Cellular protein synthesis is suppressed during influenza virus infection, allowing for preferential production of viral proteins. To explore the impact of polymerase subunits on protein synthesis, we co-expressed eGFP or luciferase together with each polymerase component or NS1 of A/California/04/2009 (Cal) and found that PA has a significant impact on the expression of eGFP and luciferase. Comparison of the suppressive activity on co-expressed proteins between various strains revealed that avian virus or avian-origin PAs have much stronger activity than human-origin PAs, such as the one from A/WSN/33 (WSN). Protein synthesis data suggested that reduced expression of co-expressed proteins is not due to PA´s reported proteolytic activity. A recombinant WSN containing Cal PA showed enhanced host protein synthesis shutoff and induction of apoptosis. Further characterization of the PA fragment indicates that the N-terminal domain (PANt), which includes the endonuclease active site, is sufficient to suppress co-transfected gene expression. By characterizing various chimeric PANt, we found that multiple regions of PA, mainly the helix α4 and the flexible loop of amino acids 51-74, affect the activity. The suppressive effect of PANt cDNA was mainly due to PA-X, which was expressed by ribosomal frameshifting. In both Cal and WSN viruses, PA-X showed a stronger effect than the corresponding PANt, suggesting that the unique C-terminal sequences of PA-X also play a role in suppressing co-transfected gene expression. Our data indicate strain variations in PA gene products, which play a major role in suppression of host protein synthesis.