利用siRNA高通量测序技术筛查蚊子体内携带的RNA病毒

昆虫可以利用小干扰RNA(siRNA)机制干扰体内RNA病毒的增殖,从而获得对该病毒的免疫能力。通过第二代测序技术对蚊子的小RNA进行高通量测序,再通过生物信息学方法寻找其中的RNA病毒序列,发现在我国云南地区的白纹伊蚊和致倦库蚊体内存在Marayo,Omsk hemorrhagic fever和Ilheus等RNA病毒的基因片段。至此建立了发现媒介昆虫体内携带病毒的一种新方法,可用于媒介昆虫携带RNA病毒的本底调查。     

Down-regulation of free riboflavin content induces hydrogen peroxide and a pathogen defense in Arabidopsis

Riboflavin mediates many bioprocesses associated with the generation of hydrogen peroxide (H?O?), a cellular signal that regulates defense responses in plants. Although plants can synthesize riboflavin, the levels vary widely in different organs and during different stages of development, indicating that changes in riboflavin levels may have physiological effects. Here, we show that changing riboflavin content affects H?O? accumulation and a pathogen defense in Arabidopsis thaliana. Leaf content of free riboflavin was modulated by ectopic expression of the turtle gene encoding riboflavin-binding protein (RfBP). The RfBP-expressing Arabidopsis thaliana (REAT) plants produced the RfBP protein that possessed riboflavin-binding activity. Compared with the wild-type plant, several tested REAT lines had >70% less flavins of free form. This change accompanied an elevation in the level of H?O? and an enhancement of plant resistance to a bacterial pathogen. All the observed REAT characters were eliminated due to RfBP silencing (RfBPi) under REAT background. When an H?O? scavenger was applied, H?O? level declined in all the plants, and REAT no longer exhibited the phenotype of resistance enhancement. However, treatment with an NADPH oxidase inhibitor diminished H?O? content and pathogen defense in wild-type and RfBPi but not in REAT. Our results suggest that the intrinsic down-regulation of free flavins is responsible for NADPH oxidase-independent H?O? accumulation and the pathogen defense.     

An analysis of the influence of protein intrinsic dynamical properties on its thermal unfolding behavior

The influence of the protein topology-encoded dynamical properties on its thermal unfolding motions was studied in the present work. The intrinsic dynamics of protein topology was obtained by the anisotropic network model (ANM). The ANM has been largely used to investigate protein collective functional motions, but it is not well elucidated if this model can also reveal the preferred large-scale motions during protein unfolding. A small protein barnase is used as a typical case study to explore the relationship between protein topology-encoded dynamics and its unfolding motions. Three thermal unfolding simulations at 500 K were performed for barnase and the entire unfolding trajectories were sampled and partitioned into several windows. For each window, the preferred unfolding motions were investigated by essential dynamics analysis, and then associated with the intrinsic dynamical properties of the starting conformation in this window, which is detected by ANM. The results show that only a few slow normal modes imposed by protein structure are sufficient to give a significant overlap with the preferred unfolding motions. Especially, the large amplitude unfolding movements, which imply that the protein jumps out of a local energy basin, can be well described by a single or several ANM slow modes. Besides the global motions, it is also found that the local residual fluctuations encoded in protein structure are highly correlated with those in the protein unfolding process. Furthermore, we also investigated the relationship between protein intrinsic flexibility and its unfolding events. The results show that the intrinsic flexible regions tend to unfold early. Several early unfolding events can be predicted by analysis of protein structural flexibility. These results imply that protein structure-encoded dynamical properties have significant influences on protein unfolding motions.     

