A chymotrypsin-like serine protease from Portunus trituberculatus involved in pathogen recognition and AMP synthesis but not required for prophenoloxidase activation. Issue 66 (July 2017)

• Record Type: Journal Article
• Authors: Liu, Hourong; Liu, Yuan; Song, Chengwen; Cui, Zhaoxia
• Is Part Of: Fish & shellfish immunology. Issue 66(2017)
• Page Start: 307
• Page End: 316
• Publication Date: 2017-07
• Subjects: Portunus trituberculatus -- Clip domain serine protease -- Antimicrobial activity -- Microbial-binding activity -- Prophenoloxidase activating system; Fishes -- Immunology -- Periodicals; Shellfish -- Immunology -- Periodicals; Poissons -- Immunologie -- Périodiques; Crustacés -- Immunologie -- Périodiques; 571.9617
• DOI: 10.1016/j.fsi.2017.05.031
• Languages: English
• ISSNs: 1050-4648

Abstract

Abstract: Clip domain serine proteases (clip-SPs) play critical roles in various immune responses in arthropods, such as hemolymph coagulation, antimicrobial peptide (AMP) synthesis, cell adhesion and melanization. In the present study, we report the molecular and functional characterization of a clip domain serine protease (PtcSP2) from the swimming crab Portunus trituberculatus . The N-terminal clip domain and the C-terminal SP-like domain of PtcSP2 were expressed in Escherichia coli system, and assayed for their activities. Sequence similarity and phylogenetic analysis revealed that PtcSP2 may belong to the chymotrypsin family, which was confirmed by protease activity assay of the recombinant SP-like domain. The clip domain of PtcSP2 exhibited strong antibacterial activity and microbial-binding activity, suggesting the potential role in immune defense and recognition. Knockdown of PtcSP2 by RNA interference could significantly reduce PtcSP2 transcript levels, but neither decrease the total phenoloxidase (PO) activity in crab nor significantly alter the expression levels of serine protease inhibitors PtPLC and PtSerpin. These results indicate that PtcSP2 is not involved in the proPO system. However, suppression of PtcSP2 led to a significant change in the expression of AMP genes PtALFs and PtCrustin but not PtALF5. All these findings suggest that PtcSP2 is a multifunctional chymotrypsin-like serine protease and may participate in crab innate immunity by its antibacterial activity, immune recognition or regulation of AMP expression. Highlights: PtcSP2 is the first chymotrypsin-like serine protease reported in swimming crab. The clip domain of PtcSP2 had strong antibacterial and microbial-binding activity. Suppression of PtcSP2 transcript did not decrease the total PO activity in crab. Knockdown of PtcSP2 could significantly induce the expression of AMPs except PtALF5.