We also characterized the role of C-terminus acidic residues in NSs functions. We found that both NSs-E253-255A/D257-259A and NSsD257-259A were not abundantly accumulated in infected cells. On the other hand, the NSs-E253-255A accumulated efficiently in cells, and showed a similar phenotype with authentic NSs. Thus, the aspartic acids at 257-259 but not glutamic acid at 253-255 must be important for the stability of NSs. Since the accumulation of NSs-E253-255A/D257-259A and NSsD257-259A were very low, we could not determine whether those mutants are lacking the functions to degrade PKR or inhibit IFNb mRNA synthesis.