昆虫谷胱甘肽S-转移酶的多样性及其介导的抗药性

谷胱甘肽S-转移酶(GSTs)是一类广泛分布于生物体的多功能解毒酶系,参与许多内外源有毒物质的代谢。昆虫GSTs目前主要分为6个已知亚族,其中Delta和Epsion是昆虫特异的亚族,已鉴定的抗性相关基因主要分属于这两个亚族。作为重要的解毒酶,它主要参与昆虫对有机磷、拟除虫菊酯和有机氯等杀虫剂的抗性形成。本文主要对昆虫细胞质GSTs的分类、基因多样性及其在抗药性中的作用等相关研究进展进行综述。     

杨树WRKY基因家族鉴定及其干旱胁迫响应模式分析

WRKY是近年来研究较广泛的植物转录因子,其序列的氨基(N)末端含有高度保守的七肽WRKYGQK,能够与顺式作用元件W盒[(T)(T)TGAC(C/T)]发生特异性结合,从而调控下游靶基因的表达。该研究利用生物信息学方法,对杨树WRKY转录因子进行了序列鉴定、结构分析、染色体定位、结构域分析、系统进化分析和胁迫响应模式分析,鉴定出杨树WRKY基因家族有122个成员,分为3大类(34个Ⅰ类成员、78个Ⅱ类成员和10个Ⅲ类成员); WRKY基因染色体定位分析发现,位于每条染色体的数目各不相同,并且在9号染色体上没有分布,表明WRKY基因在染色体上呈现出不均匀分布;系统发育分析表明, WRKY家族成员形成3个主要进化支,此外,结合基因结构分析发现,位于相同进化支的WRKY基因通常具有相似的基序组成和外显子/内含子结构模式,表明WRKY基因的功能相似性;转录组分析发现, 62个家族成员表现出上调或下调的差异表达,可将其划分为5个基因聚类(Ⅰ、Ⅱ、Ⅲ、Ⅳ、Ⅴ),其中Ⅰ、Ⅲ、Ⅳ、Ⅴ这4个聚类中绝大多数基因在干旱胁迫8 h左右表现明显上调, Ⅱ类在干旱胁迫2～4 h表现明显上调,而PtWRKY70、PtWRKY81、PtWRKY104、PtWRKY108等在干旱胁迫处理后开始明显下调,表明WRKY基因在响应干旱胁迫的过程中具有重要作用。通过上述研究,极大丰富了杨树WRKY基因家族以及其应对干旱胁迫的功能,并为杨树抗旱育种计划提供了候选基因。     

昆虫启动子的研究及应用

启动子(promotor)作为基因时空表达调控的重要元件,与RNA聚合酶结合形成转录起始复合体,控制着基因表达的起始时间和表达水平。对启动子的研究可以更好地阐明基因表达调控的机制,为解析基因调控网络奠定基础。本文对昆虫启动子研究方法进行综述,主要阐述昆虫启动子及其特定元件的鉴定方法,以及昆虫启动子的应用。     

昆虫抗药性相关细胞色素P450基因的表达调控机制

杀虫剂的频繁持续使用,必然导致昆虫产生抗药性。大量研究事例表明参与杀虫剂解毒的细胞色素P450(简称P450)过量表达是昆虫对不同类型杀虫剂产生抗性的重要原因,但目前人们对P450基因过量表达机制的认识还非常有限。近十年来,随着生命科学与相关研究技术的发展,有关昆虫P450基因表达调控机制的研究取得了实质性的进展。本文综述了这一研究领域的重要发现。除了基因重复或基因扩增导致的P450基因拷贝数增加外,P450基因在转录层面的上调表达是P450介导抗药性的普遍且重要的机制。P450基因的转录上调由顺式调控元件与反式作用因子相互作用得以实现。现已发现了几种不同类型的转录因子(CncC, CREB和核受体等)对昆虫P450表达的直接调控,也鉴定了间接调控P450表达的作用因子如G蛋白偶联受体及其下游效应子。ncC:Maf/Keap1是抗药性相关P450基因表达的重要而普遍的调控途径。越来越多的事例表明小RNA在昆虫P450的表达调控中起重要作用。现有的研究结果揭示了昆虫P450基因调控因子和信号转导通路的多样性及调控机制的复杂性。     

白癜风的HLA等位基因遗传学研究进展

白癜风的病因尚不明,国内外研究表明其发生与遗传因素相关,其遗传特征不符合孟德尔遗传规律,而是属于多基因遗传的范畴.主要与人类白细胞抗原HLA-Ⅰ类、Ⅱ类及Ⅲ类基因及其产物相关,以往文献报道认为HLA-Ⅱ类基因与白癜风的关系最为密切.HLA基因的多态性决定HLA分子的多样性.充分认识HLA基因水平的多态性,有助于白癜风的基因诊断和治疗.     

