Cloning, in vitro expression and bioactivity of duck interleukin-2.
In this report, the cDNA sequences of Shaoxing (SX) and Muscovy (MV) duck IL-2 were cloned, then recombinant duck IL-2 (rduIL-2) was produced in prokaryotic expression system. In vitro bioactivity of rduIL-2 was determined by lymphocyte proliferation assay and in vivo bioactivity of rduIL-2 was assessed by vaccine immunization. Monoclonal antibodies (mAbs) and polyclonal antibodies (pAbs) specific for rduIL-2 were generated and characterized by ELISA, Western blot and neutralizing assays. The cDNA contains an open reading frame (ORF) of 420-base pairs encoding a protein of 140 amino acids (aa) with a putative signal peptide of 21aa. The His-duIL-2 fusion protein was recognized in Western blot by mAb against chicken IL-2 (chIL-2), but not by mAbs against human IL-2 and mouse IL-2. Recombinant duIL-2 induces in vitro proliferation of Con A-stimulated duck splenocytes in MTT assay and strengthens duck immune responses induced by vaccinating the inactivated oil emulsion vaccine against avian influenza virus. Polyclonal antibodies and mAb 2B3 against rduIL-2 were shown to have effective neutralizing ability by inhibiting the biological activities of both recombinant duIL-2 and endogenous duIL-2. Despite the fact that duck and chicken IL-2s only share identity of 55.0-56.7% in amino acid sequence, duck and chicken IL-2 molecules displayed similar cross-priming activity in in vitro lymphocyte proliferation assays. The results, at the first time, indicated that rduIL-2 has the potential to be used as an immunoadjuvant for enhancing vaccine efficacy and an immunotherapeutic, and the mAbs against rduIL-2 further facilitate basic immunobiological studies of the role of IL-2 in avian immune system.
Zhou
Ji-Yong
JY
Laboratory of Virology and Immunology, Institute of Preventive Veterinary Medicine, Zhejiang University, Hangzhou 310029, China. jyzhou@zju.edu.cn
Wang
Jin-Yong
JY
Chen
Ji-Gang
JG
Wu
Jian-Xiang
JX
Gong
Hui
H
Teng
Qiao-Yang
QY
Guo
Jun-Qing
JQ
Shen
Hui-Gang
HG
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D001681:Biological Assay; D002645:Chickens; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004372:Ducks / Q000235:genetics* / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D007114:Immunization; D066298:In Vitro Techniques; D009980:Influenza A virus / Q000276:immunology; D007252:Influenza Vaccines / Q000276:immunology; D007376:Interleukin-2 / Q000032:analysis / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009500:Neutralization Tests; D010802:Phylogeny; D011817:Rabbits; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D007252:Influenza Vaccines; D007376:Interleukin-2; D011994:Recombinant Proteins