Emodin induces defensive autophagy against white spot syndrome virus infection via targeting the hemocyanin-mediated endoplasmic reticulum-mitochondrial crosstalk in shrimp

White spot syndrome virus (WSSV) devastates shrimp aquaculture, yet safe antivirals remain scarce. Here we identify a druggable host-directed pathway centered on hemocyanin (HMC). This pathway coordinates endoplasmic reticulum (ER)-mitochondrial crosstalk to promote WSSV replication, which could be counteracted by the natural anthraquinone emodin. In Litopenaeus vannamei, emodin suppressed viral replication with a half maximal inhibitory concentration (IC₅₀) of 1.174 μM, improved survival in both therapeutic and prophylactic regimens, remained effective after per os administration, and retained Antiviral activity in water for up to 4 d. Target fishing, orthogonal biophysics, and docking analyses show that emodin binds HMC (K_d = 4.49 μM) and interferes with HMC-HSPA5/BiP (heat shock protein family A (HSP70) member 5) association. Mechanistically, emodin weakens the ITPR/IP₃R (inositol 1,4,5-trisphosphate receptor)-VDAC (voltage dependent anion channel)-MCU (mitochondrial calcium uniporter) conduit at ER-mitochondrial contact sites, thereby limiting CA²⁺ transfer, restoring mitochondrial membrane potential and organelle spacing, and attenuating ER stress. Untargeted metabolomics revealed that WSSV induced phosphoinositide and amino acid dysregulation, whereas emodin selectively normalizes phosphatidylinositol (PtdIns) and phosphatidylinositol 1,4,5-trisphosphate (PtdIns[1,4,5]P₃) signaling and rebalances amino acid, tricarboxylic acid (TCA) intermediates. Correspondingly, emodin inhibited phosphoinositide 3-kinase (PI3K)-AKT/protein kinase B-MTOR (mechanistic target of rapamycin kinase) complex 1 (mTORC1) signaling and restored lysosome-dependent macroautophagy/Autophagy. HMC RNAi or ITPR and MCU inhibition phenocopied emodin, whereas exogenous HMC or ER stress activation exacerbated Infection and was mitigated by emodin or MCU blockade. These findings establish HMC-anchored ER-mitochondrial contact as a central proviral vulnerability and position emodin as a practical scaffold for next-generation antivirals in aquaculture.Abbreviations: 2-APB: 2-aminoethyl diphenylborinate; 3-MA: 3-methyladenine; ΔΨm: mitochondrial membrane potential; Akt/protein kinase B: Akt serine/threonine kinase; ATG1/ULK1: Autophagy related 1; ATG13: Autophagy related 13; BSA: bovine serum albumin; CETSA: cellular thermal shift assay; Co-IP: co-immunoprecipitation; DAPI: 4',6-diamidino-2-phenylindole; DDIT3/Chop: DNA damage inducible transcript 3; DMSO: dimethyl sulfoxide; EAS6B: epoxy-activated sepharose 6B; EGTA: ethylene glycol tetraacetic acid; EIF4EBP1: eukaryotic translation initiation factor 4E binding protein 1; EPC: epithelioma papulosum cyprini; ER: endoplasmic reticulum; HMC: hemocyanin; HPLC-MS: high performance liquid chromatography-mass spectrometry; HRP: horseradish peroxidase; HSPA5/BiP: heat shock protein family A (HSP70) member 5; HSPA9/GRP75: heat shock protein family A (HSP70) member 5; ITPR/IP₃R: inositol 1,4,5-trisphosphate receptor; KEGG: Kyoto Encyclopedia of Genes and Genomes; LAMP1: lysosome associated membrane protein 1; LC-MS: liquid chromatography-mass spectrometry; MAMs: mitochondria-associated ER membranes; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCU: mitochondrial calcium uniporter; MTOR: mechanistic target of rapamycin kinase; MTORC1: mTOR complex 1; OPLS-DA: orthogonal partial least squares-discriminant analysis; PA: phosphatidic acid; PBS: phosphate-buffered saline; PC: phosphatidylcholine; PCA: principal component analysis; PE: phosphatidylethanolamine; PI3K: phosphoinositide 3-kinase; PLS-DA: partial least squares-discriminant analysis; PS: phosphatidylserine; PtdIns: phosphatidylinositol; PtdIns(1,4,5)P₃: phosphatidylinositol (1,4,5)-trisphosphate; PtdIns(3,4,5)P₃: phosphatidylinositol (3,4,5)-trisphosphate; PtdIns(4,5)P₂: phosphatidylinositol (4,5)-bisphosphate; RPS6KB1/S6K1: ribosomal protein S6 kinase B1; SDS-PAGE: sodium dodecyl sulfate-polyacrylamide gel electrophoresis; SQSTM1/p62: sequestosome 1; TCA: tricarboxylic acid cycle; TEM: transmission electron microscopy; TFEB: transcription factor EB; UPR: unfolded protein response; VDAC: voltage dependent anion channel; WSSV: white spot syndrome virus; XBP1: X-box binding protein 1.