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Elife. 2019 Jan 11;8. pii: e42037. doi: 10.7554/eLife.42037.

Determination of host proteins composing the microenvironment of coronavirus replicase complexes by proximity-labeling.

Author information

1
Institute of Virology and Immunology IVI, Bern, Switzerland.
2
Department of Infectious Diseases and Pathobiology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
3
Graduate School for Biomedical Science, University of Bern, Bern, Switzerland.
4
Interfaculty Bioinformatics Unit, SIB Swiss Institute of Bioinformatics, University of Bern, Bern, Switzerland.
5
Mass Spectrometry and Proteomics Core Facility, Department for BioMedical Research (DBMR), University of Bern, Bern, Switzerland.
6
Department of Clinical Research, University of Bern, Bern, Switzerland.
7
Division of Veterinary Anatomy, Vetsuisse Faculty, University of Bern, 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.

KEYWORDS:

coronavirus; human; infectious disease; microbiology; mouse; proximity labeling; replicase complex; translation; vesicular transport; virus; virus-host interaction

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