Molecular cloning of rock bream (Oplegnathus fasciatus) tumor necrosis factor-alpha and its effect on the respiratory burst activity of phagocytes.
Rock bream (Oplegnathus fasciatus) tumor necrosis factor-alpha (rbTNF-alpha) gene was cloned, recombinantly produced, and the effect of the recombinant rbTNF-alpha on the respiratory burst activity of rock bream phagocytes was analyzed. Structurally, genomic DNA of rbTNF-alpha was comprised with four exons and three introns, and deduced amino acid sequence of its cDNA possessed the TNF family signature, a transmembrane domain, a protease cleavage site, and two cysteine residues, which are the typical characteristics of TNF-alpha gene in mammals and fish. The chemiluminescent (CL) response of rock bream phagocytes was significantly enhanced by pre-incubation with recombinant rbTNF-alpha, when opsonized zymosan was used as a stimulant of the respiratory burst. However, CL enhancing effect of the recombinant rbTNF-alpha was very weak when the respiratory burst activity of phagocytes was triggered with phorbol-12-myristate-13-acetate (PMA) instead of zymosan. These results suggest that rock bream TNF-alpha might have an ability to prime the respiratory burst activity of phagocytes against receptor-mediated phagocytosis inducing stimulants, such as zymosan, but have little ability against stimulants not accompanying receptor-mediated phagocytosis.
Kim
Min Sun
MS
Department of Aquatic Life Medicine, Pukyong National University, Nam-gu 599-1, Busan 608-737, South Korea.
Hwang
Yoon Jung
YJ
Yoon
Ki Joon
KJ
Zenke
Kosuke
K
Nam
Yoon Kwon
YK
Kim
Sung Koo
SK
Kim
Ki Hong
KH
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D008969:Molecular Sequence Data; D010473:Perciformes / Q000235:genetics / Q000378:metabolism*; D010586:Phagocytes / Q000187:drug effects / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique / Q000662:veterinary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D016897:Respiratory Burst / Q000187:drug effects / Q000502:physiology*; D016415:Sequence Alignment; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
D012333:RNA, Messenger; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha