The first cloned echinoderm tumor necrosis factor receptor from Holothuria leucospilota: Molecular characterization and functional analysis.

In this study, an echinoderm tumor necrosis factor receptor named HLTNFR-16 was first cloned from the tropical sea cucumber Holothuria leucospilota. The full-length cDNA of HLTNFR-16 is 3675 bp in size, containing a 415 bp 5'-untranslated region (UTR), a 2024 bp 3'-UTR and a 1236 bp open reading frame (ORF) encoding a protein of 411 amino acids with a deduced molecular weight of 45.63 kDa. The HLTNFR-16 protein contains a signal peptide, four TNFR domains (the last three were identified as extracellular cysteine-rich domains), a transmembrane region and a death domain. Phylogenetic analysis showed that HLTNFR-16 was clustered into a clade with TNFR-16s in other species, indicating that this echinoderm TNFR may be a new member of the TNFR-16 subfamily. The results of TUNEL assay showed that the over expression of HLTNFR-16 could induce apoptosis in HEK293T cells. When HLTNFR-16 was silenced by siRNA, the apoptosis of sea cucumber coelomocytes induced by inactivated Vibrio harveyi was suppressed significantly, indicating that HLTNFR-16 is important for apoptosis induction. Additionally, luciferase reporter assay exhibited that the over-expressed HLTNFR-16 in HEK293T cells could activate the transcription factors nuclear factor-κB (NF-κB) and activator protein-1 (AP-1). Moreover, the secretion of proinflammatory cytokines interleukin (IL)-1β, IL-6 and IL-18 in HEK293T cells was increased by the over-expression of HLTNFR-16. This study provides evidences for the potential roles of sea cucumber TNFR in the innate immunity.