Immunohistochemical demonstration of airway epithelial cell markers of guinea pig

The guinea pig (Cavea porcellus) is a mammalian non-rodent species in the Caviidae family. The sensitivity of the respiratory system and the susceptibility to infectious diseases allows the guinea pig to be a useful model for both infectious and non-infectious lung diseases such as asthma and tuberculosis. In this report, we demonstrated for the first time, the major cell types and composition in the guinea pig airway epithelium, using cell type-specific markers by immunohistochemical staining using the commercial available immunological reagents that cross-react with guinea pig. Our results revealed the availability of antibodies cross-reacting with airway epithelial cell types of basal, non-ciliated columnar, ciliated, Clara, goblet and alveolar type II cells, as well as those cells expressing Mucin 5AC, Mucin 2, Aquaporin 4 and Calcitonin Gene Related Peptide. The distribution of these various cell types were quantified in the guinea pig airway by immunohistochemical staining and were comparable with morphometric studies using an electron microscopy assay. Moreover, this study also demonstrated that goblet cells are the main secretory cell type in the guinea pig's airway, distinguishing this species from rats and mice. These results provide useful information for the understanding of airway epithelial cell biology and mechanisms of epithelial–immune integration in guinea pig models.     

非洲猪瘟病毒的非必需基因：真的可有可无吗？

非洲猪瘟(African swine fever,ASF)是由非洲猪瘟病毒(African swine fever virus,ASFV)引起的一种出血性、致死性的猪烈性传染病。ASF在全球广泛传播,给养猪业造成重大的经济损失。ASFV基因组庞大,可编码150多种蛋白,一些非必需基因编码的蛋白与调控病毒毒力、复制和免疫逃逸等相关。通过删除ASFV毒力相关的非必需基因所构建的减毒株是当前比较有前景的疫苗,然而其安全性有待提高。系统地鉴定ASFV非必需基因及其功能,不仅有助于ASF基因缺失疫苗的研发,也有益于ASFV致病机制研究。本文对目前已鉴定的ASFV非必需基因及其功能研究进行了总结分析,着重讨论了影响ASFV毒力、调控病毒复制、参与免疫逃逸的非必需基因及其编码蛋白的功能,旨在加深对ASFV病原学的认识,为新的ASFV非必需基因的鉴定和功能研究提供参考。     

湖南张家界天门山种子植物区系新资料

基于对湖南省张家界市天门山国家森林公园种子植物标本的采集鉴定,报道了湖南省新记录13种2变种,隶属于11科15属。即小山飘风Sedum filipes Hemsl．、绿花石莲Sinocrassula indica（Decne．）Berger var．viridiflora K．T．Fu、扇叶虎耳草Scocifraga rufescens Balf．var．flabellifolia C．Y.Wu et J．T．Pan、滇池海棠Malus yunnanensis（Franch．）Schneid．、北枳棋Hovenia dulcis Thtmb．、橄榄槭Acer olivaceum W．P．Fang et P．L．Chiu、大叶柴胡Bupleurum longiradiaum Turcz．、重齿当归Angelica biserrata（R．H．Shan et C．Q．Yuan）C．Q．Yuan et R．H．Shan、羌活Notopterygium incisum T．T．Ting ex H．T．Chang、直瓣苣苔Ancylostemon sarasilia（Hemsl．）Craib、野丁香Leptodermis potanini Batalin、毛华菊Dendranthema vestitum（Hemsl．）Ling、长冠鼠尾草Salvia pleetranthoides Griff．、茖葱Allium victorialis Linn．、节根黄精Polygonatum nodosum Hua。其中羌活属和野丁香属为湖南分布新记录属。凭证标本均存放在中南林业科技大学树木标本室（CSFI）。     

一色齿毛菌对刚果红的脱色优化及其毒性变化研究

一色齿毛菌Cerrena unicolor是分离自野外的一株能够降解木质素的白腐真菌。为明确一色齿毛菌对染料的脱色能力及脱色前后染料毒性的变化,本研究利用一色齿毛菌对固体条件下4种染料进行脱色能力的检测,筛选出较易脱色的染料后,对该染料的脱色条件进行优化,并以3种豆类发芽率为指标测定该染料脱色前后的毒性变化。结果表明,一色齿毛菌对4种染料均可脱色,其中对刚果红的脱色效果最为明显;一色齿毛菌对刚果红脱色条件的优化结果为：20g/L麦芽糖,1g/L硝酸铵,1mmol/L硫酸镁,接种9块直径1cm菌饼,10mg/L染料浓度,pH 7时脱色效果最好;刚果红染料脱色前后毒性测试结果显示：染料脱色前发酵液毒性>染料脱色后发酵液毒性>清水处理毒性,表明刚果红染料存在一定的毒性,但在被一色齿毛菌脱色后,染料毒性有所降低。本研究为一色齿毛菌在染料废水脱色方面的应用及降低染料废水毒性提供一定的参考依据。     

