title: Characterization and expression analysis of Calmodulin (CaM) in orange-spotted grouper (Epinephelus coioides) in response to Vibrio alginolyticus challenge.
abstract: Vibrio alginolyticus containing the highly toxic extracellular product is one of the most serious threats to grouper survival and its minimum lethal dose is approximately 500 CFU/g fish body weight in grouper. To study the toxic effects of V. alginolyticus on the immune system in teleost, Calmodulin (CaM), an important molecular indicator gene, was cloned from the orange-spotted grouper (Epinephelus coioides). The full-length Ec-CaM consisted of a 5'-UTR of 103 bp, an ORF of 450 bp and a 3'-UTR of 104 bp. The Ec-CaM gene encoded a protein of 149 amino acids with an estimated molecular mass of 16.4 kDa and a predicted isoelectric point of 3.93. The deduced amino acid sequence showed that Ec-CaM contained four highly conserved EF-hand domains known to be critical for the function of CaM. Ec-CaM was widely expressed and the highest expression level was observed in liver. Following V. alginolyticus challenge, a sharp increase level of respiratory burst activity and apoptosis ratio were observed. Further analyses of CaM expression and p53 expression in liver, kidney and spleen by qRT-PCR demonstrated that the up-regulated expression of CaM and p53 were observed in the vibrio challenge group. Western blotting analysis confirmed that the Ec-CaM protein was strongly induced in liver at 12 h post-injection, while a sharp increase of p53 protein expression was observed at 24 h post-injection. These results showed CaM expression serving as a potential molecular indicator may help to assess the toxicological effects of V. alginolyticus on the ROS generation and apoptotic process 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, People's Republic of China.
authors.forename: Sheng-Wei
authors.initials: SW
authors.lastname: Luo
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, People's Republic of China.
authors.forename: Fu-Xing
authors.initials: FX
authors.lastname: Xie
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, People's Republic of 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, People's Republic of China. Weina63@aliyun.com.
authors.forename: Wei-Na
authors.initials: WN
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D001492:Bass; D002147:Calmodulin / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D005393:Fish Diseases / Q000235:genetics* / Q000276:immunology / Q000382:microbiology; D029941:Fish Proteins / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D009928:Organ Specificity; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016897:Respiratory Burst; D016415:Sequence Alignment; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D014735:Vibrio Infections / Q000235:genetics / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D044149:Vibrio alginolyticus / Q000502:physiology*
chemical_list: D002147:Calmodulin; D018076:DNA, Complementary; D029941:Fish Proteins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53
keywords: Calmodulin; Epinephelus coioides; Gene expression; Vibrio alginolyticus