Interaction of the influenza virus nucleoprotein with the cellular CRM1-mediated nuclear export pathway

Influenza virus transcription occurs in the nuclei of infected cells, where the viral genomic RNAs are complexed with a nucleoprotein (NP) to form rib onucleoprotein (RNP) structures. Prior to assembly into progeny virions, th ese RNPs exit the nucleus and accumulate in the cytoplasm. The mechanisms r esponsible for RNP export are only partially understood but have been propo sed to involve the viral M1 and NS2 polypeptides. We found that the drug le ptomycin B (LMB), which specifically inactivates the cellular CRM1 polypept ide, caused nuclear retention of NP in virus-infected cells, indicating a r ole for the CRM1 nuclear export pathway in RNP egress. However, no alterati on was seen in the cellular distribution of M1 or NS2, even in the case of a mutant virus which synthesizes greatly reduced amounts of NS2. Furthermor e, NP was distributed throughout the nuclei of infected cells at early time s postinfection but, when retained in the nucleus at late times by LMB trea tment, was redistributed to the periphery of the nucleoplasm. No such chang e was seen in the nuclear distribution of M1 or NS2 after drug treatment. S imilar to the behavior of NP, M1 and NS2 in infected cells, LMB treatment o f cells expressing each polypeptide in isolation caused nuclear retention o f NP but not M1 or NS2. Conversely, overexpression of CRM1 caused increased cytoplasmic accumulation of NP but had little effect on M1 or NS2 distribu tion. Consistent with this, NP bound CRM1 in vitro. Overall, these data rai se the possibility that RNP export is mediated by a direct interaction betw een NP and the cellular CRM1 export pathway.