Viruses take advantage of host post-translational modifications for their own benefit. It was recently reported that influenza A virus proteins interact extensively with the host sumoylation system. Thereby, several viral proteins, including NS1 and M1, are sumoylated to facilitate viral replication. However, to what extent sumoylation is exploited by influenza A virus is not fully understood. In this study, we found that influenza A virus nucleoprotein (NP) is a bona fide target of sumoylation in both NP-transfected cells and virus-infected cells. We further identified that NP is sumoylated at the two most N-terminal residues, lysine 4 and lysine 7, and that sumoylation at lysine 7 of NP is highly conserved across different influenza A subtypes and strains, including the recently emerged human H7N9 virus. While NP stability and polymerase activity are little affected by sumoylation, the NP-sumoylation-defective virus, WSN-NP_K4,7R virus, exhibited an early cytoplasmic localization of NP. The growth of the WSN-NP_K4,7R virus was highly attenuated compared to that of WSN-WT virus, and lysine residue at position 7 is indispensable for the virus survival as illustrated by the rapid emergence of revertant viruses. Thus, sumoylation of influenza A virus NP is essential for intracellular trafficking of NP and for virus growth, illustrating sumoylation as a crucial strategy extensively exploited by influenza A virus for survival in the host.