Functional characterization of two clip-domain serine proteases in the swimming crab Portunus trituberculatus. Issue 89 (June 2019)

• Record Type: Journal Article
• Authors: Liu, Hourong; Liu, Yuan; Song, Chengwen; Ning, Junhao; Cui, Zhaoxia
• Is Part Of: Fish & shellfish immunology. Issue 89(2019)
• Page Start: 98
• Page End: 107
• Publication Date: 2019-06
• Subjects: Portunus trituberculatus -- Clip domain serine protease -- Antimicrobial activity -- Bacterial clearance -- Prophenoloxidase activating system -- Complement-like factors; Fishes -- Immunology -- Periodicals; Shellfish -- Immunology -- Periodicals; Poissons -- Immunologie -- Périodiques; Crustacés -- Immunologie -- Périodiques; 571.9617
• DOI: 10.1016/j.fsi.2018.12.047
• Languages: English
• ISSNs: 1050-4648

Abstract

Abstract: Clip domain serine proteases (cSPs), a family of multifunctional proteins, play a crucial role in innate immune system. Here, we report the functional characterization of two clip domain serine proteases (PtcSP1 and PtcSP3) from the swimming crab Portunus trituberculatus . The recombinant N-terminal clip domains and the C-terminal SP-like domains of PtcSP1 and PtcSP3 were expressed in Escherichia coli system, and assayed for various biological functions: protease activity, antimicrobial activity, bacterial clearance and microbial-binding activity. The recombinant SP-like domains of PtcSP1 and PtcSP3 exhibited trypsin-like protease activity, while their recombinant clip domains showed strong antibacterial activity and could bind to bacteria and yeast, suggesting the potential roles of PtcSP1 and PtcSP3 in immune defense and pattern recognition. Unlike PtcSP3, PtcSP1 revealed the opsonic activity as shown by a higher bacterial clearance rate of Vibrio alginolyticus coated with the combination of the recombinant clip domain and SP-like domain of PtcSP1 as compared with V. alginolyticus only. Knockdown of PtcSP1 or PtcSP3 by RNA interference resulted in a significant decrease of total phenoloxidase (PO) activity in crab, suggesting that PtcSP1 and PtcSP3 are involved in the proPO system. In addition, suppression of PtcSP1 or PtcSP3 changed the expression of PtALFs and complement-like components. All these findings suggest that PtcSP1 and PtcSP3 are multifunctional immune molecules and perform different protective functions in crab defense. Highlights: PtcSP1 and PtcSP3 are two trypsin-like serine proteases. The clip domain of PtcSP1 and PtcSP3 had strong antibacterial and microbial-binding activity. PtcSP1 functioned as an opsonin. Suppression of PtcSP1 or PtcSP3 transcript reduce the total PO activity in crab. Knockdown of PtcSP1 or PtcSP3 could significantly change the expression levels of complement-like factors and AMPs.