Another major conclusion from this study is that the 5 0 -end of segment 2 mRNA itself has a number of overlapping functions. These include coding sequences critical for PB1 function, regulation of expression of downstream ORFs and also regions important for vRNA packaging. This has practical implications by reinforcing that this region represents an attractive target for therapeutic intervention, either by anti-viral drugs [e.g. those targeting the PB1-PA protein interface; (48) ] or through T-cell epitope immunization (49) , because the chance of finding escape mutations that maintain all functions of the protein/RNA sequence is likely to be lower than in a less functionally intricate area of the virus genome. Understanding the overlapping functional requirements also provides an interesting perspective on the evolutionary selection pressures that could be operating in this region of the influenza genome.