EB1蛋白在调节微管动态、纺锤体定位和染色体稳定性中的作用

EB1(the end-binding protein 1)蛋白家族是一群广泛存在且高度保守的微管相关蛋白,存在于从酵母到人类的广泛的生物体中.它与微管正极和中心体结合,参与了绝大部分基于微管的生理过程,包括:维持细胞极性,调节染色体稳定性,有丝分裂纺锤体的定位,将微管锚定到成核位点.自从1995年,Su等人在人细胞中发现了EB1基因,各种生物体中EB1的同源物质被相继报道.十年来,人们通过对不同生物体的研究,试图揭开EB1在细胞中的分布以及它的生理功能.然而到目前为止,对于EB1的了解还非常有限.本文结合国外的研究成果,对EB1蛋白在调节微管动态、纺锤体定位和染色体的稳定性方面以及它与APC(the adenomatous polyposis coli)之间的相互作用作以综述.     

Sister-chromatid separation at anaphase onset is promoted by cleavage of the cohesin subunit Scc1.

Cohesion between sister chromatids is established during DNA replication and depends on a multiprotein complex called cohesin. Attachment of sister kinetochores to the mitotic spindle during mitosis generates forces that would immediately split sister chromatids were it not opposed by cohesion. Cohesion is essential for the alignment of chromosomes in metaphase but must be abolished for sister separation to start during anaphase. In the budding yeast Saccharomyces cerevisiae, loss of sister-chromatid cohesion depends on a separating protein (separin) called Esp1 and is accompanied by dissociation from the chromosomes of the cohesion subunit Scc1. Here we show that Esp1 causes the dissociation of Scc1 from chromosomes by stimulating its cleavage by proteolysis. A mutant Scc1 is described that is resistant to Esp1-dependent cleavage and which blocks both sister-chromatid separation and the dissociation of Scc1 from chromosomes. The evolutionary conservation of separins indicates that the proteolytic cleavage of cohesion proteins might be a general mechanism for triggering anaphase.     

Cohesin Rec8 is required for reductional chromosome segregation at meiosis.

When cells exit from mitotic cell division, their sister chromatids lose cohesion and separate to opposite poles of the dividing cell, resulting in equational chromosome segregation. In contrast, the reductional segregation of the first stage of meiotic cell division (meiosis I) requires that sister chromatids remain associated through their centromeres and move together to the same pole. Centromeric cohesion is lost as cells exit from meiosis II and sister chromatids can then separate. The fission yeast cohesin protein Rec8 is specific to and required for meiosis. Here we show that Rec8 appears in the centromeres and adjacent chromosome arms during the pre-meiotic S phase. Centromeric Rec8 persists throughout meiosis I and disappears at anaphase of meiosis II. When the rec8 gene is deleted, sister chromatids separate at meiosis I, resulting in equational rather than reductional chromosome segregation. We propose that the persistence of Rec8 at centromeres during meiosis I maintains sister-chromatid cohesion, and that its presence in the centromere-adjacent regions orients the kinetochores so that sister chromatids move to the same pole. This results in the reductional pattern of chromosome segregation necessary to reduce a diploid zygote to haploid gametes.     

Direct observation of microtubule dynamics in living cells

The study of cell locomotion is fundamental to such diverse processes as embryonic development, wound healing and metastasis. Since microtubules play a role in establishing the leading lamellum and maintaining cell polarity, it is important to understand their dynamic behaviour. In vitro, subunits exchange with polymer by treadmilling and by dynamic instability. Disassembly events can be complete (catastrophic) or incomplete (tempered). In vivo, microtubules are in dynamic equilibrium with subunits with a half-time for turnover of 4-20 min. Microtubules grow by elongation of their ends and are replaced one by one with turnover being most rapid at the periphery. Although previous results are consistent with dynamic instability, we sought to directly test the mechanism of turnover. Direct observations of fluorescent microtubules in the fibroblast lamellum show that individual microtubules undergo rounds of assembly and disassembly from the same end. Reorganization of the microtubule network occurs by a tempered mode of dynamic instability.     

The dynamics of chromosome evolution in birds and mammals

Comparative mapping, which compares the location of homologous genes in different species, is a powerful tool for studying genome evolution. Comparative maps suggest that rates of chromosomal change in mammals can vary from one to ten rearrangements per million years. On the basis of these rates we would expect 84 to 600 conserved segments in a chicken comparison with human or mouse. Here we build comparative maps between these species and estimate that numbers of conserved segments are in the lower part of this range. We conclude that the organization of the human genome is closer to that of the chicken than the mouse and by adding comparative mapping results from a range of vertebrates, we identify three possible phases of chromosome evolution. The relative stability of genomes such as those of the chicken and human will enable the reconstruction of maps of ancestral vertebrates.     

