Institute of Hydrobiology, Chinese Academy Of Sciences

    Mga Modulates Bmpr1a Activity by Antagonizing Bs69 in Zebrafish

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    MAX giant associated protein (MGA) is a dual transcriptional factor containing both T-box and bHLHzip DNA binding domains. In vitro studies have shown that MGA functions as a transcriptional repressor or activator to regulate transcription of promotors containing either E-box or T-box binding sites. BS69 (ZMYND11), a multidomain-containing (i.e., PHD, BROMO, PWWP, and MYND) protein, has been shown to selectively recognizes histone variant H3.3 lysine 36 trimethylation (H3.3K36me3), modulates RNA Polymerase II elongation, and functions as RNA splicing regulator. Mutations in MGA or BS69 have been linked to multiple cancers or neural developmental disorders. Here, by TALEN and CRISPR/Cas9-mediated loss of gene function assays, we show that zebrafish Mga and Bs69 are required to maintain proper Bmp signaling during early embryogenesis. We found that Mga protein localized in the cytoplasm modulates Bmpr1a activity by physical association with Zmynd11/Bs69. The Mynd domain of Bs69 specifically binds the kinase domain of Bmpr1a and interferes with its phosphorylation and activation of Smad1/5/8. Mga acts to antagonize Bs69 and facilitate the Bmp signaling pathway by disrupting the Bs69-Bmpr1a association. Functionally, Bmp signaling under control of Mga and Bs69 is required for properly specifying the ventral tailfin cell fate.</p

    Rana grylio virus 43R encodes an envelope protein involved in virus entry

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    Rana grylio virus (RGV), a member of genus Ranavirus in the family Iridoviridae, is a viral pathogen infecting aquatic animal. RGV 43R has homologues only in Ranavirus and contains a transmembrane (TM) domain, but its role in RGV infection is unknown. In this study, 43R was determined to be associated with virion membrane. The transcripts encoding 43R and the protein itself appeared late in RGV-infected EPC cells and its expression was blocked by viral DNA replication inhibitor, indicating that 43R is a late expressed protein. Subcellular localization showed that 43R-EGFP fusion protein distributed in cytoplasm of EPC cells and that TM domain is essential for its distribution in cytoplasm. 43R-EGFP fusion protein colocalized with viral factories in RGV-infected cells. A recombinant RGV deleting 43R (43R-RGV) was constructed by homologous recombination to investigate its role in virus infection. Compared with wild type RGV, the ability of 43R-RGV to induce the cytopathic effect and its virus titers were significantly reduced. Furthermore, it is revealed that 43R deletion significantly inhibited viral entry but did not influence viral DNA replication by measuring and comparing the DNA levels of RGV and 43R-RGV in the infected cells at the early stage of infection. RGV neutralization with anti-43R serum reduced the virus titer. Therefore, these data showed that RGV 43R is a late gene that encodes an envelope protein involved in RGV entry.</p

    Effects of Salt Concentrations and Nitrogen and Phosphorus Starvations on Neutral Lipid Contents in the Green Microalga Dunaliella tertiolecta

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    Dunaliella tertiolecta, a halotolerant alga, can accumulate large amounts of neutral lipid, which makes it a potential biodiesel feedstock. In this study, neutral lipids of D. tertiolecta induced by different salinities or N or P starvation were analyzed by thin-layer chromatography (TLC), flow cytometry (FCM), and confocal laser scanning microscopy (CLSM). High salinities or N or P starvation resulted in a decrease in cell growth and chlorophyll contents of D. tertiolecta. Neutral lipid contents increased markedly after 3-7 days of N starvation or at low NaCl concentrations (0.5-2.0 M). N starvation had a more dramatic effect on the neutral lipid contents of D. tertiolecta than P starvation. Four putative ME isozymes in different conditions can be detected by using isozyme electrophoresis. Two alternative acetyl-CoA producers, ACL and ACS genes, were up-regulated under low salinities and N starvation. It was suggested that low salinities and N starvation are considered efficient ways to stimulate lipid accumulation in D. tertiolecta.</p

    Zebrafish foxo3b Negatively Regulates Antiviral Response through Suppressing the Transactivity of irf3 and irf7

