真核细胞中多重表达框mRNA的选择性翻译起始

扫描模型和遗漏扫描模型是真核生物mRNA翻译起始的两种主要机制,但其仍存在某些例外情况,如对具有多顺反子结构的mRNA,选择性翻译起始的发生机制目前仍不清楚.本研究基于GFP蛋白开放表达框(ORF)构建了一系列重组表达载体,用以转录在移码翻译顺序及同一翻译顺序下,AUG起始密码子处于不同序列背景,以及间隔不同距离的多顺反子结构mRNA.通过转染人Bel 7402细胞系,研究了这些多顺反子结构mRNA的翻译起始模式.结果表明,在移码翻译顺序下,多顺反子mRNA可翻译出对应的不同蛋白质,而在同一翻译顺序下,GFP蛋白表达框中的多个AUG密码子,仅有首位起始密码子可发挥作用,提示核糖体在从首位起始密码子开始翻译的同时,可能会有部分核糖体继续向下扫描并识别下游的起始密码子,而这种选择性的翻译起始效率,主要取决于密码子所处的序列背景及间隔距离等因素.     

Contingent Conclusions: Year of Initiation Influences Ecological Field Experiments,but Temporal Replication is Rare

Interannual variation in experimental field conditions produce variability in the results of experiments monitored over multiple years, termed here “year effects.” When experimental treatments are replicated in separate years, interannual variation may influence treatment effects and produce significant treatment by initiation‐year interactions. Understanding the frequency and strength of these effects requires initiating identical experiments across years. We conducted a review of literature covering more than 500 experimental articles published in 7 journals between 1966 and 2008. Only 5% of the 276 general ecological field studies initiated experiments in multiple years. This rarity was even more evident in the journal Restoration Ecology, in which none of the 173 surveyed experimental studies initiated experiments in multiple years. In contrast, 48% of the 58 field experiments published in an agronomy journal were replicated across years. We found only 17 studies that tested treatment by initiation‐year interactions. Despite their rarity, 76% of these studies found significant interactions between treatment and initiation year. We conclude that the results of many ecological field experiments are likely to be contingent on the year in which they are implemented. We discuss the importance of treatment by initiation‐year interactions in ecology and restoration, factors that have hindered the inclusion of temporal replication in the past, and some suggestions for the appropriate design and analysis of temporally replicated experiments. We argue for more deliberate investigation of temporal contingency in ecological experimentation, especially in the field of restoration ecology, which may be particularly sensitive to treatment by initiation‐year interactions.     

真核生物DNA复制起始调控

复制起始调控是真核生物复制调控机制的重要环节,也是细胞生长调控的核心问题．对SV40病毒和酵母体系的研究为阐明真核生物的复制起始机制及其与细胞周期的关系提供了线索．目前,与DNA复制起始有关的多种蛋白质因子（如核蛋白P₁,DNA单链结合蛋白,DNA聚合酶α,增殖细胞核抗原等）的作用机理逐渐明朗,周期依赖的调控特点得到了证实文章着重介绍了DNA复制起始在细胞周期中的两个调控点及各种周期蛋白在该点的作用,文中还涉及复制起始异常与肿瘤发生的关系.     

A Consideration of Alternative Models for the Initiation of Translation in Eukaryotes

Abstract

Although recent biochemical and genetic investigations have produced some insights into the mechanism of initiation of translation in eukaryotic cells, two aspects of the initiation process remain controversial. One unsettled issue concerns a variety of functions that have been proposed for mRNA binding proteins, including some initiation factors. The need to distinguish between specific and nonspecific binding of proteins to mRNA is discussed herein. The possibility that certain initiation factors might act as RNA helicases is evaluated along with other ideas about the functions of mRNA- and ATP-binding factors. A second controversial issue concerns the universality of the scanning mechanism for initiation of translation. According to the conventional scanning model, the initial contact between eukaryotic ribosomes and mRNA occurs exclusively at the 5' terminus of the message, which is usually capped. The existence of uncapped mRNAs among a few plant and animal viruses has prompted a vigorous search for other modes of initiation. An “internal initiation” mechanism, first proposed for picornaviruses, has received considerable attention. Although a large body of evidence has been adduced in support of such a mechanism, many of the experiments appear flawed or inconclusive. Some suggestions are given for improving experiments designed to test the internal initiation hypothesis.     

