The vaccinia virus F11L gene product facilitates cell detachment and promotes migration.

We previously showed that infection with vaccinia virus (VV) induces cell motility, characterized by contractility and directed migration. Motility is temporally regulated because cells are motile immediately after infection, whereas late in infection motility ceases and cells resettle. Motility and its cessation are accompanied by temporal rearrangements of both the microtubule and the actin networks. Because the F11L gene has previously been implicated in VV-induced migration, we now explore the role of F11L in contractility, migration, the cessation of motility and the cytoskeletal rearrangements. By live cell imaging using a VV that lacks an intact F11L gene, we show that F11L facilitates cell detachment and is required for migration but not for contractility. By light microscopy, F11L expression induces a remodeling of the actin, but not the microtubule, network. The lack of migration correlates with smaller plaques, indicating that this process facilitates cell-to-cell spreading of VV. Late in infection, when motility ceases, cells re-establish cell-to-cell contacts in an F11L-independent manner. We finally show that VV-induced motility and its cessation correlate with a temporal regulation of the guanosine triphosphatase RhoA as well as the expression levels of F11L during the infectious cycle.

An alternative genetic method to test essential vaccinia virus early genes.

The vaccinia virus F11L gene product was identified during search for additional factors involved in the control of post-replicative viral gene transcription elongation. F11L is a 1065 base pairs (354 aminoacids) gene expressed early during infection with no attributed function. The F11L gene is conserved in many but not all poxviruses. The essential presence of the F11L gene was tested using two different genetic methods. F11L gene disruption by insertion of a selectable cassette containing the Escherichia coli guanine phosphoribosyl transferase gene driven by the viral early-late 7.5K transcriptional promoter resulted exclusively in recombinant viruses containing both the wild type and disrupted alleles, indicating that the F11L gene was essential. However, an alternative test, using transient dominant selection to insert nonsense mutations into the F11L gene, proved that the F11L gene was non-essential for growth in culture. These experiments suggest that misleading results can be obtained using gene insertional mutagenesis as a test of essential presence of the gene. The experiments also provide genetic data on the probability of co-insertion of linked mutations in vaccinia virus genome using transient dominant selection.