细胞骨架在植物细胞周期进程中的动态变化及其调控?

细胞周期,即细胞生长与分裂的周期,是生命得以世代繁衍而生生不息的基础.真核细胞有丝分裂周期进程调控的分子机制高度保守.其间,微管和微丝骨架进行有规律的动态变化,顺次组成各种细胞生长和分裂装置,主动参与细胞周期进程的调节.然而,高等植物细胞周期不同时相分别有着与动物细胞不完全相同的、独特的细胞骨架列阵.而这些列阵的产生和维持直接依赖于众多细胞骨架结合蛋白以及上游信号分子的调控.本文重点综述了植物细胞周期进程中微管和微丝骨架的动态变化规律以及参与植物细胞骨架动态和有丝分裂装置组装调控的细胞骨架结合蛋白的最新研究进展,同时对细胞骨架在植物细胞周期进程中研究进行总结和展望.     

小鼠腹水瘤巨噬细胞对大肠杆菌O157免疫应答的膜蛋白质组学研究

采用膜亚蛋白质组学方法研究小鼠腹水瘤巨噬细胞Raw 264.7对致病性大肠杆菌O157的免疫应答反应,筛选并鉴定到24个差异表达的膜蛋白质.这些膜蛋白质与细胞免疫、细胞周期、细胞发育、细胞骨架、细胞生长等有关.在蛋白质水平上对巨噬细胞与大肠杆菌间的相互作用关系进行了一些探索性研究,为进一步揭示机体对病原反应的作用机制提供理论基础.     

Proteomic characterization of the human centrosome by protein correlation profiling

The centrosome is the major microtubule-organizing centre of animal cells and through its influence on the cytoskeleton is involved in cell shape, polarity and motility. It also has a crucial function in cell division because it determines the poles of the mitotic spindle that segregates duplicated chromosomes between dividing cells. Despite the importance of this organelle to cell biology and more than 100 years of study, many aspects of its function remain enigmatic and its structure and composition are still largely unknown. We performed a mass-spectrometry-based proteomic analysis of human centrosomes in the interphase of the cell cycle by quantitatively profiling hundreds of proteins across several centrifugation fractions. True centrosomal proteins were revealed by both correlation with already known centrosomal proteins and in vivo localization. We identified and validated 23 novel components and identified 41 likely candidates as well as the vast majority of the known centrosomal proteins in a large background of nonspecific proteins. Protein correlation profiling permits the analysis of any multiprotein complex that can be enriched by fractionation but not purified to homogeneity.     

KAx-3细胞和AK127细胞发育期间细胞骨架蛋白组分比较研究

采用生化抽提骨架蛋白和SDS-聚丙烯酰胺凝胶电泳等方法比较盘基网柄菌AK127细胞和KAx-3细胞骨架蛋白的结果显示，发育各时期的KAX-3和AK127细胞骨架蛋白在含量和组分方面存在一定的差异.在整个发育阶段两种类型细胞共有的蛋白组分是103.4kD、49.6kD、44.2kD、30.8kD、19.8kD和13.5kD条带，其中44.2kD和30.8kD是最稳定的细胞骨架成分，并不因AK127细胞缺失gp150分子而有所改变.它们与其他骨架蛋白组分一起为细胞提供有力的支持，保证发育的顺利进行.差别最为明显的蛋白条带是87.2kD、 68.2kD和40.7kD蛋白，由于同时期的突变型细胞不能显示这些蛋白条带；说明这些条带是发育所需的蛋白.值得注意的是突变型细胞也出现一条特征性的21.1kD的蛋白条带.分析认为这些变化一方面与盘基网柄菌发育过程有关，另一方面与突变细胞缺乏gp150分子有密切关系.     

