Hemocyanin regulates phosphatidylcholine metabolism to enhance reactive oxygen species–mediated immunity in Pacific white shrimp ( Penaeus vannamei )

Acute hepatopancreatic necrosis disease (AHPND) is one of the most severe threats to global shrimp aquaculture. Recent studies report a significant reduction in hepatopancreas hemocyanin levels in Penaeus vannamei (PvHMC) by AHPND. To investigate the functional implications, we performed liquid chromatography-tandem mass spectrometry-based metabolomics and found that PvHMC knockdown caused a marked accumulation of phosphatidylcholine (PC). qPCR and ELISA analyses further showed that PvHMC suppression upregulated the mRNA expression, protein abundance, and enzymatic activity of PvCPT1, a key enzyme in PC synthesis. Co-immunoprecipitation and confocal immunofluorescence analyses confirmed a direct interaction between PvHMC and PvCPT1, indicating that PvHMC regulates PC metabolism by binding to PvCPT1. Functional assays demonstrated that exogenous PC supplementation significantly increased reactive oxygen species (ROS) production, whereas simultaneous knockdown of PvHMC and PvCPT1 reduced plasma PC levels and ROS activity. Under pathogenic challenge, PC administration inhibited Vibrio parahaemolyticus (VpAHPND) proliferation by enhancing intracellular ROS-mediated antimicrobial responses, ultimately improving shrimp survival. In summary, PvHMC regulates PC metabolism to modulate ROS levels, strengthening shrimp's immune defense against pathogenic infections. These findings reveal a novel metabolic regulatory role of PvHMC in response to VpAHPND infection and suggest potential strategies for controlling AHPND in shrimp aquaculture.