Determination of host proteins composing the microenvironment of coronavirus replicase complexes by proximity-labeling
- Institute of Virology and Immunology IVI, Switzerland
- University of Bern, Switzerland
Abstract
Positive-sense RNA viruses hijack intracellular membranes that provide niches for viral RNA synthesis and a platform for interactions with host proteins. However, little is known about host factors at the interface between replicase complexes and the host cytoplasm. We engineered a biotin ligase into a coronaviral replication/transcription complex (RTC) and identified >500 host proteins constituting the RTC microenvironment. siRNA-silencing of each RTC-proximal host factor demonstrated importance of vesicular trafficking pathways, ubiquitin-dependent and autophagy-related processes, and translation initiation factors. Notably, detection of translation initiation factors at the RTC was instrumental to visualize and demonstrate active translation proximal to replication complexes of several coronaviruses. Collectively, we establish a spatial link between viral RNA synthesis and diverse host factors of unprecedented breadth. Our data may serve as a paradigm for other positive-strand RNA viruses and provide a starting point for a comprehensive analysis of critical virus-host interactions that represent targets for therapeutic intervention.
Data availability
The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD009975.All other data generated or analysed during this study are included in the manuscript and supporting files.
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Proximity biotinylation and LC-MSMS identification of the molecular microenvironment of coronavirus replication complexesProteomeXchange Consortium, PXD009975.
Article and author information
Author details
Funding
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (173085)
- Philip V'kovski
- Volker Thiel
European Commission (748627)
- Stephanie Pfaender
- Volker Thiel
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (160780)
- Jenna Kelly
- Nadine Ebert
- Volker Thiel
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, V'kovski et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Determination of host proteins composing the microenvironment of coronavirus replicase complexes by proximity-labeling
eLife 8:e42037.
https://doi.org/10.7554/eLife.42037
