There were also discrepancies between virus infection and in vitro data for the AUG3/sORF2 mutant A78T virus regarding PB1-F2 expression, as the mutation decreased accumulation of the protein to $50% of normal instead of increasing it. Equally however, there was no evidence of expression of a larger form of PB1-F2 ( Figure 4A, lane 16) . Here, we surmise that the larger product was unstable in infected cells, and that the remaining F2 expression came from ribosomes initiating normally at AUG4 via leaky scanning. Consistent with this hypothesis, combining the A78T and T120C mutations (the latter removing AUG4) led to the loss of all detectable PB1-F2 accumulation ( Figure 4A, lane 26) .