A trypsin-like serine protease domain of masquerade gene in crayfish Procambarus clarkii could activate prophenoloxidase and inhibit bacterial growth

Masquerade (Mas) is a secreted trypsin-like serine protease (SPs) and involved in immune response in some arthropods. However, according to previous studies, Mas presents different functional activities. In the present study, the functional mechanisms of Mas in crayfish Procambarus clarkii immune defense were studied. A fragment cDNA sequence of PcMas was identified and characterized. From the structural analysis, it contains a trypsin-like serine protease domain. The highest expression level of PcMas was detected in hepatopancreas. The infection of A. hydrophila could induce the expression of PcMas, while the WSSV infection did not cause changes in the expression of PcMas. Through the prokaryotic expression system, the PcMas protein was expressed in E. coli. It was verified that PcMas can bind to bacteria in vitro and inhibit the growth of the bacteria. By dsRNA interference with the expression of PcMas, the decrease expression of PcMas led to a decrease in the activity of phenoloxidase in hemolymph and an increase of mortality caused by A. hydrophila infection. The injection of recombinant protein can enhance the activity of phenoloxidase and reduce mortality caused by A. hydrophila infections. Therefore, the present study confirmed that PcMas could improve the body's immune response to eliminate bacterial pathogens by binding with bacteria and activating the prophenoloxidase system. The results will enrich the molecular mechanisms of crustaceans immune defense.