Molecular identification and functional characterization of a tumor necrosis factor (TNF) gene in Crassostrea hongkongensis

Tumor necrosis factor superfamily (TNFSF) represents a group of multifunctional inflammatory cytokines that have been shown to participate in a variety of pathological and immunological process. However, the functions of these proteins in oyster are still poorly understood. In the present study, an oyster TNF homolog (named ChTNF) was identified from a cDNA library of Crassostrea hongkongensis. The complete cDNA of ChTNF was 2457 bp in length containing an open reading frame (ORF) of 1044 bp, a 5′-untranslated region (UTR) of 381 bp and a 3′-UTR of 1032 bp. The deduced ChTNF protein consisted of 347 amino acids with a characteristic transmembrane domain and a typical TNF homology domain (THD). Quantitative real-time PCR analysis revealed that ChTNF was broadly expressed in various oyster tissues and different stages of embryonic development. In addition, transcriptional analysis indicated that ChTNF transcription levels in hemocytes were increased significantly in pathogen challenge groups (Vibrio alginolyticus and Staphylococcus haemolyticus) compared to that in the control. Moreover, in vitro PAMP (lipopolysaccharide and peptidoglycan) treatments showed a stimulatory effect on the expression of ChTNF in the primary cultured hemocytes of C. hongkongensis. Finally, dual-luciferase reporter assays revealed that ChTNF could activate the NF-κB-Luc reporter in a dose-dependent manner in HEK293T cells. Altogether, these findings may provide valuable information regarding oyster TNFs and its role in innate immunity.