沙丁胺醇对黑腹果蝇基因组稳定性、细胞凋亡和蛋白表达的影响

【目的】已有研究表明,食用了饲喂以沙丁胺醇为主要成分的瘦肉精的动物肉类后,瘦肉精成分会在人体内富集,摄入过量沙丁胺醇会对生物体造成不良影响,但是,其具体毒性作用机理目前尚不明确。作为一种模式生物,黑腹果蝇Drosophila melanogaster与哺乳动物的基因具有较高的同源性,且具有繁殖周期短、方便进行遗传操作等优势。因此,我们通过研究过量沙丁胺醇对黑腹果蝇基因组稳定性、细胞凋亡和蛋白表达的影响,来探究它对生物体毒性作用的机理。【方法】将野生型黑腹果蝇3龄幼虫用含沙丁胺醇（120 μg/mL）的饲料饲喂2 h后,对幼虫翅成虫盘进行H2Av抗体免疫染色。选取rpr-lacZ转基因黑腹果蝇1龄幼虫用含沙丁胺醇（40 和120 μg/mL）的饲料饲喂,对幼虫翅成虫盘进行lacZ活性测定。提取沙丁胺醇处理后的野生型3龄幼虫总蛋白,采用SDS-PAGE比较对照组和实验组蛋白表达的差异,并通过质谱分析差异蛋白的氨基酸序列。【结果】沙丁胺醇处理后,经免疫荧光染色发现野生型黑腹果蝇幼虫翅成虫盘部分细胞中组蛋白H2Av的量有显著增加。随着沙丁胺醇浓度的增加,转rpr-lacZ报告基因黑腹果蝇成虫盘细胞lacZ活性增强。采用SDS-PAGE和质谱分析表明,沙丁胺醇处理后黑腹果蝇肌动蛋白（Actin-87E）和异柠檬酸脱氢酶表达量上升。【结论】沙丁胺醇处理会引起黑腹果蝇细胞核DNA损伤,对基因稳定性有显著影响,并且会促进细胞凋亡和蛋白表达的改变。沙丁胺醇可能通过促进肌肉收缩和加速生物体能量代谢这两方面来减少脂肪积蓄。     

果蝇基因组与功能基因研究进展

黑腹果蝇Drosophila melanogaster是生物科学研究中重要的模式动物之一。2000年,黑腹果蝇全基因组测序完成,随后基因组序列质量不断完善,对其功能基因进行深入研究,为其他高等动物基因组和功能基因的研究提供了巨大帮助。本文综述了近年来基因组功能元件、比较基因组学等方面的最新研究成果,着重介绍了功能基因在Hh信号通路、细胞凋亡方面的研究进展,并对最新的功能基因研究技术进行了简要概述。     

线粒体与细胞凋亡机制

细胞凋亡是生理性的细胞死亡过程,受到多种基因的精确调节,一类被统称为ｃａｓｐａｓｅ的半胱氨酸蛋白酶是细胞凋亡程序的执行者,综们被激活后作用于细胞内的一些蛋白质,经起细胞凋亡。线粒体中含有许多凋亡相关因子,在凋亡信号转导中起着重要作用。细胞受到凋亡刺激后,细胞色素ｃ、ＡＩＦ、ｃａｓｐａｓｅ－９等凋亡相关因子从线粒体中释放出来。细胞色素ｃ通过和Ａｐａｆ－１、ｃａｓｐａｓｅ－９相互作用,激活ｃａｓｐａｓ     

果蝇转座因子对基因组进化的影响

真核生物基因组织有很多可移动ＤＮＡ片段为称转座因子,果蝇是大量系统研究的最好实验材料之一,其基因组的１０％－１２％是由转座因子组成,在宿主中,ＴＥs也许改变基因表达模型,也许改变ＯＲＦs编码序列,也许对细胞功能产生影响,这此因子遗传的可动性也可能使它们适于建造载体产生转基因生物。因此,对ＴＥs进化的动态研究以及对宿主基因组进化影响的探索将有助于TEs作为载体的细胞工程研究。     

细胞凋亡的信号转导机制与调节

本文综述近年来细胞凋亡信号转导机制的有关研究进展,重点概述了凋亡信号转导的死亡受体途径,线粒体－细胞色素Ｃ途径的信号转导机制及信号转导的有关调节机制的研究进展。     

细胞凋亡机制的研究进展

以线虫和哺乳动物为实验材料,综述近年来对细胞凋亡机制的研究进展。     

ced,rpr和病毒基因与细胞凋亡

本简要综述了线虫ｃｅｄ（ｃｅｄ－３、ｃｅｄ－４、ｃｅｄ－９）基因、果蝇ｒｐｒ基因和一些病毒基因对相关程序性细胞死亡（ＰＣＤ）的调控及其可能机制,并初步探讨了这些基因诱导或抑制ＰＣＤ的可能生物学或医学意义。深入研究这些ＰＣＤ相关的低等运行动物和病毒基因的作用机制,将对高等动物乃至于人的ＰＣＤ基因调控的探讨产生重大影响。     

