Background

Poxviruses, including cowpox (smallpox, variola) and Mpox, are large, double-stranded DNA (dsDNA) viruses that replicate in the host cell cytoplasm. After infection, the genome remains protected within a dense protein core, and viral messenger RNA (mRNA) produced by early transcription is released to the outside. However, the structure and molecular composition of the channel that penetrates the core wall have remained elusive. Existing structural biology techniques have only captured fragmented forms of isolated viruses, failing to elucidate the actual activity of the complex during cell infection. Therefore, to fundamentally understand the poxvirus replication mechanism, it is necessary to determine the actual three-dimensional structure of the core wall nanogate.

Key Findings

A research team from the Francis Crick Institute in the United Kingdom captured the structure of the Portal Complex of the Vaccinia Virus (VACV) core wall using cryo-electron tomography (cryo-ET) and sub-tomogram averaging (STA) techniques. The analysis revealed the complex at an overall resolution of 7.1 Ångströms (Å), and the internal pore at a resolution of 4.9 Å. This complex is a hexameric structure composed of three conserved viral proteins, E8, E6, and L3, bound together. The E6 protein, which forms the center of the pore, interacts with the scaffold layer of the core wall, forming a framework. The E8 protein forms a ring on the outside of this cylinder, in the cytoplasmic direction, and the L3 protein is bound in the form of a hexamer of dimers on the inside. In particular, the L3 protein forms the narrowest region of the channel, serving as a target for the host cell's immune factor, TRIM5α. The geometry and electrostatic properties of the pore are designed to selectively allow only single-stranded RNA to pass through. This filter system protects the internal dsDNA while allowing mRNA to escape into the cytoplasm. The research team confirmed that there are approximately 20 portals per core, and that the D5 helicase, which is responsible for genome release, binds to the E8 ring, triggering the opening of the channel.

Significance and Prospects

The three-dimensional structure revealed in this study provides a key to understanding the physical secrets by which poxviruses evade host cell immune sensors such as cGAS and survive. The virus thoroughly conceals its genome with a rigid core wall and a fine-scale filter in the portal, while releasing only transcripts into the cytoplasm. This mechanism provides new clues for the development of smallpox and Mpox therapeutics. By designing compounds that interfere with the binding of the E8 ring and D5 helicase or target the L3 protein, it is possible to develop candidate substances that can fundamentally block the release of the viral genome. However, the vaccinia virus structure in this model may not fully represent the subtle differences in variant poxviruses or Mpox viruses. A limitation that needs to be overcome in the future is the inability to observe the dynamic changes of the pore opening and closing in real-time in an actual infection environment.