Heat shock protein A1L restricts influenza A virus by ubiquitination of NA.

Neuraminidase (NA) protein of influenza A virus (IAV), a tetrameric envelope glycoprotein critical for viral release and spread, has well-characterized enzymatic functions but poorly resolved host-pathogen interactions. However, host factors targeted by NA or those regulating NA activity are largely uncharacterized. Here, we identified host heat shock protein A1L (HSPA1L) as a novel NA-binding partner that suppresses IAV replication. Subsequent studies revealed that HSPA1L promotes NA ubiquitination at K242, resulting in its NBR1-dependent autophagic degradation. Moreover, a recombinant H5N1 virus carrying the NAK242R mutation exhibited enhanced replication and resistance to HSPA1L-mediated viral suppression compared to the wild-type (WT) virus. Collectively, we unveil an intrinsic defense mechanism, wherein HSPA1L antagonizes IAV by promoting NBR1-dependent autophagic degradation of NA via site-specific ubiquitination.IMPORTANCEIAV, especially avian influenza virus H5N1, begins to cause the infection and/or death for its new uncommon host, especially sea mammals and ruminants, indicating that the virus is adapting to mammalian infections. These developments substantially elevate concerns about pandemic potential and zoonotic risk escalation, necessitating a deeper understanding of host-IAV interactions to develop effective antiviral countermeasures. HSP70 family proteins are known to modulate viral infections. However, the specific role of HSPA1L, a member of the HSP70 family, in IAV infection remains poorly characterized. In this study, we demonstrate that HSPA1L defends against IAV by regulating NA proteostasis. Mechanistically, HSPA1L directly binds NA, promotes its ubiquitination at K242, and mediates NBR1-dependent autophagic degradation, thereby suppressing viral replication. Our study identifies HSPA1L as a promising target for antiviral strategies against IAV.