Dynamics of ATP-dependent chromatin assembly by ACF

The assembly of DNA into chromatin is a critical step in the replication and repair of the eukaryotic genome. It has been known for nearly 20 years that chromatin assembly is an ATP-dependent process. ATP-dependent chromatin-assembly factor (ACF) uses the energy of ATP hydrolysis for the deposition of histones into periodic nucleosome arrays, and the ISWI subunit of ACF is an ATPase that is related to helicases. Here we show that ACF becomes committed to the DNA template upon initiation of chromatin assembly. We also observed that ACF assembles nucleosomes in localized arrays, rather than randomly distributing them. By using a purified ACF-dependent system for chromatin assembly, we found that ACF hydrolyses about 2#150;4 molecules of ATP per base pair in the assembly of nucleosomes. This level of ATP hydrolysis is similar to that used by DNA helicases for the unwinding of DNA. These results suggest that a tracking mechanism exists in which ACF assembles chromatin as an ATP-driven DNA-translocating motor. Moreover, this proposed mechanism for ACF may be relevant to the function of other chromatin-remodelling factors that contain ISWI subunits.     

In vitro centromere and kinetochore assembly on defined chromatin templates

During cell division, chromosomes are segregated to nascent daughter cells by attaching to the microtubules of the mitotic spindle through the kinetochore. Kinetochores are assembled on a specialized chromatin domain called the centromere, which is characterized by the replacement of nucleosomal histone H3 with the histone H3 variant centromere protein A (CENP-A). CENP-A is essential for centromere and kinetochore formation in all eukaryotes but it is unknown how CENP-A chromatin directs centromere and kinetochore assembly. Here we generate synthetic CENP-A chromatin that recapitulates essential steps of centromere and kinetochore assembly in vitro. We show that reconstituted CENP-A chromatin when added to cell-free extracts is sufficient for the assembly of centromere and kinetochore proteins, microtubule binding and stabilization, and mitotic checkpoint function. Using chromatin assembled from histone H3/CENP-A chimaeras, we demonstrate that the conserved carboxy terminus of CENP-A is necessary and sufficient for centromere and kinetochore protein recruitment and function but that the CENP-A targeting domain--required for new CENP-A histone assembly--is not. These data show that two of the primary requirements for accurate chromosome segregation, the assembly of the kinetochore and the propagation of CENP-A chromatin, are specified by different elements in the CENP-A histone. Our unique cell-free system enables complete control and manipulation of the chromatin substrate and thus presents a powerful tool to study centromere and kinetochore assembly.     

Preferential electrical coupling regulates neocortical lineage-dependent microcircuit assembly

Radial glial cells are the primary neural progenitor cells in the developing neocortex. Consecutive asymmetric divisions of individual radial glial progenitor cells produce a number of sister excitatory neurons that migrate along the elongated radial glial fibre, resulting in the formation of ontogenetic columns. Moreover, sister excitatory neurons in ontogenetic columns preferentially develop specific chemical synapses with each other rather than with nearby non-siblings. Although these findings provide crucial insight into the emergence of functional columns in the neocortex, little is known about the basis of this lineage-dependent assembly of excitatory neuron microcircuits at single-cell resolution. Here we show that transient electrical coupling between radially aligned sister excitatory neurons regulates the subsequent formation of specific chemical synapses in the neocortex. Multiple-electrode whole-cell recordings showed that sister excitatory neurons preferentially form strong electrical coupling with each other rather than with adjacent non-sister excitatory neurons during early postnatal stages. This preferential coupling allows selective electrical communication between sister excitatory neurons, promoting their action potential generation and synchronous firing. Interestingly, although this electrical communication largely disappears before the appearance of chemical synapses, blockade of the electrical communication impairs the subsequent formation of specific chemical synapses between sister excitatory neurons in ontogenetic columns. These results suggest a strong link between lineage-dependent transient electrical coupling and the assembly of precise excitatory neuron microcircuits in the neocortex.     