真核翻译起始因子与肿瘤

真核翻译起始因子(eukaryotic translation initiation factors,eIFs)是一类在蛋白质翻译起始的过程中发挥各自不同作用的蛋白质。近年来的研究发现,eIFs除了在蛋白质翻译起始中起作用外,还具有其他的作用,而且多种eIFs均与肿瘤的发生和进展相关。现就eIFs、eIFs与肿瘤的相关性及其在肿瘤治疗方面的应用等研究进展作一综述。     

真核生物起始因子5

真核生物起始因子5(eIF-5)是一种重要的翻译起始因子,过去人们认为它只是GTP酶活化因子,催化eIF-2上的GTP水解,促进80 S起始复合体的形成.近年来人们发现它不仅可以催化eIF-2上的GTP水解,还参与eIF-3功能的发挥,与eIF-2、eIF-3同时结合,促进起始因子复合体的形成.     

激素信号在调节果树花芽发端假说的概述

概述了Lavee和Bangerth两个激素信号调节花芽发端假说的内容和证据,并分析了这两个假说的优缺点。认为旺盛营养生长的梢尖或正在发育果实的种子产生的极性运输的生长素（IAA）可能是抑制果树花芽发端的信号。尚不能确定来自正在发育果实种子的赤霉素（GA）自身是抑制花芽发端的信号,还是参与调节花芽发端信号的产生。营养芽中高水平的细胞分裂素（CTK）促进花芽发端,可能与较弱的IAA信号有关。同时指出,激素信号调节花芽发端的机理还有待完善。     

Initiation of DNA replication in eukaryotes is an intriguing cascade of protein interactions

Initiation of eukaryotic DNA replication is a complex process including the recognition of initiation sites on DNA, multi-step DNA preparation for duplication, and assembly of multi-protein complexes capable of beginning DNA synthesis at initiation sites. The process starts at the late M phase and lasts till the appropriate time of the S phase for each initiation site. A chain of interesting interactions between Orc1p-6p, Cdc6p, Mcm2p-7p, Mcm10p, Cdt1, Cdc45p, Dbf4/Cdc7p, RPA, and DNA polymerase takes place during this period. The sequence of these interactions is controlled by cyclin-dependent kinases, as well as by ubiquitin-dependent proteolysis in the proteasome. This review summarizes the data on proteins initiating DNA replication and factors controlling their activities.     

真核翻译起始因子3与肿瘤

真核翻译起始因子3是由多个亚基组成的,在真核翻译起始中发挥重要作用,近年来的研究表明其多个亚基在多种肿瘤细胞中存在异常表达的现象且与肿瘤的侵袭性、转移能力、分化程度及预后相关,使其有望成为肿瘤治疗的新靶点。     

Sigma54依赖性转录起始

作为细菌RNA聚合酶(RNAP)的组成型辅助因子,sigma70和sigma54分别参与了原核细胞不同类型基因的转录起始调控。sigma70负责管家基因的自发转录起始;而sigma54负责应激信号相关的基因转录起始。sigma54与RNAP形成复合物后,会通过空间阻滞的方式阻碍DNA进入RNAP中,抑制基因转录起始。当细胞环境变化后,特定应激信号会通过细菌增强子结合蛋白(bEBP)诱发sigma54的构象发生变化,解除sigma54对RNAP的抑制,启动sigma54依赖的基因转录。最近的结构生物学研究揭示了sigma54依赖性转录起始的若干复合物结构,包括全酶、封闭式复合物、2个中间状态复合物及开放式复合物。通过分析这些转录起始复合物的结构,本文阐述了转录起始过程中复合物的结构变化。描述并分析了sigma54和bEBP在转录起始过程中所发挥的功能。本文有助于了解转录起始分子水平的变化,为深入理解sigma54和bEBP促进转录起始的分子机制提供了参考。     

