S293 phosphorylation of hemocyanin boosts shrimp antimicrobial immunity via p38 MAPK signaling

Scaffold proteins serve as essential modulators of the mitogen-activated protein kinase (MAPK) signaling pathway, coordinating the assembly of kinase hierarchies to maintain accurate spatiotemporal modulation of phosphorylation-driven signal transduction. This study identifies the large subunit of Penaeus vannamei hemocyanin ( Pv HMCL) as a novel scaffold protein regulating shrimp innate immunity. Phosphorylation of Pv HMCL at the S293 site is a critical molecular switch that enhances the interaction and phosphorylation of Pv MKK4 and Pv MEKK4, two key components of the p38 MAPK signaling pathway. This modification triggers the activation of the p38 MAPK cascade, which in turn upregulates the synthesis of major antimicrobial peptides (AMPs) including crustins, anti-lipopolysaccharide factors, and penaeidins. Functional analysis confirms that the phosphorylation-mimetic mutant Pv HMCL-S293D significantly boosts P. vannamei resistance to Vibrio parahaemolyticus and Staphylococcus aureus infections, reducing bacterial load in the hepatopancreas. Collectively, these findings reveal that S293 phosphorylation enables Pv HMCL's scaffold function to orchestrate p38 MAPK pathway activation, highlighting a pivotal role of hemocyanin phosphorylation in enhancing this shrimp's antimicrobial defense and providing a theoretical basis for targeted disease control strategies in aquaculture.