The RCAF complex mediates chromatin assembly during DNA replication and repair

Chromatin assembly is a fundamental biological process that is essential for the replication and maintenance of the eukaryotic genome. In dividing cells, newly synthesized DNA is rapidly assembled into chromatin by the deposition of a tetramer of the histone proteins H3 and H4, followed by the deposition of two dimers of histones H2A and H2B to complete the nucleosome-the fundamental repeating unit of chromatin. Here we describe the identification, purification, cloning, and characterization of replication-coupling assembly factor (RCAF), a novel protein complex that facilitates the assembly of nucleosomes onto newly replicated DNA in vitro. RCAF comprises the Drosophila homologue of anti-silencing function 1 protein ASF1 and histones H3 and H4. The specific acetylation pattern of H3 and H4 in RCAF is identical to that of newly synthesized histones. Genetic analyses in Saccharomyces cerevisiae demonstrate that ASF1 is essential for normal cell cycle progression, and suggest that RCAF mediates chromatin assembly after DNA replication and the repair of double-strand DNA damage in vivo.     

催化氧化合成过氧乙酸的本征动力学

过氧乙酸是一种用途广泛的有机过氧化物.利用固体酸催化剂催化氧化乙酸从而间接氧化环己酮合成ε-己内酯是一种绿色生产过程,其中催化氧化乙酸生成过氧乙酸是关键步骤.借助化学滴定分析法,使用Mg/Sn/W复合氧化物为催化剂,以H2O2为氧化剂,以乙酸丁酯为共沸剂研究氧化乙酸生成过氧乙酸过程的本征动力学及过程中H2O2分解动力学.当催化剂粒径为160～200目时,内扩散影响可基本消除,当搅拌速度大于800 r·min－1时,外扩散可以忽略.使用幂函数型模型对动力学过程进行描述,研究结果表明,过氧乙酸生成反应本征动力学速率方程对H2O2、乙酸均为一级,反应活化能为6.76141×104 J·mol－1,指前因子为6.78090×106;适用于该反应过程体系的H2O2分解反应动力学的反应级数为1.59345,反应活化能为7.59041×104 J·mol－1,指前因子为1.08795×108.     

四种缩合试剂对含N-Me二肽的合成

用N,N′-二环己基碳二亚胺(DCC)/1-羟基-苯并-三氮唑(HOBt)、1-乙基-(3-二甲基氨基丙基)碳酰二亚胺(EDC)/HOBt、2-乙氧基-1-乙氧碳酰基-1,2-二氢喹啉(EEDQ)、3-(二乙氧基磷酰基)-氧-1,2,3-苯并三嗪-4(3H)-酮(DEPBT)4种常用缩合试剂合成了含N-Me二肽Boc-Leu-N-Me-Leu-OBzl,产率分别为73.5%～92.8%,65.2%～85.9%,38.9%,87.8%.比较发现反应物Boc-Leu-OH、H-N-Me-Leu-OBzl与缩合试剂DCC/HOBt的物质的量比在2.0:1.0:2.0时产率最高.利用此比例合成了3个立体异构体,产率较高.     

考虑多公差耦合的几何要素误差建模及产品装配精度预测

为了准确有效地实现零件几何要素误差建模,提出了一种基于多公差耦合作用下的几何要素误差建模方法.该方法采用小位移旋量(SDT,small displacement torsor)描述公差,针对平动公差与固定公差耦合建立几何要素误差模型,通过利用约束方程的不合格率p,推导各个误差分量参数的实际的分布规律,建立几何要素误差模型.采用雅可比旋量模型将该方法扩展应用于复杂装配体的装配误差建模中,实现了装配精度的预测.以顶尖尾座装配精度预测为例,验证了该方法的合理性及可行性.     

The histone H3.3 chaperone HIRA is essential for chromatin assembly in the male pronucleus

In sexually reproducing animals, a crucial step in zygote formation is the decondensation of the fertilizing sperm nucleus into a DNA replication-competent male pronucleus. Genome-wide nucleosome assembly on paternal DNA implies the replacement of sperm chromosomal proteins, such as protamines, by maternally provided histones. This fundamental process is specifically impaired in sésame (ssm), a unique Drosophila maternal effect mutant that prevents male pronucleus formation. Here we show that ssm is a point mutation in the Hira gene, thus demonstrating that the histone chaperone protein HIRA is required for nucleosome assembly during sperm nucleus decondensation. In vertebrates, HIRA has recently been shown to be critical for a nucleosome assembly pathway independent of DNA synthesis that specifically involves the H3.3 histone variant. We also show that nucleosomes containing H3.3, and not H3, are specifically assembled in paternal Drosophila chromatin before the first round of DNA replication. The exclusive marking of paternal chromosomes with H3.3 represents a primary epigenetic distinction between parental genomes in the zygote, and underlines an important consequence of the critical and highly specialized function of HIRA at fertilization.     