Initiation of protein synthesis in eukaryotes

The study of the regulation of initiation of protein synthesis has recently gained momentum because of the established relationship between translation initiation, cell growth and tumorigenesis. Therefore much effort is devoted to the role of protein kinases which are activated in signal transduction cascades and which are responsible for the phosphorylation of a number of initiation factors. These specific factors are mainly involved in the binding of messenger RNA to the 40S ribosome, a process that makes the unwinding of the 5 untranslated region necessary. It appears that the phosphorylation of these factors increases their ability for cap recognition and helicase activity. The enhanced phosphorylation of the messenger binding factors results not only in an overall stimulation of translation, but especially weak messengers are positively discriminated. The above mechanisms mainly deal with qualitative control of translation, i.e., messenger selection, but phosphorylation also plays a role in quantitative regulation of protein synthesis. The generation of active eIF-2, the initiation factor that binds the Met-tRNA _i and GTP, is dependent on a factor involved in the GDP-GTP exchange. Phosphorylation of eIF-2 results in sequestration of the exchange factor and a slowing down of the rate of initiation.Abbreviations eIF eukaryotic initiation factor - 5 UTR 5 untranslated region     

小麦茎顶端原基分化的综合模式

研究了小麦 (TriticumaestivumL .)茎顶端不同类型原基分化的动态过程 ,以明确原基分化的综合模式 ,并建立了不同原基分化之间的定量关系。结果表明 ,小麦叶原基和苞叶原基分化与播后累积生长度日 (GDD ,growingdegreedaysaftersowing)的关系呈S形曲线 ,而小穗原基和小花原基为上升段抛物曲线。从分化模式看 ,苞叶原基具备营养器官原基特征 ;小穗和小花原基的分化进程能较好地反映基因型和生态条件对顶端发育的影响。小麦茎顶端原基分化的综合模式为由三段子模式构成的近似S曲线。叶原基数由基因型和环境条件共同决定 ,而苞叶原基、小穗原基和小花原基数以环境因子的影响为主。以平均热间距来衡量 ,适期播种处理的叶片、苞叶和小穗原基分化速率最高 ;而小花原基数与小花分化持续期之间的数量关系最为密切。研究结果有助于揭示和理解小麦茎顶端发育的生物学规律。     

小麦茎顶端原基分化的综合模式

研究了小麦(Triticum aestivum L.)茎顶端不同类型原基分化的动态过程,以明确原基分化的综合模式,并建立了不同原基分化之间的定量关系.结果表明,小麦叶原基和苞叶原基分化与播后累积生长度日(GDD, growing degree days after sowing)的关系呈S形曲线,而小穗原基和小花原基为上升段抛物曲线.从分化模式看,苞叶原基具备营养器官原基特征;小穗和小花原基的分化进程能较好地反映基因型和生态条件对顶端发育的影响.小麦茎顶端原基分化的综合模式为由三段子模式构成的近似S曲线.叶原基数由基因型和环境条件共同决定,而苞叶原基、小穗原基和小花原基数以环境因子的影响为主.以平均热间距来衡量,适期播种处理的叶片、苞叶和小穗原基分化速率最高;而小花原基数与小花分化持续期之间的数量关系最为密切.研究结果有助于揭示和理解小麦茎顶端发育的生物学规律.     

Lateral Root Initiation or the Birth of a New Meristem

Root branching happens through the formation of new meristems out of a limited number of pericycle cells inside the parent root. As opposed to shoot branching, the study of lateral root formation has been complicated due to its internal nature, and a lot of questions remain unanswered. However, due to the availability of new molecular tools and more complete genomic data in the model species Arabidopsis, the probability to find new and crucial elements in the lateral root formation pathway has increased. Increasingly more data are supporting the idea that lateral root founder cells become specified in young root parts before differentiation is accomplished. Next, pericycle founder cells undergo anticlinal asymmetric, divisions followed by an organized cell division pattern resulting in the formation of a new organ. The whole process of cell cycle progression and stimulation of the molecular pathway towards lateral root initiation is triggered by the plant hormone auxin. In this review, we aim to give an overview on the developmental events taking place from the very early specification of founder cells in the pericycle until the first anticlinal divisions by combining the knowledge originating from classical physiology studies with new insights from genetic-molecular analyses. Based on the current knowledge derived from recent genetic and developmental studies, we propose here a hypothetical model for LRI.     

