title: Molecular cloning, characterization and expression analysis of PPAR gamma in the orange-spotted grouper (Epinephelus coioides) after the Vibrio alginolyticus challenge.
abstract: PPAR gamma was a key nuclear receptor, playing an important role in the immune defense and the anti-inflammatory mechanism. In this study, the full-length PPAR gamma (EcPPAR gamma) was obtained, containing a 5'UTR of 133 bp, an ORF of 1602 bp and a 3'UTR of 26 bp besides the poly (A) tail. The EcPPAR gamma gene encoded a protein of 533 amino acids with an estimated molecular mass of 60.02 KDa and a predicted isoelectric point (pI) of 6.26. The deduced amino acid sequence showed that EcPPAR gamma consisted of the conserved residues and the domains known to be critical for the PPAR gamma function. The quantitative real-time PCR analysis revealed that EcPPAR gamma transcript was expressed in all the examined tissue, while the strong expression was observed in intestine, followed by the expression in liver, gill, spleen heart, kidney and muscle. Vibrio challenge could stimulate the inflammatory response in grouper and induce a sharp increase of pro-inflammatory cytokines expression, lipid peroxidation and DNA damage, while the up-regulation of vibrio-induced inflammation could also increase the non-specific immune defense. The groupers challenged with Vibrio alginolyticus showed a sharp increase of EcPPAR gamma transcript in immune tissues. Subcellular localization analysis revealed that EcPPAR gamma was distributed in the nucleus. Furthermore, overexpression of EcPPAR gamma could down-regulated the expression of IL1b, IL6, TNF1 and TNF2. In addition, the administration of PPAR gamma antagonist, GW9662, could up-regulate the expression of pro-inflammatory genes, including IL1b, IL6, TNF1 and TNF2. Together, these results indicated that EcPPAR gamma serving as a negative regulator of pro-inflammatory cytokines may play an important role in the immune defense against vibrio-induced inflammation in grouper.
authors.affiliation: Key Laboratory of Ecology and Environmental Science in Guangdong Higher Education, Guangdong Provincial Key Laboratory for Healthy and Safe Aquaculture, College of Life Science, South China Normal University, Guangzhou 510631, PR China.
authors.forename: Shengwei
authors.initials: S
authors.lastname: Luo
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Science, 164 West Xinggang Road, Guangzhou 510301, PR China.
authors.forename: Youhua
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Key Laboratory of Ecology and Environmental Science in Guangdong Higher Education, Guangdong Provincial Key Laboratory for Healthy and Safe Aquaculture, College of Life Science, South China Normal University, Guangzhou 510631, PR China.
authors.forename: Fuxing
authors.initials: F
authors.lastname: Xie
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Science, 164 West Xinggang Road, Guangzhou 510301, PR China.
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Huang
authors.affiliation: Key Laboratory of Ecology and Environmental Science in Guangdong Higher Education, Guangdong Provincial Key Laboratory for Healthy and Safe Aquaculture, College of Life Science, South China Normal University, Guangzhou 510631, PR China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Key Laboratory of Ecology and Environmental Science in Guangdong Higher Education, Guangdong Provincial Key Laboratory for Healthy and Safe Aquaculture, College of Life Science, South China Normal University, Guangzhou 510631, PR China. Electronic address: wangwn@scnu.edu.cn.
authors.forename: Weina
authors.initials: W
authors.lastname: Wang
authors.affiliation: Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Science, 164 West Xinggang Road, Guangzhou 510301, PR China. Electronic address: qinqw@scsio.ac.cn.
authors.forename: Qiwei
authors.initials: Q
authors.lastname: Qin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001492:Bass / Q000145:classification / Q000235:genetics*; D002460:Cell Line; D003001:Cloning, Molecular; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D005393:Fish Diseases / Q000235:genetics* / Q000276:immunology / Q000382:microbiology; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D017665:Hydroxyl Radical / Q000378:metabolism; D008969:Molecular Sequence Data; D009928:Organ Specificity; D047495:PPAR gamma / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D016415:Sequence Alignment / Q000662:veterinary; D013482:Superoxide Dismutase / Q000378:metabolism; D014735:Vibrio Infections / Q000235:genetics / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D044149:Vibrio alginolyticus / Q000502:physiology*
chemical_list: D016207:Cytokines; D018076:DNA, Complementary; D029941:Fish Proteins; D047495:PPAR gamma; D017665:Hydroxyl Radical; D013482:Superoxide Dismutase
keywords: Epinephelus coioides; Gene expression; PPAR gamma; Vibrio alginolyticus