模拟失重对拟南芥下胚轴细胞生长和微管动态性的影响

空间飞行中产生的失重环境会影响植物的一系列生命活动过程,因此,为保证植物正常生长,揭示失重环境下植物的生长发育规律至关重要。利用大梯度强磁场超导磁体产生的抗磁悬浮模拟失重环境,对拟南芥幼苗进行处理。在相同外界条件下培养4 d后,拍照记录不同重力环境下拟南芥下胚轴细胞的生长发育及其周质微管骨架的排列情况,并进行图像定量分析和统计分析。结果显示,与对照组相比,重力改变对幼苗的形态、株高、下胚轴细胞形态及其周质微管骨架的排列有不同程度的影响。尤其是在μg(模拟失重),幼苗弯曲明显,株高比其他组显著降低,细胞长宽比较小,下胚轴周质微管发生了重排。而对照组、1g组、2g组(模拟超重)三组之间相比显示,除了在微管骨架排列方面存在极显著差异,其余指标均无明显差异。结果提示,模拟失重对植物生长发育有显著影响,微管骨架参与了重力信号的传感和转导,推测微管骨架可能通过其重排调控下胚轴细胞生长,参与模拟失重抑制下胚轴伸长来影响植株形态建成。     

In vitro effects on microtubule dynamics of purified Xenopus M phase-activated MAP kinase.

The protein kinase MAP kinase, also called MAP2 kinase, is a serine/threonine kinase whose activation and phosphorylation are induced by a variety of mitogens, and which is thought to have a critical role in a network of protein kinases in mitogenic signal transduction. A burst in kinase activation and protein phosphorylation may also be important in triggering the dramatic reorganization of the cell during the transition from interphase to mitosis. The interphase-metaphase transition of microtubule arrays is under the control of p34cdc2 kinase, a central control element in the G2-M transition of the cell cycle. Here we show that a Xenopus kinase, closely related to the mitogen-activated mammalian MAP kinase, is phosphorylated and activated during M phase of meiotic and mitotic cell cycles, and that the interphase-metaphase transition of microtubule arrays can be induced by the addition of purified Xenopus M phase-activated MAP kinase or mammalian mitogen-activated MAP kinase to interphase extracts in vitro.     

Regulation of microtubule dynamics by cdc2 protein kinase in cell-free extracts of Xenopus eggs

Microtubules are involved in the transport of vesicles in interphase and of the chromosomes during mitosis. Their arrangement and orientation in the cell are therefore of prime importance and specific patterns are believed to be generated by modulations of the intrinsic dynamic instability of microtubules. Here it is shown that the interphase-metaphase transition of microtubule arrays is under the control of the cdc2 kinase that precisely regulates the dynamics and steady-state length of microtubules.     

North American ice-sheet dynamics and the onset of 100,000-year glacial cycles

The onset of major glaciations in the Northern Hemisphere about 2.7 million years ago was most probably induced by climate cooling during the late Pliocene epoch. These glaciations, during which the Northern Hemisphere ice sheets successively expanded and retreated, are superimposed on this long-term climate trend, and have been linked to variations in the Earth's orbital parameters. One intriguing problem associated with orbitally driven glacial cycles is the transition from 41,000-year to 100,000-year climatic cycles that occurred without an apparent change in insolation forcing. Several hypotheses have been proposed to explain the transition, both including and excluding ice-sheet dynamics. Difficulties in finding a conclusive answer to this palaeoclimatic problem are related to the lack of sufficiently long records of ice-sheet volume or sea level. Here we use a comprehensive ice-sheet model and a simple ocean-temperature model to extract three-million-year mutually consistent records of surface air temperature, ice volume and sea level from marine benthic oxygen isotopes. Although these records and their relative phasings are subject to considerable uncertainty owing to limited availability of palaeoclimate constraints, the results suggest that the gradual emergence of the 100,000-year cycles can be attributed to the increased ability of the merged North American ice sheets to survive insolation maxima and reach continental-scale size. The oversized, wet-based ice sheet probably responded to the subsequent insolation maximum by rapid thinning through increased basal-sliding, thereby initiating a glacial termination. Based on our assessment of the temporal changes in air temperature and ice volume during individual glacials, we demonstrate the importance of ice dynamics and ice-climate interactions in establishing the 100,000-year glacial cycles, with enhanced North American ice-sheet growth and the subsequent merging of the ice sheets being key elements.     

