돌돔 (Oplegnathus fasciatus) tumor necrosis factor receptor -associated factor 2 클로닝 및 NF-κB 활성화에 미치는 영향

Abstract

Molecular cloning of rock bream’s (Oplegnathus fasciatus) tumor necrosis factor receptor-associated factor 2 and its function on NF-κB activation. Woo Li Kim Department of fish pathology, The Graduate School, Pukyong National University Abstract Tumor necrosis factor receptor-associated factor 2 (TRAF2) acts as a transducer of tumor necrosis factor-α (TNF-α) triggered cell signals which results in inflammation, cell proliferation and anti-apoptotic response. In this study, we have cloned cDNA of rock bream (Oplegnathus fasciatus) TRAF2, and analyzed its function in activation of NF-κB. The open reading frame (ORF) of rock bream TRAF2 consisted of 1563 bp encoding 520 amino acids that contained N-terminal RING-type and TRAF-type zinc finger domains and C-terminal TRAF domain. The rock bream TRAF2 protein showed more than 75% identity with other fish TRAF2s, and even as high as 56% identity with mouse and human TRAF2 proteins. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed that rock bream TRAF2 was expressed in all tested tissues of unstimulated fish, and, interestingly, tissues of spleen, gill, leukocytes, intestine, and skin showed higher expression than other tissues. TRAF2 is essential for TNF-α-induced NF-κB activation. To know whether the rock bream TRAF2 involves in NF-κB activation, Epithelioma papulosum cyprini (EPC) cells harboring an NF-κB reporting vector were transfected with a vector expressing rock bream TRAF2 or a control empty vector. EPC cells transfected with the vector expressing rock bream TRAF2 showed significantly higher NF-κB activity than cells transfected with a control empty vector, suggesting rock bream TRAF2 acts as a transducer of TNF-α-mediated cell signals that enhance NF-κB activation.