重金属诱导细胞凋亡的分子机制

重金属诱导的细胞凋亡是一个十分复杂的过程,不同种类的重金属以及同类重金属离子的不同价态所诱导的凋亡效应及其分子机制不尽相同。目前的研究表明,重金属与DNA形成加合物而导致DNA损伤可能是引发细胞凋亡的重要步骤：多种重金属能通过激活内质网、线粒体钙通道,使Ca^2 释放进入细胞质而引发凋亡;重金属还能使细胞中ROS升高,在直接导致DNA损伤的同时,启动与线粒体相关的细胞凋亡信号通路。此外,ROS还能通过MAPKs增强JNK介导的：FasL和Fas表达,最终使caspase-3和caspase-7激活,从而促进凋亡的发生。重金属诱导细胞凋亡还涉及一系列重要基因和蛋白质的参与,包括促进凋亡的Src家族酪氨酸激酶、bax、fas和p53等基因及相关蛋白,抑制凋亡的Sp1锌指转录因子、bcl-2和myc等基因及相关蛋白。部分重金属如镉、锌等对细胞凋亡具有诱导和拮抗双重效应,其中拮抗效应主要是通过与自由钙离子协同进行的,而诱导效应则可能是通过调节caspase-3活性而实现的。     

HCV核心蛋白诱导Cos-7细胞凋亡

将丙型肝炎病毒(HCV)核心蛋白(Core)基因的全长序列插入到真核表达载体pCDNA3 CMV启动子下游,构建真核表达载体pCDNA3-Core,用脂质体LipoVecTM转染Cos-7细胞系进行瞬时表达;DNA转染24h后用免疫斑点试验检测在细胞中表达的Core蛋白;转染72h后用Hoechst染色和DNA Ladder检测Cos-7细胞的凋亡情况;荧光染色观察到了细胞凋亡核碎裂,琼脂糖凝胶电泳也呈现出180-200bp整数倍的梯形带,呈现典型的细胞凋亡特征.这些结果表明HCV Core蛋白的表达能引起Cos-7细胞凋亡,Core蛋白的这种功能可能在HCV的持续感染过程中起着一定的作用.     

The Drosophila melanogaster genome sequencing and annotation projects: a status report.

The sequence and genome annotations of Drosophila melanogaster were initially published in late 1999 and early 2000. Since then, the Berkeley Drosophila Genome Project (BDGP) and FlyBase have improved the quality of the sequence and reviewed the annotations by hand, respectively, to produce an account of the fruit fly genome that is of the highest quality. This review discusses the main features of this process, both from the point of view of the biology revealed in the end result and in the development of software that has been central to this genome sequencing and annotation project.     

CoreCruncher: Fast and Robust Construction of Core Genomes in Large Prokaryotic Data Sets

The core genome represents the set of genes shared by all, or nearly all, strains of a given population or species of prokaryotes. Inferring the core genome is integral to many genomic analyses, however, most methods rely on the comparison of all the pairs of genomes; a step that is becoming increasingly difficult given the massive accumulation of genomic data. Here, we present CoreCruncher; a program that robustly and rapidly constructs core genomes across hundreds or thousands of genomes. CoreCruncher does not compute all pairwise genome comparisons and uses a heuristic based on the distributions of identity scores to classify sequences as orthologs or paralogs/xenologs. Although it is much faster than current methods, our results indicate that our approach is more conservative than other tools and less sensitive to the presence of paralogs and xenologs. CoreCruncher is freely available from: https://github.com/lbobay/CoreCruncher. CoreCruncher is written in Python 3.7 and can also run on Python 2.7 without modification. It requires the python library Numpy and either Usearch or Blast. Certain options require the programs muscle or mafft.     

黑腹果蝇种组的核型与进化研究

观察了国内黑腹果蝇种组34种果蝇的有丝分裂中期核型,其中首次描述了一些新核型。系统地分析了黑腹果蝇种组8个种亚组之间的核型进化关系及种间亲缘关系。结果是:elegans种亚组的核型为A型;eugracilis、melanogaster和ficusphila种亚组的核型为C型;takahashii和suzukii种亚组的核型为C型和D型;montium种亚组的核型为B、C、C’、D、D’、和E型;ananassae种亚组的核型为F、G和H型。从核型分化的角度可以将黑腹果蝇种组分为5个谱系:elegans,eugracilis-melanogaster-ficusphila,takkahashii-suzukii,montium,ananassae。这与2004年Yang等的观点基本一致,正好从核型进化的角度验证了Yang通过DNA序列分析所得到的结果。差别只在于elegans种亚组,作者把它单独列为一支,认为是祖先种亚组。通过选取同一种果蝇的几个不同地域单雌系的核型分析,结果表明:同一种果蝇的核型存在地域差异。这种差异可能是由于不同生境造成,也可能是本身进化程度的差异,或是两种因素相互作用的结果。     

Evolution of paralogous genes: Reconstruction of genome rearrangements through comparison of multiple genomes within Staphylococcus aureus

