Assembly and Budding of Ebolavirus

Abstract

Ebolavirus is responsible for highly lethal hemorrhagic fever. Like all viruses, it must reproduce its various components and assemble them in cells in order to reproduce infectious virions and perpetuate itself. To generate infectious Ebolavirus, a viral genome-protein complex called the nucleocapsid (NC) must be produced and transported to the cell surface, incorporated into virions, and then released from cells. To further our understanding of the Ebolavirus life cycle, we expressed the various viral proteins in mammalian cells and examined them ultrastructurally and biochemically. Expression of nucleoprotein alone led to the formation of helical tubes, which likely serve as a core for the NC. The matrix protein VP40 was found to be critical for transport of NCs to the cell surface and for the incorporation of NCs into virions, where interaction between nucleoprotein and the matrix protein VP40 is likely essential for these processes. Examination of virus-infected cells revealed that virions containing NCs mainly emerge horizontally from the cell surface, whereas empty virions mainly bud vertically, suggesting that horizontal budding is the major mode of Ebolavirus budding. These data form a foundation for the identification and development of potential antiviral agents to combat the devastating disease caused by this virus.

Structural dissection of Ebola virus and its assembly determinants using cryo-electron tomography

“…These narrow structures can be recruited into VLPs by coexpression of VP40 (14). If NP is expressed together with VP24 and VP35, cytoplasmic clusters of NC-like structures are formed that are similar to those seen in infected cells (13). These structures are also recruited into VLPs when VP40 is coexpressed (7,(13)(14)(15).…”

“…Expression of VP40 alone in mammalian cells can lead to formation and release of enveloped, filamentous viruslike particles (VLPs) (9)(10)(11)(12). Expression of NP alone leads to the formation of narrow, tubular structures in the cytoplasm of the cell (13). These narrow structures can be recruited into VLPs by coexpression of VP40 (14).…”

Filovirus Replication and Transcription

“…For EBOV, the minimal protein requirements for the formation of nucleocapsid-like structures are NP, VP35 and VP24 [17,35]. Recently, Noda and colleagues have proposed a model for EBOV nucleocapsid formation and subsequent budding [23]. According to this model, NP forms helical tubes which differ from nucleocapsid-like structures.…”

“…VP35 and VP24 interact with the pre-formed NP-derived helices, resulting in the formation of nucleocapsid-like structures [36]. Both transport of the nucleocapsids to the plasma membrane and incorporation into virions are mediated by the interaction of NP with VP40 [23,37]. In addition, Johnson and colleagues have shown that EBOV VP35 interacts with the matrix protein VP40 and EBOV minigenomes, thus mediating the packaging of the minigenome RNA into virus-like particles [38].…”

“…These are the NP, the polymerase cofactor VP35, the transcription activator VP30, and the major component of the RNA-dependent RNA polymerase, L ( Figure 1C). The nucleocapsid proteins play a dual role in the viral replication cycle; they are structural components of the nucleocapsid complex and are, therefore, involved in viral morphogenesis and catalyze the replication and transcription of the RNA genome [22][23][24][25].…”