Observation of rare-earth segregation in silicon nitride ceramics at subnanometre dimensions

Silicon nitride (Si3N4) ceramics are used in numerous applications because of their superior mechanical properties. Their intrinsically brittle nature is a critical issue, but can be overcome by introducing whisker-like microstructural features. However, the formation of such anisotropic grains is very sensitive to the type of cations used as the sintering additives. Understanding the origin of dopant effects, central to the design of high-performance Si3N4 ceramics, has been sought for many years. Here we show direct images of dopant atoms (La) within the nanometre-scale intergranular amorphous films typically found at grain boundaries, using aberration corrected Z-contrast scanning transmission electron microscopy. It is clearly shown that the La atoms preferentially segregate to the amorphous/crystal interfaces. First-principles calculations confirm the strong preference of La for the crystalline surfaces, which is essential for forming elongated grains and a toughened microstructure. Whereas principles of micrometre-scale structural design are currently used to improve the mechanical properties of ceramics, this work represents a step towards the atomic-level structural engineering required for the next generation of ceramics.     

Mediation of meiotic and early mitotic chromosome segregation in Drosophila by a protein related to kinesin.

Contrary to the traditional view that microtubules pull chromosomes polewards during the anaphase stage of meiotic and mitotic cell divisions, new evidence suggests that the chromosome movements are driven by a motor located at the kinetochore. The process of chromosome segregation involves proper arrangement of kinetochores for spindle attachment, followed by spindle attachment and chromosome movement. Mechanisms in Drosophila for chromosome segregation in meiosis differ in males and females, implying the action of different gene products in the two sexes. A product encoded at the claret locus in Drosophila is required for normal chromosome segregation in meiosis in females and in early mitotic divisions of the embryo. Here we show that the predicted amino-acid sequence of this product is related to the heavy chain of kinesin. The conserved region corresponds to the kinesin motor domain and includes the ATP-binding site and a region that can bind microtubules. A second region contains a leucine repeat motif which may mediate protein-subunit interactions necessary for attachment of chromosomes to the spindle. The mutant phenotype of chromosome nondisjunction and loss, and its similarity to the kinesin ATP-binding domain, suggest that the product encoded at claret not only stabilizes chromosome attachments to the spindle, but may also be a motor that drives chromosome segregation in female meiosis.     

Assembly dynamics of microtubules at molecular resolution

Microtubules are highly dynamic protein polymers that form a crucial part of the cytoskeleton in all eukaryotic cells. Although microtubules are known to self-assemble from tubulin dimers, information on the assembly dynamics of microtubules has been limited, both in vitro and in vivo, to measurements of average growth and shrinkage rates over several thousands of tubulin subunits. As a result there is a lack of information on the sequence of molecular events that leads to the growth and shrinkage of microtubule ends. Here we use optical tweezers to observe the assembly dynamics of individual microtubules at molecular resolution. We find that microtubules can increase their overall length almost instantaneously by amounts exceeding the size of individual dimers (8 nm). When the microtubule-associated protein XMAP215 (ref. 6) is added, this effect is markedly enhanced and fast increases in length of about 40-60 nm are observed. These observations suggest that small tubulin oligomers are able to add directly to growing microtubules and that XMAP215 speeds up microtubule growth by facilitating the addition of long oligomers. The achievement of molecular resolution on the microtubule assembly process opens the way to direct studies of the molecular mechanism by which the many recently discovered microtubule end-binding proteins regulate microtubule dynamics in living cells.     

纳米尺度下对黏滑现象的分子动力学模拟

采用分子动力学模拟的方法研究了硅针尖在金刚石基底滑动的黏滑现象,讨论了纳米尺度下温度、滑动速度、载荷等因素对黏滑摩擦的影响.模拟结果表明,在纳米尺度下,原子排列规则的两固体间的滑动摩擦力呈现出周期性的锯齿型变化.摩擦力曲线的波动周期取决于滑动过程中基体沿滑动方向的晶格常数,同时受接触面原子排列结构变化的影响.在较低温度范围内,滑动摩擦力随温度的升高近似呈线性减小,对滑动摩擦力的波动周期和振幅影响不大.在一定速度范围内,滑动摩擦力主要受黏着作用的影响,滑动摩擦力的大小随速度的增大近似呈线性增大.在一定载荷范围内,滑动摩擦力随载荷的增大近似呈线性增大,振动周期变大.     

Grain boundary segregation of minor arsenic and nitrogen at elevated temperatures in a microalloyed steel

Auger electron spectroscopy (AES) was used to investigate the grain boundary segregation of arsenic and nitrogen in a kind of microalloyed steel produced by a compact strip production (CSP) technology at 950 to 1100℃, which are similar to the hot working temperature of the steel on a CSP production line. It was discovered that arsenic segregated on grain boundaries when the steel was annealed at 950℃ for 2 h. When the annealing temperature increased to 1100℃, arsenic was also found to have segregated on grain boundaries in the early annealing stage, for instance, within the first 5 min annealing time. However, if the holding time of the steel at this temperature increased to 2 h, arsenic diffused away from grain boundaries into the matrix again. Nitrogen was not found to have segregated on grain boundaries when the steel was annealed at a relatively low temperature, such as 950℃. But when the annealing temperature increased to 1100℃, nitrogen was detected to have segregated at grain boundaries in the steel.     

