The vaccinia virus (VV) A27L gene encodes a 14 kDa protein that is required for the formation of intracellular enveloped virus (IEV) and, consequently, normal sized plaques. Data presented here show that A27L plays an additional role in VV assembly. When cells were infected with the VV WR32-7/Ind 14K, under conditions that repress A27L expression, transport of intracellular mature virus (IMV) from virus factories was inhibited and some IMV was found in aberrant association with virus crescents. In contrast, other VV mutants (vΔB5R and vΔF13L) that are defective in IEV formation produce IMV particles that are transported out of virus factories. This indicated a specific role for A27L in IMV transport. Induction of A27L expression at 10 h post-infection promoted the dispersal of clustered IMV particles, but only when microtubules were intact. Formation of IEV particles was also impaired when cells were infected with WR32-7/14K, a VV strain expressing a mutated form of the A27L protein; however, this mutation did not inhibit intracellular transport of IMV particles. Collectively, these data define two novel aspects of VV morphogenesis. Firstly, A27L is required for both IMV transport and the process of envelopment that leads to IEV formation. Secondly, movement of IMV particles between the virus factory and the site of IEV formation is microtubule-dependent.


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