Analysis of evolution of paralogous genes in a genome is central to our understanding of genome evolution. Comparison of closely related bacterial genomes, which has provided clues as to how genome sequences evolve under natural conditions, would help in such an analysis. With species Staphylococcus aureus, whole-genome sequences have been decoded for seven strains. We compared their DNA sequences to detect large genome polymorphisms and to deduce mechanisms of genome rearrangements that have formed each of them. We first compared strains N315 and Mu50, which make one of the most closely related strain pairs, at the single-nucleotide resolution to catalogue all the middle-sized (more than 10 bp) to large genome polymorphisms such as indels and substitutions. These polymorphisms include two paralogous gene sets, one in a tandem paralogue gene cluster for toxins in a genomic island and the other in a ribosomal RNA operon. We also focused on two other tandem paralogue gene clusters and type I restriction-modification (RM) genes on the genomic islands. Then we reconstructed rearrangement events responsible for these polymorphisms, in the paralogous genes and the others, with reference to the other five genomes. For the tandem paralogue gene clusters, we were able to infer sequences for homologous recombination generating the change in the repeat number. These sequences were conserved among the repeated paralogous units likely because of their functional importance. The sequence specificity (S) subunit of type I RM systems showed recombination, likely at the homology of a conserved region, between the two variable regions for sequence specificity. We also noticed novel alleles in the ribosomal RNA operons and suggested a role for illegitimate recombination in their formation. These results revealed importance of recombination involving long conserved sequence in the evolution of paralogous genes in the genome.     

生命科学与人类疾病研究的重要模型——果蝇

黑腹果蝇(Drosophilamelanogaster)是生物学研究中最重要的模式生物之一,它在遗传的染色体理论建立中起到非常重要的作用。由于果蝇自身独特的优势,20世纪70年代以来,它又在发育生物学、神经科学、人类疾病研究等领域得到广泛应用,作出许多新的重要贡献。果蝇在神经退行性疾病研究中是非常有用的模型。可以预期,随着研究手段的丰富及科学的发展,果蝇将作为一种理想的模式生物在生物医学中发挥更大的作用。     

Comparison between the molecular characteristics and the potential activity of X and Y nucleolar organizers from various Drosophila melanogaster laboratory lines

The molecular characteristics of nucleolar organizers from X and Y chromosomes of different Drosophila melanogaster lines have previously been studied (17). By analysis of appropriate genetic crosses we show in the present study that the X and Y chromosomes of these lines can confer different degrees of resistance on an inhibitor of ribosomal RNA synthesis (beta exotoxin or thuringiensin) present in the thermostable supernatant of Bacillus thuringiensis cultures. None of the lines studied gives rise to any particular phenotype under normal culture conditions; variations in the degree of supernatant resistance of these lines provide a relative measure of what can be called the potential activity of the nucleolar organizers of the different X and/or Y chromosomes. The potential activity of the Y nucleolar organizers is found to be generally higher than that of the X organizers. This result can be correlated with the fact that the number of uninterrupted ribosomal units is much greater on the Y chromosomes than on the Xs. Significant variations in potential activity have been shown to occur among the X as also among the Y nucleolar organizers. Comparison between the molecular characteristics of the nucleolar organizers and their level of activity shows that among the different ribosomal units, only those devoid of insertion interfere with the level of activity. However, some of our results could also indicate that not all the uninterrupted units have the same level of activity; this level could be related to the size of the nontranscribed spacer of the ribosomal units.(ABSTRACT TRUNCATED AT 250 WORDS)     

Native Molecular Forms of Head Acetylcholinesterase from Adult Drosophila melanogaster: Quaternary Structure and Hydrophobic Character

The native molecular forms of acetylcholinesterase (AChE) present in adult Drosophila heads were characterized by sedimentation analysis in sucrose gradients and by nondenaturing electrophoresis. The hydrophobic properties of AChE forms were studied by comparing their migration in the presence of Triton X100, 10-oleyl ether, or sodium deoxycholate, or in the absence of detergent. We examined the polymeric structure of AChE forms by disulfide bridge reduction. We found that the major native molecular form is an amphiphilic dimer which is converted into hydrophilic dimer and monomer on autolysis of the extracts, or into a catalytically active amphiphilic monomer by partial reduction. The latter component exists only as trace amounts in the native enzyme. Two additional minor native forms were identified as hydrophilic dimer and monomer. Although a significant proportion of AChE was only solubilized in high salt, following extractions in low salt, this high salt-soluble fraction contained the same molecular forms as the low salt-soluble fractions: thus, we did not detect any molecular form resembling the asymmetric forms of vertebrate cholinesterases.     

Drosophila subpulchrella, a new species of the Drosophila suzukii species subgroup from Japan and China (Diptera: Drosophilidae)

Drosophila (Sophophora) subpulchrella Takamori and Watabe, sp. nov., of the D. suzukii subgroup in the D. melanogaster species group, is described from Japan and southern China, and compared with its sibling species, D. pulchrella Tan et al. distributed in the Yun‐Gui Highland, south‐western China. The results of cross‐experiments show a complete pre‐mating isolation between D. subpulchrella and D. pulchrella.