Cloning, Expression and Bioactivity of Wild Ruddy Shelduck (Tadorna ferruginea) Interleukin-2

Abstract

Interleukin-2 (IL-2) is one of the most actively studied interleukins. It is a powerful growth factor for a variety of cell types, plays a central role in the cell differentiation and stimulates the activity of many kinds of killer cells. However, little information is available on the IL-2 genes in migratory waterfowl, ecologically important species potentially involved in virus transmission. In this study, the cDNA of wild ruddy shelduck IL-2 was cloned by RT-PCR from peripheral blood lymphocytes stimulated with Concanavalin A (ConA), and the recombinant ruddy shelduck IL-2 (rRSIL-2) mature protein was expressed in Escherichia coli. In vitro bioactivity of rRSIL-2 was demonstrated in a lymphocyte proliferation assay. The results showed that RSIL-2 cDNA contained an open reading frame (ORF) of 423 bp encoding a protein of 140 amino acids (aa) with a putative signal peptide of 21 aa. Phylogenetic analysis based on nucleotide sequences showed that RSIL-2 was most closely related to IL-2s of the muscovy duck and mallard (homology of 97.4% and 96.2%), and to a lesser extent to spot-billed duck and swan goose sequences (homology of 95.8% and 93.6%). The RSIL-2 showed genetic diversity compared with other birds of anseriformes. Fusion protein expressed in E. coli had the molecular weight of around 31.7 kDa, as shown by SDS-PAGE and western blotting. The rRSIL-2 affected the duck, chicken, and goose lymphocyte proliferation in the same concentration, although the effect on goose and chicken lymphocytes was relatively weak. Our study reported the molecular signatures and bioactivity of IL-2 in a migratory waterfowl species.