Influenza A Virus Induces Autophagosome by Inhibiting LTCC/Calpain 2/LC3A Signaling to Promote Viral Replication

l-type voltage-gated calcium channels (LTCC), which are accountable for the influx of extracellular Ca²⁺, have been discovered to play a crucial regulatory role in the process of autophagy. However, the regulatory role of LTCC in autophagy process induced by influenza A virus (IAV) infection remains largely unknown. Here, we found that IAV (H1N1/PR8) induced autophagosome accumulation consistent with previous studies but blocked the fusion of autophagosomes with lysosomes. Meanwhile, viral infection led to a persistent decline of the cytoplasmic calcium signal in A549 cells. Interestingly, activation of LTCC partially restored the cytoplasmic calcium signal, impeded the formation of autophagosomes, and hindered the replication of IAV. Conversely, hindering LTCC or suppressing Cav1.3, the primary isoform of LTCC in A549 cells, significantly enhanced autophagosome formation and IAV replication. Mechanistically, calpain 2, a calcium-dependent cysteine protease, mediated the inhibition of LTCC/Cav1.3 on autophagosome formation and IAV replication by cleaving the carboxyl-terminal (112-118aa) of Microtubule-associated protein 1 light chain 3A(MAP1LC3A). Our findings reveal that IAV infection inhibits the LTCC/Cav1.3-calpain 2-LC3A axis to induce autophagosome formation, contributing to better understanding of viral infection process and providing potential target for combating IAV infection.