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Article

The Cellular Protein CAD is Recruited into Ebola Virus Inclusion Bodies by the Nucleoprotein NP to Facilitate Genome Replication and Transcription

Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institut, 17493 Greifswald-Insel Riems, Germany
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Cells 2020, 9(5), 1126; https://doi.org/10.3390/cells9051126
Received: 30 March 2020 / Revised: 28 April 2020 / Accepted: 29 April 2020 / Published: 1 May 2020
(This article belongs to the Special Issue Cell Biology of Viral Infections)
Ebola virus (EBOV) is a zoonotic pathogen causing severe hemorrhagic fevers in humans and non-human primates with high case fatality rates. In recent years, the number and extent of outbreaks has increased, highlighting the importance of better understanding the molecular aspects of EBOV infection and host cell interactions to control this virus more efficiently. Many viruses, including EBOV, have been shown to recruit host proteins for different viral processes. Based on a genome-wide siRNA screen, we recently identified the cellular host factor carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) as being involved in EBOV RNA synthesis. However, mechanistic details of how this host factor plays a role in the EBOV life cycle remain elusive. In this study, we analyzed the functional and molecular interactions between EBOV and CAD. To this end, we used siRNA knockdowns in combination with various reverse genetics-based life cycle modelling systems and additionally performed co-immunoprecipitation and co-immunofluorescence assays to investigate the influence of CAD on individual aspects of the EBOV life cycle and to characterize the interactions of CAD with viral proteins. Following this approach, we could demonstrate that CAD directly interacts with the EBOV nucleoprotein NP, and that NP is sufficient to recruit CAD into inclusion bodies dependent on the glutaminase (GLN) domain of CAD. Further, siRNA knockdown experiments indicated that CAD is important for both viral genome replication and transcription, while substrate rescue experiments showed that the function of CAD in pyrimidine synthesis is indeed required for those processes. Together, this suggests that NP recruits CAD into inclusion bodies via its GLN domain in order to provide pyrimidines for EBOV genome replication and transcription. These results define a novel mechanism by which EBOV hijacks host cell pathways in order to facilitate genome replication and transcription and provide a further basis for the development of host-directed broad-spectrum antivirals. View Full-Text
Keywords: Ebola virus; filovirus; inclusion bodies; CAD; pyrimidine synthesis Ebola virus; filovirus; inclusion bodies; CAD; pyrimidine synthesis
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MDPI and ACS Style

Brandt, J.; Wendt, L.; Bodmer, B.S.; Mettenleiter, T.C.; Hoenen, T. The Cellular Protein CAD is Recruited into Ebola Virus Inclusion Bodies by the Nucleoprotein NP to Facilitate Genome Replication and Transcription. Cells 2020, 9, 1126. https://doi.org/10.3390/cells9051126

AMA Style

Brandt J, Wendt L, Bodmer BS, Mettenleiter TC, Hoenen T. The Cellular Protein CAD is Recruited into Ebola Virus Inclusion Bodies by the Nucleoprotein NP to Facilitate Genome Replication and Transcription. Cells. 2020; 9(5):1126. https://doi.org/10.3390/cells9051126

Chicago/Turabian Style

Brandt, Janine, Lisa Wendt, Bianca S. Bodmer, Thomas C. Mettenleiter, and Thomas Hoenen. 2020. "The Cellular Protein CAD is Recruited into Ebola Virus Inclusion Bodies by the Nucleoprotein NP to Facilitate Genome Replication and Transcription" Cells 9, no. 5: 1126. https://doi.org/10.3390/cells9051126

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