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    Forkhead box O (FOXO)3, a member of the FOXO family of transcription factors, plays key roles in various cellular processes, including development, longevity, reproduction, and metabolism. Recently, FOXO3 has also been shown to be involved in modulating the immune response. However, how FOXO3 regulates immunity and the underlying mechanisms are still largely unknown. In this study, we show that zebrafish (Danio rerio) foxo3b, an ortholog of mammalian FOXO3, is induced by polyinosinic-polycytidylic acid stimulation and spring viremia of carp virus (SVCV) infection. We found that foxo3b interacted with irf3 and irf7 to inhibit ifr3/irf7 transcriptional activity, thus resulting in suppression of SVCV or polyinosinic-polycytidylic acid-induced IFN activation. By suppressing expression of key antiviral genes, foxo3b negatively regulated the cellular antiviral response. Furthermore, upon SVCV infection, the expression of the key antiviral genes was significantly enhanced in foxo3b-null zebrafish larvae compared with wild-type larvae. Additionally, the replication of SVCV was inhibited in foxo3b-null zebrafish larvae, leading to a higher survival rate. Our findings suggest that by suppressing irf3/irf7 activity, zebrafish foxo3b negatively regulates the antiviral response, implicating the vital role of the FOXO gene family in innate immunity.</p

    Divergent DNA Methylation Provides Insights into the Evolution of Duplicate Genes in Zebrafish

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    The evolutionary mechanism, fate and function of duplicate genes in various taxa have been widely studied; however, the mechanism underlying the maintenance and divergence of duplicate genes in Danio rerio remains largely unexplored. Whether and how the divergence of DNA methylation between duplicate pairs is associated with gene expression and evolutionary time are poorly understood. In this study, by analyzing bisulfite sequencing (BS-seq) and RNA-seq datasets from public data, we demonstrated that DNA methylation played a critical role in duplicate gene evolution in zebrafish. Initially, we found promoter methylation of duplicate genes generally decreased with evolutionary time as measured by synonymous substitution rate between paralogous duplicates (Ks). Importantly, promoter methylation of duplicate genes was negatively correlated with gene expression. Interestingly, for 665 duplicate gene pairs, one gene was consistently promoter methylated, while the other was unmethylated across nine different datasets we studied. Moreover, one motif enriched in promoter methylated duplicate genes tended to be bound by the transcription repression factor FOXD3, whereas a motif enriched in the promoter unmethylated sequences interacted with the transcription activator Sp1, indicating a complex interaction between the genomic environment and epigenome. Besides, body-methylated genes showed longer length than body-unmethylated genes. Overall, our results suggest that DNA methylation is highly important in the differential expression and evolution of duplicate genes in zebrafish.</p

    欧洲鳗鲡淋巴细胞转化试验条件的建立

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    通过对培养时间、培养温度、刀豆蛋白A(ConA)质量浓度和鳗鲡血清质量分数4个参数的测定,确定欧洲鳗鲡(Anguilla anguilla)外周血淋巴细胞转化试验四甲基偶氮唑(MTT)法的实验条件。结果表明,在培养时间为42-90 h,培养温度分别为15.0℃、20.0℃、25.0℃和30.0℃时,细胞培养液中ConA质量浓度分别为0、10&mu;g/mL、20&mu;g/mL和30&mu;g/mL,鳗鲡血清质量分数分别为0、5%、10%和15%时,淋巴细胞于20.0℃、含10&mu;g/mL ConA,10%鳗鲡血清的RPM

    湘中山地土壤线虫(Nematoda)生态地理群的研究

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    作者对湖南南岳衡山和长沙岳麓山土壤线虫进行了两年多的定位研究,共获得18511条土壤线虫标本.经分类鉴定,计有24个种,分属6目10科18届

    Construction and Characterization of Two Bacterial Artificial Chromosome Libraries of Grass Carp

