Cryo-ET of infected cells reveals that a succession of two lattices drives vaccinia virus assembly

Abstract

During replication, vaccinia virus and other poxviruses such as monkeypox first assemble non-infectious spherical immature virions (IV) in the cytoplasm of infected cells. The IV membrane, which is fully coated by a viral D13 lattice, encapsidates both viral proteins and the DNA genome. Subsequently, the D13 lattice is lost and IV transitions into the brick-shaped intracellular mature virus (IMV) with an organised core enclosing the genome. We have an overview of the maturation of IV into IMV but our structural understanding remains limited. Here, we performed cryo-electron tomography of frozen-hydrated vaccinia-infected cells to structurally characterise the maturation process in situ. We found that D13-coated IV have similar diameters and lattice dimensions to spherical D13 assemblies formed in vitro, confirming the size of IV is determined by D13. During IMV formation a new pseudohexagonal lattice forms inside IV to produce the viral core. This lattice is composed of trimeric pillars with outward projections that appears as a palisade in cross-section. Strikingly, the diameter of IV precisely matches the IMV long axis, suggesting the length of the core, and hence the IMV, is determined by the D13 lattice. During maturation the viral membrane becomes corrugated as it adapts to the shape of the core. This suggests that the 50% reduction in volume during the transition of IV to IMV does not require membrane removal. Our observations suggest that the consecutive D13 and palisade lattices structurally define vaccinia assembly and maturation.