Molecular characterization and cellular localization of a transmembrane C-type lectin receptor in hemocytes from the scallop Argopecten purpuratus

Recognition of potential pathogens is a crucial function of the immune system of every organism. In this context, great efforts have been made in the last years to characterize the diversity of pattern recognition receptors (PRRs) present in cultured aquatic species, such as bivalve mollusks. Among the great diversity of PRRs described so far, C-type lectin receptors (CLRs) comprise a large family of innate immune receptors that can recognize different carbohydrates present on the cell surface of microorganisms. CLRs can be found extracellularly in soluble form, or transmembrane, located on immune cell surfaces. To date, almost only soluble CLRs have been described in bivalves and so far, no transmembrane CLRs are identified in pectinids. In the present study, a sequence showing structural homologies to transmembrane CLRs (tmCLR) was characterized from scallop Argopecten purpuratus hemocytes, named ApCLR. The cDNA sequence displays 948 nucleotides, coding for a deduced protein of 316 amino acids with a molecular weight of 35.24 kDa. The protein sequence showed characteristic domains of a tmCLR, such as a signal peptide, an extracellular N-terminal carbohydrate recognition domain (CRD), one transmembrane domain and a cytoplasmic tail. Phylogenetic analysis revealed that ApCLR groups with soluble CLRs from scallops but also with transmembrane CLRs from vertebrates, such as chondrolectins and layilins. Gene expression analysis showed that ApCLR was constitutively present in gill, mantle, digestive gland, gonad and hemocyte tissues from non-immune challenged scallops and it was most strongly expressed in the gill and mantle tissues. Furthermore, ApCLR was significantly upregulated in scallop hemocytes at 6 h after scallop injection with heat-killed Vibrio splendidus. At the protein level, ApCLR is located at the plasma membrane of hemocytes, observed as punctate signals by immunofluorescence and confocal analysis. Results obtained in this study reveal the first transmembrane CLR in scallops. Also, results showed that ApCLR is present at the surface of hemocytes and increase its transcript availability during the immune challenge, suggesting a potential role in the recognition of microorganisms.