B-type allatostatin regulates immune response of hemocytes in mud crab Scylla paramamosain

B-type allatostatins (AST-B) are neuropeptides that have important physiological roles in arthropods, they have also been identified in a number of crustacean species. Recent research on neuroendocrine-immune (NEI) regulatory system in invertebrates has exploded, it reveals that the NEI network plays an indispensable role in optimizing the immune response and maintaining homeostasis. Herein, mud crab Scylla paramamosain provides a primitive and ancient model to study crosstalk between the neuroendocrine and immune systems. In this study, qRT-PCR analysis showed that the nervous system was the main production site for Sp-AST-B mRNA in S. paramamosain, while its receptor gene (Sp-AST-BR) mRNA could be detected in all the analyzed tissues including hemocytes. This reveals that AST-B might act as a pleiotropic neuropeptide. In situ hybridization further confirmed that granular cells of hemocyte subpopulations express Sp-AST-BR. Time-course analysis revealed that bacteria-analog LPS or virus-analog Poly (I:C) challenge significantly induced Sp-AST-B expression in the thoracic ganglion, and the expression of Sp-AST-BR in hemocytes were also positively changed. Furthermore, mud crabs treated with a synthetic AST-B peptide significantly increased the mRNA levels of AST-BR, nuclear factor-κB (NF-κB) transcription factor (Dorsal and Relish), pro-inflammatory cytokine (IL-16) and immune-effector molecules, and also dramatically enhanced the nitric oxide (NO) production and phagocytic activity in hemocytes. Meanwhile dsRNA-mediated knockdown of Sp-AST-B remarkably suppressed the NO concentrations, phagocytic activity and the expression of immune related genes, resulting in markedly impaired ability of crabs to inhibit bacterial proliferation in vivo. Combined, these data demonstrate that AST-B induced innate immune in the mud crab.