内皮细胞E选择蛋白的表达及p53基因的影响

为探讨血管内皮细胞Ｅ选择蛋白的表达及野生型ｐ５３ 基因对其表达的影响, 采用流式细胞术及ＲＴＰＣＲ法分别测定其Ｅ选择蛋白及其ｍ ＲＮＡ水平。结果表明： 静息状态的内皮细胞表面检测不到Ｅ选择蛋白的表达, 尽管此时细胞内存在Ｅ选择蛋白的ｍＲＮＡ。肿瘤坏死因子α（ＴＮＦα） 可浓度依赖性地诱导内皮细胞表达Ｅ选择蛋白, 内皮细胞表面Ｅ选择蛋白表达的增加伴随着细胞内Ｅ选择蛋白ｍＲＮＡ 水平的升高。ＴＮＦα刺激内皮细胞后４ ～６ ｈ ,Ｅ选择蛋白的表达达高峰。将ｐ５３ 基因导入内皮细胞后, 经ＴＮＦα诱导的内皮细胞表面Ｅ选择蛋白的表达显著下降, 同时细胞内的Ｅ选择蛋白ｍＲＮＡ 的水平也明显降低。导入ｐ５３ 基因不影响静息状态内皮细胞Ｅ选择蛋白的表达及其ｍ ＲＮＡ 的水平。提示： ｐ５３ 基因至少部分通过降低Ｅ选择蛋白的ｍＲＮＡ 水平而抑制ＴＮＦα诱导的内皮细胞表面Ｅ选择蛋白的表达。     

胰岛素受体底物1的PH结构域与PKC的结合

从大鼠的肝脏克隆胰岛素受体底物１（ＩＲＳ－１）的ＰＨ结构域基因并进行谷胱甘肽Ｓ－转移酶（ＧＳＴ）融合表达,研究该结构域与蛋白激酶Ｃ（ＰＫＣ）的结合情况,并为进一步寻找其新配基打下基础,研究采用一步法从大鼠新鲜肝组织中提取总ＲＮＡ,以ＲＴ－ＰＣＲ的方法扩增目的基因片段,测序证明序列正确,再将正确的目的基因片段定向克隆到表达载体ｐＧＥＸ－４Ｔ－１中,以ＩＰＴＧ在２６℃下诱导,获得与ＧＳＴ的融合表达,表     

Constructing the suitable initial configuration of the membrane-protein system in molecular dynamics simulations

A method for constructing the suitable initial configuration of the membrane-protein system for molecular dynamics (MD) simulations is presented. This method could provide some hydrated initial configurations and help us to determine the best surface area of the system by contracting the surface area and comparing the optimized lowest energy of the system by energy minimization. The gramicidin A (GA) channel in;the fully hydrated dimyristoylphosphatidylcholine (DMPC) bilayer was used as our model. Three configurations with different surface areas were selected and applied for one 400 ps and two 300 ps MD simulations at constant pressure and temperature. All simulations were fairly stable without any constraints. Through analysis of the MD trajectories we found that the system with the best surface area was more stable than the other two systems, whose sizes were changed in the simulations. Further analysis of the bilayer normal length and the order parameters of the lipid alkyl tails indicates that the system with the best surface area shows some characteristics of the L_α phase, while both the smaller and the larger size systems have distinct deviations from the L_α phase that we expect. This illustrates that the correct surface area and the suitable initial configuration have an important influence on the phase of the membrane in the MD simulation. In addition, by comparing the root mean square differences of GA relative to the initial structure and interaction energy between different components of the system for all three systems, we find that the state of the DMPC bilayer has exerted a significant influence on the structure of GA. All these results demonstrate the validity of our method for constructing the initial configuration of the membrane-protein system for MD simulations. Received: 10 September 1998 / Revised version: 19 March 1999 / Accepted: 19 March 1999