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    Bacterial artificial chromosome (BAC) library is an important tool in genomic research. We constructed two libraries from the genomic DNA of grass carp (Ctenopharyngodon idellus) as a crucial part of the grass carp genome project. The libraries were constructed in the EcoRI and HindIII sites of the vector CopyControl pCC1BAC. The EcoRI library comprised 53,000 positive clones, and approximately 99.94% of the clones contained grass carp nuclear DNA inserts (average size, 139.7 kb) covering 7.4x haploid genome equivalents and 2% empty clones. Similarly, the HindIII library comprised 52,216 clones with approximately 99.82% probability of finding any genomic fragments containing single-copy genes; the average insert size was 121.5 kb with 2.8% insert-empty clones, thus providing genome coverage of 6.3x haploid genome equivalents of grass carp. We selected gene-specific probes for screening the target gene clones in the HindIII library. In all, we obtained 31 positive clones, which were identified for every gene, with an average of 6.2 BAC clones per gene probe. Thus, we succeeded in constructing the desired BAC libraries, which should provide an important foundation for future physical mapping and whole-genome sequencing in grass carp

    Microbial diversity of intestinal contents and mucus in yellow catfish (Pelteobagrus fulvidraco)

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    In this study, traditional culture-based techniques and the 16S rDNA sequencing method were used to investigate the microbial community of the intestinal contents and mucosal layer in the intestine of yellow catfish (Pelteobagrus fulvidraco). Eleven phylotypes were detected from culturable microbiota, and their closest relatives were Plesiomonas, Yersinia, Enterobacter, Shewanella, Aeromonas, Vibrio, and Myroides. Forty-four phylotypes were retrieved from 100 positive clones from intestinal contents (library C), and 21 phylotypes were detected in the 57 positive clones from intestinal mucus (library M), most of which were affiliated with Proteobacteria (>50% of the total). However, the bacterial groups OP10 and Actinobacteria detected in library C were not found in library M, suggesting that the abundance and diversity of bacterial populations in mucus might be different from the microbiota in gut contents, and that some microbial species poorly colonized the gut mucosal layer. (C) 2010 Published by Elsevier B.V

    A niche model to predict Microcystis bloom decline in Chaohu Lake, China

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    Cyanobacterial blooms occur frequently in lakes due to eutrophication. Although a number of models have been proposed to forecast algal blooms, a good and applicable method is still lacking. This study explored a simple and effective mathematical-ecological model to evaluate the growth status and predict the population dynamics of Microcystis blooms. In this study, phytoplankton were collected and identified from 8 sampling sites in Chaohu Lake every month from July to October, 2010. The niche breadth and niche overlap of common species were calculated using standard equations, and the potential relative growth rates of Microcystis were calculated as a weighted-value of niche overlap. In July, the potential relative growth rate was 2.79 (a.u., arbitrary units) but then rapidly declined in the following months to -3.99 a.u. in September. A significant correlation (R =0.998, P < 0.01) was found in the model between the net-increase in biomass of Microcystis in the field and the predicted values calculated by the niche model, we concluded that the niche model is suitable for forecasting the dynamics of Microcystis blooms. Redundancy analysis indicated that decreases in water temperature, dissolved oxygen and total dissolved phosphorus might be major factors underlying bloom decline. Based on the theory of community succession being caused by resource competition, the growth and decline of blooms can be predicted from a community structure. This may provide a basis for early warning and control of algal blooms.Cyanobacterial blooms occur frequently in lakes due to eutrophication. Although a number of models have been proposed to forecast algal blooms, a good and applicable method is still lacking. This study explored a simple and effective mathematical-ecological model to evaluate the growth status and predict the population dynamics of Microcystis blooms. In this study, phytoplankton were collected and identified from 8 sampling sites in Chaohu Lake every month from July to October, 2010. The niche breadth and niche overlap of common species were calculated using standard equations, and the potential relative growth rates of Microcystis were calculated as a weighted-value of niche overlap. In July, the potential relative growth rate was 2.79 (a.u., arbitrary units) but then rapidly declined in the following months to -3.99 a.u. in September. A significant correlation (R =0.998, P < 0.01) was found in the model between the net-increase in biomass of Microcystis in the field and the predicted values calculated by the niche model, we concluded that the niche model is suitable for forecasting the dynamics of Microcystis blooms. Redundancy analysis indicated that decreases in water temperature, dissolved oxygen and total dissolved phosphorus might be major factors underlying bloom decline. Based on the theory of community succession being caused by resource competition, the growth and decline of blooms can be predicted from a community structure. This may provide a basis for early warning and control of algal blooms
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