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《第十三届全国免疫学学术大会摘要汇编》2018年
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Structure and function of CXCL8 in Northeast Chinese lamprey(Lethenteron morii)

QINGHUA ZHANG  XINYUN ZHU  ZHE ZHANG  JIANFENG REN  WEIMING LI  
【摘要】:Chemokine induced chemotaxis of leukocytes is an important part of the innate immunity, and has been shown to mediate inflammation in all groups of jawed vertebrates.For jawless vertebrates, hagfish leukocytes are known to show chemotaxis toward mammalian complement anaphylotoxin and Gram negative bacteria lipopolysaccharide(LPS).However, whether chemokines mediate chemotaxis in jawless vertebrates have not been conclusively examined.To elucidate the function of CXCL8 in sea lamprey innate immunity, we cloned LmCXCL8 by PCR from Northeast Chinese lamprey(Lethenteron morii).Next, the recombinant PmCXCL8 protein was purified by Bio-Rad IMAC Cartridge and Profinia Protein Purification Instrument, and analyzed by SDS-PAGE and Western blot.At last, we test the chemotaxis of recombinant LmCXCL8 to the lamprey peripheral blood leucocytes(PBLs).Our results showed that the full-length cDNA of LmCXCL8 consisted of 1055 nucleotides(nt), including a 5'-untranslated region(UTR) of 142 nt and a 3'-UTR of 604 nt, and an open reading frame(ORF) of 309 bp encoded a polypeptide of 102 amino acid residues with a predicted molecular weight of 11.23 kDa.It also contained four cysteine residues(Cys-36, 38, 63 and 80), which were characteristic of CXC subfamily cytokines and found in all vertebrates.The identified LmCXCL8 possessed the characteristic N-terminal cysteine residues and GGR(Gly-Gly-Arg) motif.The Lmcxcl8 gene was expressed in all examined tissues of the juvenile and adult lamprey, and the expression was further inducible by challenge of an infectious bacterium, Pseudomonas aeruginosa.A recombinant LmCXCL8 protein elicited concentration dependent chemotaxis in peripheral blood leucocytes(PBLs) isolated from the Northeast Chinese lamprey.Based on these results, we confirm that the LmCXCL8 is a constitutive and inducible acute-phase cytokine that mediates immune defense, and trace the chemotactic function of chemokine to basal vertebrates.

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