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, solubilised and used in pull-down assays from total barley protein extracts. Many putative interactors were identified by LC-MS/MS, despite 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 resistance and response to pathogens. In addition, pull-down data indicated the possible interaction of BEC1054 with ribosomal proteins, including an elongation factor 1 gamma. This data supports the idea that BEC1054 binds to barley ribosomal proteins. The biological significance of this remains to be defined.