Identification of RUVBL1 and RUVBL2 as novel cellular interactors of the Ebola virus nucleoprotein

M. Jane Morwitzer, Sarah R. Tritsch, Lisa H. Cazares, Michael D. Ward, Jonathan E. Nuss, Sina Bavari, St Patrick Reid

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Ebola virus (EBOV) is a filovirus that has become a global public health threat in recent years. EBOV is the causative agent of a severe, often fatal hemorrhagic fever. A productive viral infection relies on the successful recruitment of host factors for various stages of the viral life cycle. To date, several investigations have discovered specific host-pathogen interactions for various EBOV proteins. However, relatively little is known about the EBOV nucleoprotein (NP) with regard to host interactions. In the present study, we aimed to elucidate NP-host protein-protein interactions (PPIs). Affinity purification-mass spectrometry (AP-MS) was used to identify candidate NP cellular interactors. Candidate interactors RUVBL1 and RUVBL2, partner proteins belonging to the AAA+ (ATPases Associated with various cellular Activities) superfamily, were confirmed to interact with NP in co-immunoprecipitation (co-IP) and immunofluorescence (IF) experiments. Functional studies using a minigenome system revealed that the siRNA-mediated knockdown of RUVBL1 but not RUVBL2 moderately decreased EBOV minigenome activity. Super resolution structured illumination microscopy (SIM) was used to identify an association between NP and components of the R2TP complex, which includes RUVBL1, RUVBL2, RPAP3, and PIH1D1, suggesting a potential role for the R2TP complex in capsid formation. Moreover, the siRNA-mediated knockdown of RPAP3 and subsequent downregulation of PIH1D1 was shown to have no effect on minigenome activity, further suggesting a role in capsid formation. Overall, we identify RUVBL1 and RUVBL2 as novel interactors of EBOV NP and for the first time report EBOV NP recruitment of the R2TP complex, which may provide novel targets for broad-acting anti-EBOV therapeutics.

Original languageEnglish (US)
Article number372
JournalViruses
Volume11
Issue number4
DOIs
StatePublished - Apr 2019

Fingerprint

Ebolavirus
Nucleoproteins
Capsid
Small Interfering RNA
Proteins
Host-Pathogen Interactions
Virus Diseases
Life Cycle Stages
Lighting
Immunoprecipitation
Fluorescent Antibody Technique
Adenosine Triphosphatases
Microscopy
Mass Spectrometry
Fever
Down-Regulation
Public Health

Keywords

  • AAA+ proteins
  • Ebola
  • NP
  • R2TP
  • RUVBL1
  • RUVBL2

ASJC Scopus subject areas

  • Infectious Diseases
  • Virology

Cite this

Morwitzer, M. J., Tritsch, S. R., Cazares, L. H., Ward, M. D., Nuss, J. E., Bavari, S., & Reid, S. P. (2019). Identification of RUVBL1 and RUVBL2 as novel cellular interactors of the Ebola virus nucleoprotein. Viruses, 11(4), [372]. https://doi.org/10.3390/v11040372

Identification of RUVBL1 and RUVBL2 as novel cellular interactors of the Ebola virus nucleoprotein. / Morwitzer, M. Jane; Tritsch, Sarah R.; Cazares, Lisa H.; Ward, Michael D.; Nuss, Jonathan E.; Bavari, Sina; Reid, St Patrick.

In: Viruses, Vol. 11, No. 4, 372, 04.2019.

Research output: Contribution to journalArticle

Morwitzer, MJ, Tritsch, SR, Cazares, LH, Ward, MD, Nuss, JE, Bavari, S & Reid, SP 2019, 'Identification of RUVBL1 and RUVBL2 as novel cellular interactors of the Ebola virus nucleoprotein', Viruses, vol. 11, no. 4, 372. https://doi.org/10.3390/v11040372
Morwitzer MJ, Tritsch SR, Cazares LH, Ward MD, Nuss JE, Bavari S et al. Identification of RUVBL1 and RUVBL2 as novel cellular interactors of the Ebola virus nucleoprotein. Viruses. 2019 Apr;11(4). 372. https://doi.org/10.3390/v11040372
Morwitzer, M. Jane ; Tritsch, Sarah R. ; Cazares, Lisa H. ; Ward, Michael D. ; Nuss, Jonathan E. ; Bavari, Sina ; Reid, St Patrick. / Identification of RUVBL1 and RUVBL2 as novel cellular interactors of the Ebola virus nucleoprotein. In: Viruses. 2019 ; Vol. 11, No. 4.
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abstract = "Ebola virus (EBOV) is a filovirus that has become a global public health threat in recent years. EBOV is the causative agent of a severe, often fatal hemorrhagic fever. A productive viral infection relies on the successful recruitment of host factors for various stages of the viral life cycle. To date, several investigations have discovered specific host-pathogen interactions for various EBOV proteins. However, relatively little is known about the EBOV nucleoprotein (NP) with regard to host interactions. In the present study, we aimed to elucidate NP-host protein-protein interactions (PPIs). Affinity purification-mass spectrometry (AP-MS) was used to identify candidate NP cellular interactors. Candidate interactors RUVBL1 and RUVBL2, partner proteins belonging to the AAA+ (ATPases Associated with various cellular Activities) superfamily, were confirmed to interact with NP in co-immunoprecipitation (co-IP) and immunofluorescence (IF) experiments. Functional studies using a minigenome system revealed that the siRNA-mediated knockdown of RUVBL1 but not RUVBL2 moderately decreased EBOV minigenome activity. Super resolution structured illumination microscopy (SIM) was used to identify an association between NP and components of the R2TP complex, which includes RUVBL1, RUVBL2, RPAP3, and PIH1D1, suggesting a potential role for the R2TP complex in capsid formation. Moreover, the siRNA-mediated knockdown of RPAP3 and subsequent downregulation of PIH1D1 was shown to have no effect on minigenome activity, further suggesting a role in capsid formation. Overall, we identify RUVBL1 and RUVBL2 as novel interactors of EBOV NP and for the first time report EBOV NP recruitment of the R2TP complex, which may provide novel targets for broad-acting anti-EBOV therapeutics.",
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