Abstract
Crustins are a family of cysteine-rich antimicrobial peptides (AMPs), predominantly found in crustaceans, and play important roles in innate immunity. However, among the many reported crustins, few studies have explored their immunomodulatory functions. In this study, we investigated the immune function of a type I crustin (LvCrustinIa-2) in Litopenaeus vannamei, with particular emphasis on comparing the roles of its different domains. LvCrustinIa-2 possesses cationic patchy surface and amphipathic structure, and its expression was significantly induced in hemocytes after pathogen challenge. Both the recombinant LvCrustinIa-2 (rLvCrustinIa-2) and its whey acidic protein (WAP) domain (rLvCrustinIa-2-WAP) exhibited significant inhibitory activities against the tested Gram-positive bacteria. They also showed binding affinity not only for Gram-positive bacteria but also for Gram-negative bacteria. Furthermore, rLvCrustinIa-2 induced membrane leakage and structure damage in the target bacteria. Notably, chemotaxis assays revealed that rLvCrustinIa-2 and the synthetic cysteine-rich region (LvCrustinIa-2-CR) significantly enhanced the chemotactic activity of shrimp hemocytes in vitro. Knockdown of LvCrustinIa-2 triggered significant transcriptional activation of genes involved in calcium transport, inflammation, redox regulation, and NF-κB pathway. Taken together, these findings elucidate the distinct roles of the cysteine-rich region and WAP domain in type Ia crustin and provide the first evidence of a crustacean AMP with chemotactic and immunomodulatory activities.