An Intrinsically Disordered Peptide from Ebola Virus VP35 Controls Viral RNA Synthesis by Modulating Nucleoprotein-RNA Interactions.

Zbyszek Otwinowski,Manu Anantpadma,Ian B. Harvey,Daisy W. Leung,Juanli Pan,Dominika Borek,Priya Luthra,Ariel Endlich-Frazier,Robert A. Davey,Jennifer M. Binning,Gaya K. Amarasinghe,Wah Chiu,Reed S. Shabman,Zhaoming Su,Christopher F. Basler,Gai Liu

doi:10.1016/j.celrep.2015.03.034

Abstract

During viral RNA synthesis, Ebola virus (EBOV) nucleoprotein (NP) alternates between an RNA-template-bound form and a template-free form to provide the viral polymerase access to the RNA template. In addition, newly synthesized NP must be prevented from indiscriminately binding to noncognate RNAs. Here, we investigate the molecular bases for these critical processes. We identify an intrinsically disordered peptide derived from EBOV VP35 (NPBP, residues 20-48) that binds NP with high affinity and specificity, inhibits NP oligomerization, and releases RNA from NP-RNA complexes invitro. The structure of the NPBP/ΔNPNTD complex, solved to3.7Å resolution, reveals how NPBP peptide occludes a large surface area that is important for NP-NP and NP-RNA interactions and for viral RNA synthesis. Together, our results identify a highly conserved viral interface that is important for EBOV replication and can be targeted for therapeutic development.

Highlights

Ebolaviruses and marburgviruses are nonsegmented negative sense RNA viruses (NNSVs) that cause severe hemorrhagic fever (Sanchez et al, 2006)
Viral genome replication and transcription of individual genes into distinct 5′-capped, 3′-polyadenylated monocistronic mRNAs are carried out by the viral RNA-dependent RNA polymerase complex (RdRp) (Muhlberger, 2007). This complex consists of the enzymatic component of the RDRP, the large protein (L), along with the viral nucleoprotein (NP), viral protein (VP)35 and VP30 and acts upon the nucleocapsid, which consists of single-stranded viral genomic and antigenomic RNAs (Muhlberger, 2007) that are encapsidated by NP
We observed >60% monodispersed ΔNPNTD protein in solution at concentrations >10 mg/mL based on multiangle light scattering (MALS) studies

Summary

Introduction

Ebolaviruses and marburgviruses are nonsegmented negative sense RNA viruses (NNSVs) that cause severe hemorrhagic fever (Sanchez et al, 2006). Viral genome replication and transcription of individual genes into distinct 5′-capped, 3′-polyadenylated monocistronic mRNAs are carried out by the viral RNA-dependent RNA polymerase complex (RdRp) (Muhlberger, 2007). This complex consists of the enzymatic component of the RDRP, the large protein (L), along with the viral nucleoprotein (NP), viral protein (VP) and VP30 and acts upon the nucleocapsid, which consists of single-stranded (ss) viral genomic and antigenomic RNAs (Muhlberger, 2007) that are encapsidated by NP. The EBOV core nucleocapsid protects ssRNAs from degradation, similar to that observed for paramyxoviruses and rhabdoviruses (Masters and Banerjee, 1988)

Results

Discussion

Conclusion