The synthesis and in vivo assembly of functional antibodies in yeast

The yeast Saccharomyces cerevisiae can synthesize, process and secrete higher eukaryotic proteins. We have investigated the expression of immunoglobulin chains in yeast and demonstrate here the synthesis, processing and secretion of light and heavy chains, the glycosylation of heavy chain, the intracellular localization of these foreign proteins by immunofluorescence, and the detection of functional antibodies in cells co-expressing both chains. This may provide the basis of a microbial fermentation process for the production of monoclonal antibodies. The co-expression of light and heavy chains in Escherichia coli has been reported but functional antibodies were not assembled in vivo. Furthermore, only low-level assembly of these chains was found in vitro.     

论"固有色"及"固有色彩观念"

进行色彩写生必须具有科学的光色变化观念,从客观物象当时当地的具体色彩实际出发,排除“固有色彩观念”的干扰,抛弃对“固有色”的盲目追求,以自然为师,努力表现一切被画物象极为丰富的色彩变化。     

如何运用篮球运动自身固有的规律进行教学

文章认为篮球训练课不仅要遵循一定的教学原则和教学规律 ,而且要采取灵活多变的教学方法。这样才能使学生尽快地掌握篮球技术。     

氧热法电石合成的反应平衡和热匹配分析

针对氧热法电石合成中吸热的生成反应和放热的碳燃烧反应耦合,从热力学角度对电石生成途径、反应化学计量平衡以及吸、放热反应热耦合进行了分析。结果表明:(1)电石由CaO+3C→CaC2+CO一步直接生成的可能性更大;(2)不同化学计量对应4种不同反应体系,各体系电石平衡转化率都随温度升高而升高,随压强增大而减小,电石与氧化钙发生副反应的转化率大致为随温度升高而先升后降;(3)反应热匹配量和匹配条件取决于电石生成反应物料处理量和电石纯度要求。电石生成反应与燃烧供热反应耦合于同一反应器是可行的。     

Electron transfer from protein to DNA in irradiated chromatin

Irradiation of dry or fully hydrated frozen DNA systems (conditions of direct damage) has been shown by electron-spin resonance spectroscopy to give rise to electron-gain centres localized on thymine (T.-) and electron loss centres ('holes') localized on guanine (G.+) with approximately equal yields. Our parallel studies on the development of both single- and double-strand breaks under comparable conditions provide good evidence that these radical centres are the precursors to such damage, and we and others have argued that this may be of relevance to the damage pathways in vivo. We now report evidence that when DNA is complexed to proteins as it is in the nuclei of eukaryotes, electron transfer from the histone to DNA is facile, leading to a significant increase in the yield of electron-gain centres in DNA as judged from their electron-spin resonance spectra. In contrast 'holes' generated in the protein are trapped and do not lead to any detectable increase in the yields of G.+.     

基于内部信息置换的创新设计

探讨创新设计的理论框架，进而提出一种基于系统内部信息置换的创新设计方法．该方法利用超图重写系统组织系统的内部结构和创新过程．它以现有系统而不是外部需求为创新的起点，通过对现有系统的超图进行系列置换而产生新的创造．     

高斯哥本哈根获奖论文及其对内蕴微分几何学的贡献

运用文献分析研究和数学史比较研究方法,分析了高斯哥本哈根获奖论文产生的历史背景及其对内蕴微分几何学的贡献．结果表明,高斯完成于1822年12月的获奖论文,不仅解决了哥本哈根科学院的征奖问题,更重要的是它奠定了其内蕴微分几何学研究的基本思想和方法．