昆虫细胞色素P450基因的多样性、进化及表达调控

细胞色素P450单加氧酶（cytochrome P450 monooxygenases, P450s）是由多个功能相关的亚铁血红素 硫醇盐蛋白基因组成的一个基因超家族, 在各种内源和外源物质的代谢中起着主要作用。目前GenBank中注册的昆虫P450基因序列已超过1 000个, 其中双翅目占序列总数的74%, 鳞翅目占序列总数的16%。而昆虫P450基因序列已克隆的全长序列中大部分属于CYP4和CYP6家族, 两个家族成员分别占总数的20%和45%。利用GenBank中现已注册的昆虫P450基因的cDNA全长序列进行比对并绘制进化树, 揭示不同种类昆虫P450的亲缘关系。结果显示基于P450基因的昆虫部分目的进化关系与大部分先前依据其他分子数据或形态分类学得到的昆虫系统进化关系基本吻合。现有研究表明, 细胞色素P450基因的表达可能受顺式作用元件(cis-acting element)、反式作用因子(trans-acting factor)或两者共同调控, 调控可能涉及转录增强的转录机制或mRNA稳定性增加的转录后机制。     

桃WRKY基因家族全基因组鉴定和表达分析

WRKY转录因子是植物中最大的转录调控家族之一,在生物和非生物胁迫以及植物生长和发育过程中起着重要调控作用.本文利用HMMER 3.0软件,使用WRKY保守域全蛋白序列(Pfam数据库编号:PF03106)鉴定桃(Prunus persica L.)基因组中的WRKY基因;利用DNAMAN 5.0,WebLogo 3,MEGA5.1,MapInspect和MEME等软件对其蛋白序列进行生物信息学分析.本文共鉴定得到61个桃WRKY基因.进化树分析结果显示,桃WRKY蛋白分为Ⅰ,Ⅱ和Ⅲ类型,类型Ⅰ分为Ⅰ-C亚组和Ⅰ-N亚组,类型Ⅱ分为Ⅱ-a,II-b,II-c,II-d和II-e亚组.WRKY结构域分析显示,WRKY结构域高度保守,绝大多数都含有WRKYGQK七肽和锌指结构.染色体定位分析显示,桃WRKY基因分布于8条染色体中,呈不均匀分布.内含子和外显子结构分析表明,WRKY基因结构进化高度保守.保守元件分析表明,桃WRKY基因家族包含5个保守元件,元件1,2和3为WRKY盒,元件4,5为未知盒.桃WRKY基因家族都包含有WRKY盒,类型Ⅰ中含有2个WRKY盒;II-d亚组中含有未知元件5.半定量和荧光定量PCR结果显示,16个WRKY基因均在桃的根,茎,叶,花和果中表达,但其相对表达水平不同.     

昆虫性别决定机制研究进展

阐述昆虫的性别决定机制是理解昆虫性别分化调控的理论基础,也为人类有效控制害虫开辟了新方向。昆虫性别决定机制存在复杂性和多样性,但主要是内因即性别决定基因级联互作调控的结果。本文对近年来基于性别决定基因级联互作的昆虫性别决定机制研究进行了综述,主要包括性别决定基因概况和重要性别决定相关基因的分子级联互作关系。目前发现昆虫重要性别决定相关基因主要集中在常染色体上,且部分基因之间存在紧密的级联互作,如Sxl,tra,dsx,csd和fem等。在这些基因中,tra/fem→dsx的调控模式在已报道的昆虫中存在共性,即tra和dsx相对较保守且tra通过性特异剪切来调控下游dsx的转录形式。目前大多数昆虫的性别决定机制还不清楚,但近年来模式昆虫性别决定机制取得了一定进展,对非模式昆虫的研究还处于起步阶段但却越来越受到重视。     

