title: Molecular and immune response characterizations of IL-6 in large yellow croaker (Larimichthys crocea).
abstract: Interleukin-6 (IL-6) is a multifunctional inflammatory cytokine which exists in multiple tissues and cell lines. In the present study, the full-length cDNA and the genomic sequence of IL-6 (LcIL-6) were cloned from large yellow croaker, Larimichthys crocea. The full-length cDNA of LcIL-6 was 1066 base pairs (bp), containing an open reading frame (ORF) of 678 bp encoding for 225 amino acids, a 5' untranslated region (UTR) of 71 bp and a 3' UTR of 317 bp. The predicted LcIL-6 protein included a 24 amino acids (aa) signal peptide and a conserved IL-6 domain. However, the polypeptide sequence identities between LcIL-6 and its counterparts in mammals and other fish are from 12% to 45%. The genome sequence of LcIL-6 gene was composed of 2126 bp, including five exons and four introns. Phylogenetic analysis revealed that LcIL-6 showed a close relationship with the IL-6 from other bony fish. Quantitative real-time PCR (qRT-PCR) analysis revealed that LcIL-6 mRNA was expressed in most examined tissues, with the most predominant expression in stomach, followed by blood and very weak expression in other tissues. The expression levels of LcIL-6 after challenged with LPS, poly I:C and Vibrio parahaemolyticus were investigated in spleen, head-kidney and liver. LcIL-6 transcripts were induced significantly after immune challenge, with the peak-value of 33.5 times as much as the control in the head-kidney at 3 h after LPS injection (p < 0.05). Overexpression of LcIL-6 enhanced tumor necrosis factor (TNF)-α transcripts significantly (p < 0.05) in L. crocea kidney (LCK) cells. Additionally, recombinant LcIL-6 mature peptide was obtained in the supernatant of Escherichia coli BL21 (DE3). The purified recombinant LcIL-6 fusion protein was also demonstrated to improve the transcriptional expression levels of TNF-α significantly in LCK cells (p < 0.05). However, no significant changes of Mx (myxovirus resistant protein), IL-1β, janus kinase (JAK)2, signal transducers and activators of transcription (STAT)3 and STAT5 in LCK cells was detected after LcIL-6 overexpression or recombinant LcIL-6 protein stimulation. Our results indicated that LcIL-6 might be important in large yellow croaker immune response and improve the inflammatory response by through activation TNF-α expression.
authors: Fisheries College/Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Jimei University, Xiamen 361021, PR China.
authors: Qian
authors: Q
authors: Zhu
authors: Fisheries College/Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Jimei University, Xiamen 361021, PR China.
authors: Chan
authors: C
authors: Li
authors: Fisheries College/Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Jimei University, Xiamen 361021, PR China.
authors: Zhen-Xing
authors: ZX
authors: Yu
authors: Fisheries College/Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Jimei University, Xiamen 361021, PR China.
authors: Peng-Fei
authors: PF
authors: Zou
authors: Fisheries College/Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Jimei University, Xiamen 361021, PR China.
authors: Qing-Xiang
authors: QX
authors: Meng
authors: Fisheries College/Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Jimei University, Xiamen 361021, PR China. Electronic address: clyao@jmu.edu.cn.
authors: Cui-Luan
authors: CL
authors: Yao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007113:Immunity, Innate; D015850:Interleukin-6 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D009928:Organ Specificity; D010473:Perciformes / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D010802:Phylogeny; D011070:Poly I-C / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment / Q000662:veterinary; D014736:Vibrio parahaemolyticus / Q000502:physiology
chemical_list: D018076:DNA, Complementary; D029941:Fish Proteins; D015850:Interleukin-6; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D011070:Poly I-C
keywords: Gene structure; Immune challenge; Inflammatory response; Interleukin-6; Large yellow croaker; Overexpression; Recombinant expression