Many bacterial pathogens express virulence proteins that are translocated into host cells (herein referred to as effectors), where they can interact with target proteins to manipulate host cell processes. These effector–host protein interactions are often dynamic and transient in nature, making them difficult to identify using traditional interaction-based methods. Here, we performed a systematic comparison between proximity-dependent biotin labelling (BioID) and immunoprecipitation coupled with mass spectrometry to investigate a series of Salmonella type 3 secreted effectors that manipulate host intracellular trafficking (SifA, PipB2, SseF, SseG and SopD2). Using BioID, we identified 632 candidate interactions with 381 unique human proteins, collectively enriched for roles in vesicular trafficking, cytoskeleton components and transport activities. From the subset of proteins exclusively identified by BioID, we report that SifA interacts with BLOC-2, a protein complex that regulates dynein motor activity. We demonstrate that the BLOC-2 complex is necessary for SifA-mediated positioning of Salmonella-containing vacuoles, and affects stability of the vacuoles during infection. Our study provides insight into the coordinated activities of Salmonella type 3 secreted effectors and demonstrates the utility of BioID as a powerful, complementary tool to characterize effector–host protein interactions.
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Quantitative proteomic screen identifies annexin A2 as a host target for Salmonella pathogenicity island-2 effectors SopD2 and PipB2
Scientific Reports Open Access 08 December 2021
Nature Communications Open Access 19 November 2021
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The data that support the findings of this study are available from the corresponding authors upon request.
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J.H.B. holds the Pitblado Chair in Cell Biology. Infrastructure for the Brumell Laboratory was provided by a John Evans Leadership Fund grant from the Canadian Foundation for Innovation and the Ontario Innovation Trust. V.M.D. was supported by fellowships from the Canadian Institutes of Health Research and the Hospital for Sick Children Research Training Centre, and is a recipient of the L’Oreal-UNESCO Women in Science Award. K.C.B. was supported by scholarships from the Natural Sciences and Engineering Research Council of Canada and the Ontario Graduate Scholarship. We thank Paul Paroutis for help with confocal microscopy. This work was supported by operating grants from the Canadian Institutes of Health Research (grant no. FDN#154329 to J.H.B., grant no. FDN#143202 to S.G. and grant no. MOP#119289 to B.R.). B.R. holds a Canada Research Chair in Proteomics and Molecular Medicine.
The authors declare no competing interests.
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Supplementary Figs. 1–11, original source images, Supplementary Table legends and Supplementary References.
IP–MS profiling of S. Typhimurium effectors.
BioID profiling of S. Typhimurium effectors.
BioID and IP–MS datasets enrichment analysis.
BioID and IP–MS individual baits individual and comparative enrichment analysis.
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D’Costa, V.M., Coyaud, E., Boddy, K.C. et al. BioID screen of Salmonella type 3 secreted effectors reveals host factors involved in vacuole positioning and stability during infection. Nat Microbiol 4, 2511–2522 (2019). https://doi.org/10.1038/s41564-019-0580-9
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