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A fibrinogen-related protein (Mnfico3) acts as a novel pattern recognition receptor in Macrobrachium nipponense
Nan Qin1, Mengjia Wu1, Ting Tang1
1The Key Laboratory of Zoological Systematics and Application, College of Life Sciences, Hebei University, Baoding, Hebei, 071002, China.
Abstract:
Fibrinogen-related proteins (FREPs) are widely found in both vertebrates as well as invertebrates, and they play a crucial role in host immunity. In this study, we isolated a novel ficolin gene (Mnfico3) from the oriental river prawn Macrobrachium nipponense. The complete cDNA sequence of Mnfico3 was 1133 bp long, containing an open reading frame of 765 bp coding for Mnfico3, a protein consisting of 254 amino acids. The Mnfico3 protein contained a putative N-terminal signal peptide and a fibrinogen-related protein domain present at the C-terminal. Phylogenetic analysis indicated that Mnfico3 had a closer evolutionary relationship with vertebrate ficolins than with its invertebrate homologues. Tissue distribution analysis indicated that Mnfico3 was predominantly expressed in muscle, in which its transcription was increased following bacterial challenge by Aeromonas veronii. Function analysis using recombinant protein revealed that rMnFico3 had broad-spectrum binding capacity to a variety of microorganisms and pathogen-associated molecular pattern (PAMP) ligands. Furthermore, rMnFico3 exhibited Ca2+-dependent agglutinating activity against microbes in vitro, and ability to attach to the hemocyte surface which promoted phagocytosis and subsequent clearance of invasive bacteria in vivo. Silencing rMnFico3 in prawn through RNAi did not alter the expression of antimicrobial peptide genes (ALF and Crustin). These results manifested that MnFico3 functioned as a potential pattern recognition receptor (PPR) to mediate cellular immune response by recognizing PAMPs, agglutinating invasive microbes, and promoting phagocytosis of hemocytes.
Insights
Researchers identified a novel fibrinogen-related protein (FREP) in prawns, MnFico3, which acts as a pattern recognition receptor (PPR). This protein recognizes pathogens, clumps microbes, and enhances phagocytosis for improved host immunity.
Area of Science:
- Immunology
- Marine Biology
- Molecular Biology
Background:
- Fibrinogen-related proteins (FREPs) are vital for host immunity across species.
- Ficolins, a type of FREP, are key components of innate immune systems.
- Understanding invertebrate FREPs provides insights into conserved immune mechanisms.
Purpose of the Study:
- To isolate and characterize a novel ficolin gene, Mnfico3, from the oriental river prawn Macrobrachium nipponense.
- To investigate the immune function of MnFico3 in response to bacterial infection.
- To determine the evolutionary relationship of Mnfico3 with other ficolins.
Main Methods:
- cDNA sequencing and phylogenetic analysis to identify and classify Mnfico3.
- Quantitative real-time PCR to analyze Mnfico3 tissue distribution and expression levels.
- Recombinant protein expression and functional assays to assess binding, agglutination, and phagocytosis-promoting activities.
- RNA interference (RNAi) to silence Mnfico3 and evaluate its role in immune gene expression.
Main Results:
- A novel ficolin gene, Mnfico3, was identified in Macrobrachium nipponense, encoding a 254-amino acid protein with a fibrinogen-related domain.
- MnFico3 expression was highest in muscle tissue and significantly upregulated after Aeromonas veronii challenge.
- Recombinant MnFico3 (rMnFico3) demonstrated broad-spectrum microbial binding, Ca2+-dependent agglutination, and enhanced hemocyte phagocytosis, leading to reduced bacterial load in vivo.
- Silencing Mnfico3 did not affect the expression of antimicrobial peptide genes ALF and Crustin.
Conclusions:
- MnFico3 functions as a pattern recognition receptor (PPR) in prawn innate immunity.
- It recognizes pathogen-associated molecular patterns (PAMPs), mediates microbial agglutination, and promotes phagocytosis by hemocytes.
- MnFico3 plays a crucial role in cellular immune responses against bacterial pathogens, independent of certain antimicrobial peptide pathways.