二冷区电磁搅拌对连铸板坯中心偏析的影响

通过对板坯逐层刨钢屑,采用化学分析和低倍组织检验的方法,研究了板坯连铸机二冷区电磁搅拌频率参数对1 450 mm×230 mm板坯中心偏析和等轴晶率的影响. 结果表明:单独采用二冷区电磁搅拌相比不采用电磁搅拌工艺的板坯中心偏析程度明显减轻,C、P、Mn的中心正偏析系数低于无电磁搅拌工艺的板坯;电磁搅拌频率为2 Hz,5 Hz和8 Hz时板坯中心偏析都为B0.5级;电磁搅拌频率对中心等轴晶率略有影响,搅拌频率5 Hz的情况下等轴晶率最大.     

MoSi_2低温氧化动力学研究

通过热重量分析法测定了MoSi2 在低温下氧化不同时间后的重量变化 ,并根据曲线拟合分析了MoSi2 的低温氧化过程 结果表明 :在 5 0 0℃时MoSi2 材料的氧化过程受扩散和反应双重影响 ,氧化速率呈阶段式变化 :先主要受扩散控制 ,氧化增重与时间呈指数关系 ;随后反应成为主要控制因素 ,氧化增重与时间呈线性关系 ;如此反复 ,表现出比 40 0℃和 6 0 0℃时快得多的氧化速率 在 40 0℃、6 0 0℃时MoSi2 的氧化速率主要取决于氧化反应 ,且与反应进行的时间呈线性关系 ,氧化 14 4h后因钝化质量维持不变 图 3 ,表 1,参 7     

Large subglacial lakes in East Antarctica at the onset of fast-flowing ice streams

Water plays a crucial role in ice-sheet stability and the onset of ice streams. Subglacial lake water moves between lakes and rapidly drains, causing catastrophic floods. The exact mechanisms by which subglacial lakes influence ice-sheet dynamics are unknown, however, and large subglacial lakes have not been closely associated with rapidly flowing ice streams. Here we use satellite imagery and ice-surface elevations to identify a region of subglacial lakes, similar in total area to Lake Vostok, at the onset region of the Recovery Glacier ice stream in East Antarctica and predicted by ice-sheet models. We define four lakes through extensive, flat, featureless regions of ice surface bounded by upstream troughs and downstream ridges. Using ice velocities determined using interferometric synthetic aperture radar (InSAR), we find the onset of rapid flow (moving at 20 to 30 m yr(-1)) of the tributaries to the Recovery Glacier ice stream in a 280-km-wide segment at the downslope margins of these four subglacial lakes. We conclude that the subglacial lakes initiate and maintain rapid ice flow through either active modification of the basal thermal regime of the ice sheet by lake accretion or through scouring bedrock channels in periodic drainage events. We suggest that the role of subglacial lakes needs to be considered in ice-sheet mass balance assessments.     

Cascades of multisite phosphorylation control Sic1 destruction at the onset of S phase

Multisite phosphorylation of proteins has been proposed to transform a graded protein kinase signal into an ultrasensitive switch-like response. Although many multiphosphorylated targets have been identified, the dynamics and sequence of individual phosphorylation events within the multisite phosphorylation process have never been thoroughly studied. In Saccharomyces cerevisiae, the initiation of S phase is thought to be governed by complexes of Cdk1 and Cln cyclins that phosphorylate six or more sites on the Clb5-Cdk1 inhibitor Sic1, directing it to SCF-mediated destruction. The resulting Sic1-free Clb5-Cdk1 complex triggers S phase. Here, we demonstrate that Sic1 destruction depends on a more complex process in which both Cln2-Cdk1 and Clb5-Cdk1 act in processive multiphosphorylation cascades leading to the phosphorylation of a small number of specific phosphodegrons. The routes of these phosphorylation cascades are shaped by precisely oriented docking interactions mediated by cyclin-specific docking motifs in Sic1 and by Cks1, the phospho-adaptor subunit of Cdk1. Our results indicate that Clb5-Cdk1-dependent phosphorylation generates positive feedback that is required for switch-like Sic1 destruction. Our evidence for a docking network within clusters of phosphorylation sites uncovers a new level of complexity in Cdk1-dependent regulation of cell cycle transitions, and has general implications for the regulation of cellular processes by multisite phosphorylation.