The novel mitochondria localization of influenza A virus NS1 visualized by FlAsH labeling

The nonstructural protein 1 (NS1) of the influenza A virus (IAV) is a multifunctional protein that counteracts host cell antiviral responses and inhibits host cell pre‐mRNA processing. NS1 contains two nuclear localization signals that facilitate NS1 shuttling between cytoplasm and nucleus. In this study, we initially observed the novel mitochondria localization of NS1 in a subset of transfected cells. We then further monitored the localization dynamics of the NS1 protein in live cells infected with IAV expressing NS1 with insertion of a tetracysteine‐tag. The resulting mutant virus showed similar levels of infectivity and expression pattern of NS1 to those of wild‐type IAV. Pulse labeling using a biarsenical compound (fluorescein arsenical hairpin binder) allowed us to visualize the dynamic subcellular distribution of NS1 real time. We detected NS1 in mitochondria at a very early infection time point [1.5 h postinfection (hpi)] and observed the formation of a granular structure pattern in the nucleus at 4 hpi. This is the first identification of the novel mitochondria localization of NS1. The possible role of NS1 at an early infection time point is discussed.