昆虫种群的遗传调控

昆虫种群的遗传调控是利用昆虫自身生长发育的关键基因,采用性别控制开关,通过遗传转化使雄虫成为携带导致后代雌虫发育异常或雌性不育的遗传控制复合体（性别开关元件和靶标基因的复合体）．昆虫种群遗传调控是一种基于不育技术的昆虫种群控制系统,具有种类特异、环境友好和便捷高效等特点．目前为止,已经由早期的通过辐射不育方法发展到释放携带显性致死基因昆虫的方法,并在多种昆虫中获得成功．本文综述了昆虫种群遗传调控的发展历程,介绍了昆虫种群遗传调控相关的理论与方法,包括特异的调控元件、致死或缺陷基因和遗传转化体系的应用,并列举了几种昆虫种群遗传调控的实例,最后对于昆虫种群遗传调控系统中存在的问题以及可能的发展方向进行了展望．     

基因及其顺式调控元件在动物表型进化中的作用

顺式调控假说是当前进化发育生物学中重要的理论之一, 该假说认为顺式调控元件的进化是调控外表性状进化的主要遗传机制。然而越来越多的实验结果表明, 仅靠顺式调控假说远不足以解释复杂的进化发育过程, 其他因素也会导致表型的进化, 如：与顺式调控元件相联基因的蛋白序列改变; 基因及染色体组复制; 蛋白结构域与顺式调控元件的灵活性等。文章回顾了近年来顺式调控元件以及与顺式调控元件相联基因的进化发育研究, 探讨了进化发育生物学研究的新方法与新思路。     

Expression and characterization of a novel class of glutathione S-transferase from Anopheles dirus

A new Anopheles dirus glutathione S-transferase (GST) has been obtained and named adGST4-1. Both genomic DNA and cDNA for heterologous expression were acquired. The genomic sequence was 3188bp and consisted of the GST gene as well as flanking sequence. The flanking sequence was analyzed for possible regulatory elements that would control gene expression. In Drosophila several of these elements have been shown to be involved in development and cell differentiation. The deduced amino acid sequence has low identity compared with the four alternatively spliced enzymes, adGST1-1 to 1-4, from another An. dirus GST gene adgst1AS1. The percent identities are 30--40% and 11--12% comparing adGST4-1 to insect GSTs from Delta and Sigma classes, respectively. Enzyme characterization of adGST4-1 shows it to be distinct from the other An. dirus GSTs because of low enzyme activity for customary GST substrates including 1-chloro-2, 4-dinitrobenzene (CDNB). However, this enzyme has a greater affinity of interaction with pyrethroids compared to the other An. dirus GSTs.     

香樟全基因组WRKY基因家族的鉴定与分析

WRKY转录因子基因家族是植物特有的一类基因,在植物次生代谢、生物和非生物胁迫中起着重要的调节作用。本研究通过生物信息学方法,在香樟(Cinnamomum camphora (L.) Presl.)全基因组中鉴定了60个WRKY基因(CcWRKY),并将其分为Group Ⅰ~ Ⅲ,其中,Group Ⅰ和Ⅲ的成员发生了收缩现象;片段复制是CcWRKY基因扩张的主要驱动力;Group Ⅰ有完整的WRKY结构域和锌指基序,但Group Ⅱ、Ⅲ存在结构域和锌指基序的丢失和变异现象;CcWRKY基因的启动子区域具有激素类和胁迫类响应顺式作用元件;基因表达分析结果显示,在贫瘠环境(未施肥)中大多数CcWRKY基因在香樟各个组织中高表达,而环境适宜(施肥)条件下,基因表达量降低。     

Organisation and structural evolution of the rice glutathione <Emphasis Type="Italic">S</Emphasis>-transferase gene family

Glutathione S-transferases (GSTs) comprise a large family of key defence enzymes against xenobiotic toxicity. Here we describe the comprehensive characterisation of this important multigene family in the model monocot species rice [Oryza sativa (L.)]. Furthermore, we investigate the molecular evolution of the family based on the analysis of (1) the patterns of within-genome duplication, and (2) the phylogenetic relationships and evolutionary divergence among rice, Arabidopsis, maize and soybean GSTs. By in-silico screening of the EST and genome divisions of the Genbank/EMBL/DDBJ database we have isolated 59 putative genes and two pseudogenes, making this the largest plant GST family characterised to date. Of these, 38 (62%) are represented by genomic and EST sequences and 23 (38%) are known only from their genomic sequences. A preliminary survey of EST collections shows a large degree of variability in gene expression between different tissues and environmental conditions, with a small number of genes (13) accounting for 80% of all ESTs. Rice GSTs are organised in four main phylogenetic classes, with 91% of all rice genes belonging to the two plant-specific classes Tau (40 genes) and Phi (16 genes). Pairwise identity scores range between 17 and 98% for proteins of the same class, and 7 and 21% for interclass comparisons. Rapid evolution by gene duplication is suggested by the discovery of two large clusters of 7 and 23 closely related genes on chromosomes 1 and 10, respectively. A comparison of the complete GST families in two monocot and two dicot species suggests a monophyletic origin for all Theta and Zeta GSTs, and no more than three common ancestors for all Phi and Tau genes.Electronic Supplementary Material Supplementary material is available in the online version of this article at Communicated by M.-A. Grandbastien     

