Characterization and functional analysis of the serpin-10 gene from oak silkworm, Antheraea pernyi (Lepidoptera: Saturniidae)

Abstract

Serpin is a broadly distributed superfamily of proteins that have a crucial role in regulating various immune reactions. Herein we identified a serpin-10 gene from Antheraea pernyi that encodes a 1557 amino acid residue protein with a predicted molecular weight of 58.76 kDa. Recombinant Apserpin-10 protein was expressed in a prokaryotic expression system (Escherichia coli) and the purified protein was used to prepare rabbit anti-Apserpin-10 polyclonal antibodies. Quantitative real-time polymerase chain reaction and western blot analysis indicate that Apserpin-10 was transcribed in all the tissues examined, including haemolymph, malpighian tubules, fat body, silk gland, integument and mid gut; the greatest expression level of Apserpin-10 was recorded in the fat body and haemocytes. The comparison of different developmental stages showed that Apserpin-10 transcript level was highest in 5th instar larvae, while the lowest expression was recorded at the egg stage. We also investigated the expression patterns of Apserpin-10 in fat body and haemocyte samples, following administration of heat-inactivated gram-positive bacteria (Micrococcus luteus), gram negative bacteria (Escherichia coli), a fungus (Beauveria bassiana) and virus (nuclear polyhedrosis virus, NPV). A substantial up-regulation of Apserpin-10 expression was recorded following pathogen challenge in both the tissues tested. Further the knock down of Apserpin-10 led to down regulation of antimicrobial peptide genes. Altogether, our results indicate that Apserpin-10 is involved in the innate immunity of A. pernyi.