Two fibrinogen-related proteins (FREPs) in the razor clam (Sinonovacula constricta) with a broad recognition spectrum and bacteria agglutination activity

Highlights

• Two fibrinogen-related proteins from the razor clam Sinonovacula constricta.

• Exploring their roles in immune defense as potential pattern recognition receptors.

• Identification of pathogen-associated molecular patterns.

Abstract

Fibrinogen-related proteins (FREPs) that contain only the fibrinogen-related domain are likely involved in pathogen recognition. In this study, we identified two FREPs from the razor clam (Sinonovacula constricta), called ScFREP-1 and ScFREP-2, and investigated their roles in the immune response. Both ScFREP-1 and ScFREP-2 contained a fibrinogen-related domain at the C-terminal. ScFREP-1 and ScFREP-2 mRNAs were detected in all adult clam tissues tested, with the highest expression levels in the gill and mantle, respectively. Their expression levels were significantly upregulated after microbe infection. Recombinant ScFREPs could bind Gram-positive and Gram-negative bacteria as well as some pathogen-associated molecular patterns (PAMPs), and they could agglutinate those bacteria. These results showed that ScFREPs functioned as potential pattern recognition receptors to mediate immune response by recognizing PAMPs and agglutinating invasive microbes.

Introduction

Invertebrates lack lymphocytes and an antibody-based humoral immune system and therefore have to rely on innate immune reactions to prevent pathogen invasion (Loker et al., 2004). When foreign microbes come into contact with a host, the recognition of pathogens by pattern recognition receptors (PRRs) initiates the first step of defense (Janeway and Medzhitov, 2002). These PRRs are important for non-self recognition to sense specific and conserved molecules called pathogen-associated molecular patterns (PAMPs), such as peptidoglycan (PGN) of Gram-positive bacteria and lipopolysaccharide (LPS) of Gram-negative bacteria. Although important advances have been made in understanding invertebrate innate immunity in recent years, there is still no comprehensive view of the immune mechanisms in invertebrate phyla (Akira et al., 2006).

In invertebrates, fibrinogen-related proteins (FREPs) act as PRRs to participate particularly in innate immunity (Hanington and Zhang, 2011). FREPs have been found in several animal species (Dai et al., 2017; Zhang et al., 2009). Their functions in invertebrates include participation in the defense process, such as pathogen recognition and bacterial defense. In the Mediterranean mussel (Mytilus galloprovincialis), expression of FREPs increased after bacterial infection or PAMP treatment, and they exhibited opsonic activities similar to mammalian ficolins (Romero et al., 2011). Furthermore, recombinant ficolin-like proteins of the freshwater crayfish Pacifastacus leniusculus helped the organism clear Gram-negative bacteria injected into the hemolymph (Wu et al., 2011).

The razor clam (Sinonovacula constricta), a member of the Solenidae family of bivalve molluscs, is widely distributed in muddy intertidal zones along the coasts of the western Pacific Ocean. Considering its relatively short production cycle and high production efficiency, the razor clam has become one of the most valuable economic species for bivalve fisheries in China. However, pathogenic diseases frequently occur, leading to reduced harvest yields of this clam. Thus, it is necessary to gain deeper insight into the molecular mechanisms underlying innate immunity in S. constricta. In this study, we cloned and characterized two FREPs from S. constricta and provided the first evidence of their potential roles in immune defense. The results of this study provided clues about the role of this kind of PRR in the molluscan immune response.