Glutathione transferase-like proteins encoded in genomes of yeasts and fungi: insights into evolution of a multifunctional protein superfamily

Most fungal glutathione transferases (GSTs) do not fit easily into any of the previously characterised classes by immunological, sequence or catalytic criteria. In contrast to the paucity of studies on GSTs cloned or isolated from fungal sources, a screen of databases revealed 67 GST-like sequences from 21 fungal species. Comparison by multiple sequence alignment generated a dendrogram revealing five clusters of GST-like proteins designated clusters 1, 2, EFIBgamma, Ure2p and MAK16, the last three of which have previously been related to the GST superfamily. Surprisingly, a relatively small number of fungal GSTs belong to mainstream classes and the previously-described fungal Gamma class is not widespread in the 21 species studied. Representative crystal structures are available for the EFIBgamma and Ure2p classes and the domain structures of representative sequences are compared with these. In addition, there are some "orphan" sequences that do not fit into any previously-described class, but show similarity to genes implicated in fungal biosynthetic gene clusters. We suggest that GST-like sequences are widespread in fungi, participating in a wide range of functions. They probably evolved by a process similar to domain "shuffling".     

The plant glutathione transferase gene family: genomic structure, functions, expression and evolution

Glutathione transferases (GSTs) are ubiquitous, multifunctional proteins encoded by large gene families. In different plant species this gene family is comprised of 25–60 members, that can be grouped into six classes on the basis of sequence identity, gene organization and active site residues in the protein. The Phi and Tau classes are the most represented and are plant specific, while Zeta and Theta GSTs are found also in animals. Despite pronounced sequence and functional diversification, GSTs have maintained a highly conserved three-dimensional structure through evolution. Most GSTs are cytosolic and active as dimers, performing diverse catalytic as well as non-catalytic roles in detoxification of xenobiotics, prevention of oxidative damage and endogenous metabolism. Among their catalytic activities are the conjugation of electrophilic substrates to glutathione, glutathione-dependent isomerizations and reductions of toxic organic hydroperoxides. Their main non-catalytic role is as hormone and flavonoid ligandins. GST genes are predominantly organized in clusters non-randomly distributed in the genome. Phylogenetic studies indicate that plant GSTs have mainly evolved after the divergence of plants, the two prevalent Phi and Tau classes being the result of recent, multiple duplication events.     

Divergence in structure and function of tau class glutathione transferase from Pinus tabulaeformis,P. yunnanensis and P. densata

Glutathione transferases (GSTs) are a family of enzymes that play important roles in stress tolerance and detoxification in plants. The plant GSTs are divided into four classes (phi, tau, zeta and theta), among which tau is the most numerously represented. To date, studies on GSTs in plants have focused largely on crop species. There is extremely little information on the molecular characteristics of GSTs in gymnosperms. Generalization on GST characteristics unique to gymnosperms and the patterns of GST evolution in plants cannot be made before more members of the gene family in conifers are described. In this study we report three new GSTs from Pinus tabulaeformis, Pinus densata and Pinus yunnanensis. Structural and phylogenetic analyses placed these three GSTs in tau class. The tau GST class is subdivided into three clades and this subdivision seems an ancient event that may have pre-dated the gymnosperm and angiosperm split. Sequence analysis revealed a highly conserved N-terminal domain in contrast to a highly variable C-terminal domain. Mutations even outside the critical glutathione-binding site in N-terminal domain can have pronounced effect on GST catalytic property. Thus, sequence similarity does not parallel functional specificity. The high diversity in C-terminal domain determines a wide range of substrate selectivity and specificity among tau GSTs. Thus the a few conserved residues in C-terminal domain seem essential to maintain the structure of the domain and the protein dimer. More extensive data on GST family organization and a thorough gene-by-gene analysis in conifers are needed to advance our understanding of the true diversity and evolution of GST in structure and function in plants.