Identification of α1-antitrypsin as a positive acute phase protein in ayu ( Plecoglossus altivelis) associated with Listonella anguillarum infection

Abstract

α1-Antitrypsin (AAT) is implicated in the regulation of a variety of mammalian immune responses and was recently identified as a major serpin in blood plasma of some fish. However, AAT expression following bacterial infection in fish has not been well described. In this study, we cloned the full-length ayu ( Plecoglossus altivelis) AAT gene cDNA. It contained a 1368-bp coding region, which encodes a 19-amino acids (aa) signal peptide and a 437-aa mature AAT containing the serpin’s signature sequence ( 427LKFDRPFMMLV 437). PNGase F digestion confirmed that the higher molecular mass of the serum AAT was caused by N-glycosylation. Phylogenetic analysis indicated that ayu AAT was closest to that of green spotted pufferfish. AAT transcripts were present in a variety of tissues, with the highest level in the liver. The real-time quantitative PCR data showed that AAT transcripts dramatically increased in various ayu tissues after Listonella anguillarum infection. Western blot analysis revealed that the serum AAT protein level significantly increased in response to inflammation, but displayed no significant changes after cadmium exposure or salinity challenge. This work represents the first report that identifies AAT as a positive acute-phase protein in ayu fish associated with bacterial infection, suggesting that it might play a role in fish innate immunity.