| Scavenger receptors(SRs)are important pattern recognition receptors(PRRs),which play significant roles in host defense against pathogens by identifying pathogen-associated molecular patterns(PAMPs).Class B scavenger receptors have been shown to promote phagocytosis in shrimp.In this study,we investigated the ability of Class B scavenger receptors to promote phagocytosis in Eriocheir sinensis.In this study,we report the cloning and characterization of a SR from Eriocheir sinensis(EsSR-B1)which is a 500 amino acid protein encoded by a gene comprised of 2726 nucleotides with a 1503 bp open reading frame.The domains of EsSR-B1 were found to be evolutionarily conserved.EsSR-B1 was widely detected in different tissues of E.sinensis by qRT-PCR analysis,and it was abundant in intestine and hepatopancreas.EsSR-B1 was significantly up-regulated in hemocytes after stimulation by Staphyloccocus aureus or Vibrio parahaemolyticus.The recombinant EsSR-B1 protein was expressed by prokaryotic expression technology,and the protein was purified by affinity chromatography.Finally,the recombinant EsSR-B1 protein was obtained by renaturation.Recombinant EsSR-B1 protein could bind to bacteria and promote phagocytosis upon bacterial stimulation.Subsequently,based on the RNA interference technique in hemocytes of Eriocheir sinensis,we detected the phagocytosis rate by immunofluorescence assay and flow cytometry.We found that the phagocytic ability of hemocytes decreased after RNA interference of EsSR-B1.Moreover,antimicrobial peptide expression was reduced in EsSR-B1-silenced hemocytes after challenge by S.aureus or V.parahaemolyticus.Our study found EsSR-B1 has a critical role in the binding of bacteria and subsequent promotion of hemocyte phagocytosis.This study also has opened up new ideas for the study of the scavenger receptors in Chinese mitten crabs and enriched the study of innate immunity of Chinese mitten crabs. |
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