二级结构形成：蛋白质折叠起始过程的框架模型

框架模型认为二级结构形成是蛋白质起始过程的结构基础.文章介绍蛋白质同源片段的溶液构象及其构象研究法和多肽二级结构的从头设计,并综述这些研究成果应用于折叠起始过程的理论模型和蛋白质折叠起始过程的最新研究进展.     

Homologous Mammalian Brain Cell Lysate System for the Initiation and Translation of Exogenous mRNAs

Abstract: The rate of protein synthesis in mammalian brain tissue is affected by a variety of physiological conditions, both natural and induced. The process of initiation may be involved in some of the observed changes, although as yet the actual rates of initiation of natural mRNAs have not been directly measured in these circumstances. One approach to studying the regulation of protein synthesis in brain tissue would be to utilize a homologous cell-free system to examine in vitro the translation of various added mRNAs. The present report describes a micrococcal nuclease-treated cell-free lysate system derived from fetal mouse brain tissue which is capable of actively initiating and translating exogenously added mRNA. Sodium dodecyl sulfate-polyacrylamide slab gel electrophoretic analysis of the specific protein products of the reaction mixture allowed a qualitative and quantitative assessment of the translational process under a variety of experimental conditions. Optimal conditions for mRNA-dependent protein synthesis were the following: 30°C incubation temperature; 80–100 mM-KCl; 2.1 mM-Mg²⁺; 50 μM-spermhe; and 10 μg/ml poly A(+) mRNA. Incorporation of L-[³⁵S]methionine into proteins required ATP, GTP, and an energy regenerating system. The addition of saturating amounts of a homologous "initiation factors" fraction stimulated incorporation twofold during the first 20 min of incubation, while the patterns of inhibition observed upon the addition of 5 × 10^-5 M-aurin tricarboxylic acid at various periods during incubation demonstrated the occurrence of multiple rounds of initiation.     

被子植物胚乳自主发生的分子机理研究进展

双受精是被子植物的一个有效的进化策略,增加了胚和种苗成活的机会,其中胚乳在胚以及种子发育中至关重要。然而极少数进行无融合生殖的被子植物,它们的胚乳可以在假受精或不受精的情况下克服基因组印记效应的影响且正常发育。拟南芥胚乳自主发生突变体及相关基因的克隆,使人们可以研究和比较自然和突变植物胚乳自主发生的分子机制。本文着重介绍基因组印记对有性生殖和无融合生殖植物胚乳发育的影响,分析和讨论近年来发现的有关胚乳自主发生的基因（如MEA,FIS2,FIE,MSI1和PHE1）及其可能的作用机理。     

人白细胞介素－3基因翻译起始区的改造提高其在大肠杆菌中的表达水平

为了提高人白细胞介素－3(bhIL-3)在大肠杆菌中的表达,在计算机辅助下,设计合成了PCR突变引物,用于改造起始密码AUG上下游序列,并在不改变5’端氨基酸编码的前提下,尽可能选用大肠杆菌高频使用的密码子。将经改造后的Hil-3cDNA和翻译起始区置于PL启动子之下,转入大肠杆菌Tapl06,经42℃热诱导后．获得表达产物,提高表达水平近一倍,表达量达到菌体总蛋白量的30％左右。表达产物经Western blot验证,经PVDF膜转移后进行N端顺序分析,证明前15个氨基酸正确,产物经包涵体纯化后,纯度提高至80％以上,初步复性后能明显促进Hil－3依赖细胞的生长。