A novel ferritin gene from Procambarus clarkii involved in the immune defense against Aeromonas hydrophila infection and inhibits WSSV replication

Abstract

ABSTRACT Ferritin is a protein related to the storage of iron and widely distributed in animals. It participates in many biological process, including antioxidation, cell activation, angiogenesis, regulation of iron metabolic balance and immune defense. In the present study, a novel ferritin gene was identified from red swamp crayfish Procambarus clarkii, with a cDNA sequence encoding a predicted 221 amino‐acid residues. The ferritin protein contains a 19‐residue signal peptide and 145‐residue classic ferritin domain. The NJ phylogenetic analysis showed PcFer clustered with other crustacean peptides. The recombinant PcFer protein was produced and purified in E. coli, and the anti‐rabbit polyclonal antibody was obtained. The rPcFer exhibited iron binding activity at a dose‐dependent effect. The qPCR and western blot analysis revealed that PcFer was highly expressed in hemocytes, hepatopancreas, and gills. After challenged with WSSV and Aeromonas hydrophila, the mRNA and protein expression patterns of PcFer were significantly up‐regulated in hemocytes and hepatopancreas. dsRNA interfering technique was utilized to silence the expression of PcFer gene. The WSSV copy number in PcFer silenced shrimp was much higher than that in the control group. The present study indicated that PcFer was involved in the immune defense against WSSV and Aeromonas hydrophila, and might inhibit WSSV replication in P. clarkii. These results will provide important data support for further study of the functional role of the ferritin gene. HighlightsThe protein and mRNA of PcFer were highly expressed in hemocyte and hepatopancreas.PcFer could be significantly up‐regulated after WSSV and A. hydrophila infections in hemocyte and hepatopancreas.The recombinant PcFer could exhibit iron binding activity.Silencing of PcFer genes caused increase of the copy number of WSSV.