a malate dehydrogenase, and a pathogen-related-5 protein isoform, indicating a possible role for BEC1054 in compromising well-known key players of defense and response to pathogens. In addition, BEC1054 interacts with an elongation factor 1 gamma. This study already suggests that BEC1054 plays a central role in barley powdery mildew virulence by acting at several levels.
authors.affiliation: Department of Life Sciences, Imperial College London , SW7 2AZ London, United Kingdom.
authors.forename: Helen G
authors.initials: HG
authors.lastname: Pennington
authors.affiliation: Department of Life Sciences, Imperial College London , SW7 2AZ London, United Kingdom.
authors.forename: Dana M
authors.initials: DM
authors.lastname: Gheorghe
authors.affiliation: Department of Life Sciences, Imperial College London , SW7 2AZ London, United Kingdom.
authors.forename: Annabelle
authors.initials: A
authors.lastname: Damerum
authors.affiliation: Department of Life Sciences, Imperial College London , SW7 2AZ London, United Kingdom.
authors.forename: Clara
authors.initials: C
authors.lastname: Pliego
authors.affiliation: Department of Life Sciences, Imperial College London , SW7 2AZ London, United Kingdom.
authors.forename: Pietro D
authors.initials: PD
authors.lastname: Spanu
authors.affiliation: Department of Chemistry, University of Reading , Reading RG6 6AD, United Kingdom.
authors.forename: Rainer
authors.initials: R
authors.lastname: Cramer
authors.affiliation: Department of Chemistry, University of Reading , Reading RG6 6AD, United Kingdom.|School of Biological Sciences, Royal Holloway University of London , Egham TW20 0EX, United Kingdom.
authors.forename: Laurence V
authors.initials: LV
authors.lastname: Bindschedler
mesh_terms: D001203:Ascomycota / Q000472:pathogenicity; D005656:Fungal Proteins / Q000633:toxicity; D001467:Hordeum / Q000737:chemistry*; D054884:Host-Pathogen Interactions; D010940:Plant Proteins / Q000032:analysis / Q000276:immunology*; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D053719:Tandem Mass Spectrometry; D014774:Virulence; D015003:Yeasts / Q000472:pathogenicity
chemical_list: D005656:Fungal Proteins; D010940:Plant Proteins
keywords: Blumeria graminis; RNase-like effector; barley; biotrophic pathogen; elongation factor; glutathione-S-transferase (GST); malate dehydrogenase; pathogenesis-related protein-5