AUTHOR=Zhu Ke-Cheng , Guo Hua-Yang , Guo Liang , Liu Bao-Suo , Zhang Nan , Jiang Shi-Gui , Zhang Dian-Chang 

TITLE=Nuclear Factor-κB Plays a Positive Role in TNFα Expression in Golden Pompano, Trachinotus ovatus (Linnaeus 1758)

JOURNAL=Frontiers in Marine Science

VOLUME=9

YEAR=2022

URL=https://www.frontiersin.org/journals/marine-science/articles/10.3389/fmars.2022.884892

DOI=10.3389/fmars.2022.884892

ISSN=2296-7745

ABSTRACT=<p>Tumour necrosis factor-α (TNFα) is a multifarious mediator of lymphoid tissue growth and antimicrobial defence mechanisms, and it acts as a pro-inflammatory regulator. The function of TNFα in parasite infection and the underlying mechanism through which nuclear factor-κB (NF-κB) regulates TNFα remain largely unclear in teleosts. In the present study, TNFα (<italic>ToTNF</italic>α) from golden pompano (<italic>Trachinotus ovatus</italic>) was identified, and its sequence features and expression levels were determined. The genomic DNA sequence is composed of 1,130 bp, consists of four exons and three introns, and encodes 341 amino acid polypeptides. The putative protein sequence shares 34.7%–61.9% identity with fish TNFα and possesses a TNF family signature and two conserved cysteine residues. Moreover, the expressions of <italic>ToNF-</italic>κ<italic>B</italic> and <italic>ToTNF</italic>α are constitutively expressed in all examined tissues, with higher levels observed in the immune relevant tissues. Both <italic>ToNF-</italic>κ<italic>B</italic> and <italic>ToTNF</italic>α transcription was increased in the local infection sites (skin and gill) and system immune tissues (liver, spleen and head kidney) after <italic>Cryptocaryon irritans</italic> stimulation. In addition, to investigate whether ToNF-κB is a regulator of <italic>ToTNF</italic>α, promoter analysis was performed. The region from -970 to +79 bp is known as the core promoter by different truncated mutants of <italic>ToTNF</italic>α. Subsequently, the activity of the <italic>ToTNF</italic>α<italic>-p2</italic> promoter was dramatically reduced after targeted mutation of the M6-binding site. Additionally, an electrophoretic mobile shift assay (EMSA) verified that ToNF-κB interacted with the M6-binding site in the <italic>ToTNF</italic>α promoter region to control the expression of <italic>ToTNF</italic>α. In conclusion, the present study provides the positive regulation of <italic>TNF</italic>α transcription by NF-κB and contributes to a better understanding of the transcriptional mechanism of <italic>TNF</italic>α in fish.</p>