Abstract
The sialic acid specific humoral lectin, Pjlec of the freshwater crab Paratelphusa jacquemontii was investigated for its opsonin function with rabbit erythrocyte as target cell for phagocytosis by the crab’s hemocyte. The untreated or trypsin treated erythrocyte induced lectin response after challenge however failed when treated with neuraminidase evidently indicating glycan dependency for elicited immune response. Our observation of in vitro phagocytosis of the erythrocyte untreated or coated with serum, clarified serum appeared to be recognized and engulfed by hemocytes but when coated with isolated lectin Pjlec, the response was elicited. Moreover, with trypsin treated erythrocyte the response remained unchanged but neuraminidase or O-glycosidase treatment eliminated the response reaction. This suggested the sialic acid specific reaction of lectin with the erythrocyte and was essential for recognition to allow the lectin Pjlec to act as an opsonin. The flowcytometry observation affirmed the enhancement of phagocytosis by Pjlec coated hemocyte. The efficiency of in vitro hemolysis of Pjlec coated erythrocyte with hemocyte when compared to untreated erythrocyte with or without hemocyte also established the opsonic function of the lectin. The mechanism of phagocytosis and induction were dependent on specific recognition of the erythrocyte by the multivalent binding site of the lectin protein, and the elicitation of the immune response was a function of the sialoglycan surface. The pathway of the challenge suggested that after entry of nonself recognition by lectin was followed by induction and activation of phagocytosis by opsonic binding of the lectin.
Highlights
The innate immune system in crustacea primitive has evolved a complex and efficient form, comprising of humoral and cellular components to recognize the pathogens by pathogen associated molecular patterns (PAMPs) through pattern recognition receptors (PRR) that activate conserved host defense signaling pathways to trigger or control the expression of a variety of immune responsive genes (Akira et al 2006; Medzhitov and Janeway 2002)
The present study demonstrates the recognition process of nonself by Pjlec lectin based on specific ligand binding to surface glycan and the subsequently regulate the process of phagocytosis
The trypsin treated rabbit erythrocyte showed no variation in Hemagglutination assay (HA) and showed a value of 2048
Summary
The innate immune system in crustacea primitive has evolved a complex and efficient form, comprising of humoral and cellular components to recognize the pathogens by pathogen associated molecular patterns (PAMPs) through pattern recognition receptors (PRR) that activate conserved host defense signaling pathways to trigger or control the expression of a variety of immune responsive genes (Akira et al 2006; Medzhitov and Janeway 2002). The mechanism of phagocytosis consists of multistep processes and is complex varying among species based on the differences in functioning of recognition, binding and ingestion. It has been a matter of speculation if the hemocyte receptors are able to recognize the non self independent to opsonin or are dependent on opsonin for recognition and subsequent internalization of foreign entity. The C-type lectin, FcLec, from the Chinese white shrimp Fenneropenaeus chinensis was reported to interact with the pathogenic bacterium Vibrio anguillarum and facilitate bacterial clearance in vivo and functioned as an opsonin by protein interaction with β-integrin on hemocyte surface during clearance (Wang et al 2009). A galectin from the kuruma shrimp Marsupenaeus japonicas functions as an opsonin for microbial pathogens, interacting with hemocyte surface promoting their phagocytosis (Wang et al 2014)
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