Two novel ficolin‐like proteins act as pattern recognition receptors for invading pathogens in the freshwater crayfish Pacifastacus leniusculus

Abstract

To isolate pathogen-associated molecular patterns (PAMPs)-binding molecules, the bacterium, Staphylococcus aureus was used as an affinity matrix to find bacteria-binding proteins in the plasma of the freshwater crayfish, Pacifastacus leniusculus. Two new bacteria-binding ficolin-like proteins (FLPs) were identified by 2-DE and MS analysis. The FLPs have a fibrinogen-related domain (FReD) in their C-terminal and a repeat region in their N-terminal regions with putative structural similarities to the collagen-like domain of vertebrate ficolins and mannose binding lectins (MBLs). Phylogenetic analysis shows that the newly isolated crayfish FLP1 and FLP2 cluster separately from other FReD-containing proteins. A tissue distribution study showed that the mRNA expression of FLP occurred mainly in the hematopoietic tissue (Hpt) and in the hepatopancreas. Recombinant FLPs exhibited agglutination activity of Gram-negative bacteria Escherichia coli and Aeromonas hydrophila in the presence of Ca(2+) . The FLPs could bind to A. hydrophila, E. coli as well as S. aureus as judged by bacteria adsorption. Moreover, the FLPs may help crayfish to clear Gram-negative bacteria, but not Gram-positive bacteria which had been injected into the hemolymph. When Gram-negative bacteria coated with FLPs were incubated with Hpt cells, a lower death rate of the cells was found compared with control treatment. Our results suggest that FLPs function as pattern recognition receptors in the immune response of crayfish.

Highly diverse fibrinogen-related proteins in the Pacific oyster Crassostrea gigas

“…Far fewer studies exist on invertebrate FREPs compared to studies conducted on vertebrates [27e29]. From what we do know, the primary role of FREPs in invertebrates appears to be defense against pathogens [8,9,11,27,30,31]. This implies that invertebrate FREPs must have a large degree of flexibility so that the immune system can adequately respond to the numerous and rapidly evolving pathogens they encounter.…”

“…Specifically, we uncovered 193 FREP proteins with 205 FBG domains in C. gigas, Previous studies have reported that FREP proteins share highly conserved C-terminal FBG domains, while their N-terminal regions vary significantly, resulting in a highly diverse gene family. For example, the immunoglobulin superfamily domains in the N-terminal regions of B. glabrata FREPs [3,32] differ from FREPs in the sea squirt (Halocynthia roretzi) [6] and crayfish (Pacifastacus leniusculus) [31], which both have one short collagen-like domain. Our study further demonstrates the diversity of this gene family by uncovering multiple, overlapping FBG domains in the C. gigas FREPs we screened.…”

Characteristic and functional analysis of a ficolin-like protein from the oyster Crassostrea hongkongensis

“…These differences might reflect the selective binding of ChFCN to E. coli K-12 but . Furthermore, these proteins could bind to A. hydrophila, E. coli as well as Staphylococcus aureus [34]. To confirm the interaction between ChFCN and E. coli K-12, an agglutination assay was performed in the presence of Ca 2þ , and the results indicated that ChFCN interacts with E. coli K-12.…”

Variable immune molecules in invertebrates

“…forming aggregates (Gokudan et al, 1999;Kairies et al, 2001). In a crustacean, two ficolin-like proteins were recently demonstrated to bind to Gramnegative bacteria and this event was followed by the clearance of the bacteria (Wu et al, 2011).…”