Isozyme Analysis of Spotted Halibut Verasper variegatus Temminck et Schlegel

To investigate the tissue-specificities of isozymes and the genetic structureof wild spotted halibut ( Verasper variegatus) population, horizontal starch gel electrophoresiswas performed on 45 individuals collected in part of the Yellow Sea. The performances of 17 isozymesin 8 kinds of tissues or organs were screened preliminarily in a TC-7.0 buffer system. The resultsshowed that the screened isozymes displayed remarkable tissue-specificities. Finally, 14 enzymes(AAT, ADH, EST, OPI, G3PDH, IDHP, LAP, LDH, MDH, MPI, PGDH, PGM, SDH and SOD) and 4 kinds of tissues(eye, skeleton muscle, liver and heart) were selected for genetic analysis. Fourteen isozymes areencoded by 20 loci, and 9 of them are polymorphic. The polymorphic loci are AAT-1~*, GPI-2~*,G3PDH~*, IDHP-1~*, LDH~*, MPI~*, PGM-1~*. PGM-2~* and SDH~*, and the proportion of polymorphic lociis 0.4500 (P_(0.99)) ? The mean values of observed and expected heterozygosities are 0.0278and 0.0265, respectively and the average effective number of alleles is 1.0675.

Tumor necrosis factor-alpha(TNF-α) in spotted halibut Verasper variegatus at the embryonic and metamorphic stages

As an aquatic fish,the spotted halibut Verasper variegatus is highly susceptible to bacterial and virus infections.Tumor necrosis factor-alpha(TNF-α)as a cytokine could control the inflammatory responses.The functions of TNF-αin many species have been widely studied,particularly in mammals.However,little is known about the TNF-αfunctions in V.variegatus.We first cloned and sequenced the TNF-αgene in V.variegatus(VvTNF-α).The two conserved cysteine residues,transmembrane sequence,Thr-Leu motif,and TNF family signature,as well as the TA-rich motifs of its proteins related to inflammatory responses had high similarity to those of the other teleost and mammalian TNF-α.The phylogenetic analysis showed that VvTNF-αwas consistent with TNF-αgenes of other vertebrates.The VvTNF-αtranscripts were extensively distributed in the peripheral blood leukocytes(PBLs),spleen,and gill,indicating that the VvTNF-αhad a role in immune function.Furthermore,treatment with pathogen-associated molecular patterns(PAMPs)could induce a rapid and significant increase of VvTNF-αin the PBLs,which reveals that VvTNF-αdoes participate in the host immune responses against bacterial and viral pathogens.We found that VvTNF-αhad an interesting expression pattern during metamorphosis,showing that the flatfish TNF-αmay have some novel functions during specific developmental stages.In addition,the 3 D structure prediction of VvTNF-αprovided an indication of how it is likely to interact with other proteins.Therefore,VvTNF-αhas multiple functions,and provides valuable information to explore novel functions of TNF-α.

Novel Functions of Interleukin-1 Beta in Spotted Halibut(Verasper variegatus) at Its Embryonic and Metamorphic Stages

Interleukin-lbeta （IL-1β） is a cytokine involved in the control of inflammatory responses. The functions of IL-1β have been extensively studied in many vertebrates. However, few studies focused on the functions of IL-1β in the spotted halibut （Verasper variegatus）, an endangered aquatic fish that easily suffers from bacterial and viral infections. We first cloned and sequenced the IL-1β gene from V. variegatus （VvIL-1β）. Its conservative interleukin-1 propeptide and interleukin-1/18 domains corresponding regions, characteristics of the IL-1 protein family and TA-rich motifs relating to inflammatory responses were similar to those of the IL-1β genes from other teleost and mammals. The phylogenetic analysis showed that VvIL-1β was homologous to the IL-1β gene of other vertebrates. Treatment with pathogen-associated molecular patterns （PAMPs） induced a significant and rapid up-regulation of VvIL-1β expression in the peripheral blood leukocytes （PBLs）, revealing that VvIL-1β is actively involved in the host ilmnune responses against bacterial and viral pathogens. The expression level of VvIL-1β was relatively high in the PBLs, gill, and intestine, indicating the immunological function of VvIL-1β . VvIL-1β also had interesting expression patterns at specific developmental stages, implying that the IL-1β of flatfishes may have some novel functions during embryonic development and metamorphosis. The 3D structure prediction also provided information about how VvIL-113 interacts with its receptor protein. These results suggested that VvIL-1β may have multiple functions in V. variegatus and provided us valuable information for understanding the novel functions of IL- 1β.