Infection with the F11-VK virus, however, did not result in a reduction in the level of GTP-bound RhoA at 8 hours post infection as seen with WR. Moreover, the level of GTP-bound RhoA in F11-VK infected cells is similar to that observed during DF11L infections. Consistent with this, we observed that F11-VK infected cells also had prominient actin stress fibres at 8 hours post infection. Our observations demonstrate that the direct binding of F11 to GTP bound RhoA and subsequent inhibition of its down stream signalling is required for the remodelling of the actin cytoskeleton observed during vaccinia infection. We observed that the DF11L and F11-VK viruses were still capable of inducing the formation of actin tails. However, it was noticeable that cells infected with the DF11L and F11-VK viruses had significantly fewer actin tails than those infected with WR. This similar reduction in the number of actin tails could be due to defects in virus replication and assembly. However, F11 is not required for virus replication and assembly. An alternative explanation for the reduction in number of actin tails is that the inability of the DF11L and F11-VK viruses to down regulate RhoA signalling impairs the ability of IEV particles to reach and fuse with the plasma membrane. In agreement with this, we found that there is a significant reduction in the percentage of infectious virus particles released into the media from cells infected DF11L and F11-VK viruses as compared to WR. These data, which agree with our previous observations using RNAi and over expression approaches in WR infected cells, suggest that F11-mediated inhibition of RhoA is likely to play an important role in the cell-to-cell spread of vaccinia. To directly examine if this is the case, we Terazosin HCl imaged the spread of WR, DF11L and F11-VK viruses expressing YFP-A3, an abundant viral core protein, in confluent monolayers of BS-C-1 cells during plaque formation. The cell monolayer was imaged for 48 hours immediately following the detection of YFP-A3 expression in the perinuclear Naftidrofuryl oxalate region of a single infected cell,8 hours post infection.