白藜芦醇诱导HepG2细胞凋亡中线粒体差异蛋白鉴定

为研究线粒体在白藜芦醇诱导人肝癌细胞系HepG2细胞凋亡中的作用机制。分离提取白藜芦醇处理的HepG2细胞及对照组细胞的线粒体蛋白质,双向电泳分离差异蛋白质,飞行时间质谱鉴定差异蛋白点,摸索并建立了一种有效分离提取细胞线粒体蛋白质和双向电泳的方法。初步分析鉴定了四个显著性差异蛋白,着丝粒蛋白Kinesin protein和CENP-E降低,证明白藜芦醇对细胞周期及细胞骨架的调节作用,Peptidase (mitochondrial processing)表达降低,线粒体核糖体蛋白L7/L12（Mitochondrial ribosomal protein L7/L12, MRP L7/L12）表达升高,表明白藜芦醇诱导HepG2细胞与其对线粒体功能的影响有关。     

Homology of a yeast actin-binding protein to signal transduction proteins and myosin-I

In yeast, the cortical actin cytoskeleton seems to specify sites of growth of the cell surface. Because the actin-binding protein ABP1p is associated with the cortical cytoskeleton of Saccharomyces cerevisiae, it might be involved in the spatial organization of cell surface growth. ABP1p is localized to the cortical cytoskeleton and its overproduction causes assembly of the cortical actin cytoskeleton at inappropriate sites on the cell surface, resulting in delocalized surface growth. We have now cloned and sequenced the gene encoding ABP1p. ABP1p is a novel protein with a 50 amino-acid C-terminal domain that is very similar to the SH3 domain in the non-catalytic region of nonreceptor tyrosine kinases (including those encoded by the proto-oncogenes c-src and c-abl), in phospholipase C gamma and in alpha-spectrin. We also identified an SH3-related motif in the actin-binding tail domain of myosin-I. The identification of SH3 domains in a family of otherwise unrelated proteins that associate with the membrane cytoskeleton indicates that this domain might serve to bring together signal transduction proteins and their targets or regulators, or both, in the membrane cytoskeleton.     

Formins： Bringing new insights to the organization of actin cytoskeleton

The actin cytoskeleton is an important component of eukaryotic cell cytoskeleton and is temporally and spatially controlled by a series of actin binding proteins (ABPs). Among ABPs, formin family proteins have attracted much attention as they can nucleate unbranched actin filament from the profilin bound actin pool in vivo. In recent years, a number of formin family members from different organisms have been reported, and their characteristics are known more clearly, although some questions are still to be clarified. Here, we summarize the structures, func-tions and nucleation mechanisms of different formin family proteins, intending to compare them and give some new clues to the study of formins.     

纤毛虫皮层细胞骨架的研究进展

纤毛虫皮层的研究对探讨细胞的发育、分化、结构模式及其遗传控制有着极其重大的生物学意义,纤毛虫细胞皮层中含有非常丰富的细胞骨架成分,它们在结构上表现出来的特殊性有利于更深地探索皮层分化和皮层模式的控制机理,本综述了有关纤毛虫皮层细胞骨架的结构和形态发生方面的研究及皮层细胞骨架蛋白质构成的生化和分子生物学研究进展。     

Cdc42的生物活性

Cdc42是Rho家族蛋白(Rho GTPase)中的一种,是Rho家族中研究得较多的一个,具有GTP酶活性。Cdc42参与细胞周期调控和基因转录的调节,在调节细胞骨架、肿瘤表达、细胞极性等方面发挥重要作用。     

Diacylglycerol in large alpha-actinin/actin complexes and in the cytoskeleton of activated platelets

The interaction of the cytoskeleton with plasma membranes may be mediated by vinculin, alpha-actinin and other proteins; alpha-actinin can interact specifically with model membranes only if they contain diacylglycerol and palmitic acid. On stimulation of platelets by thrombin, which leads to a reorganization of the cytoskeleton, diacylglycerol is produced rapidly, simultaneously with the disappearance of phosphatidylinositol. One important function of the diacylglycerol produced in platelets may be the activation of the Ca2+-and phospholipid-dependent protein kinase C. We show here that, in the presence of diacylglycerol and palmitic acid, a supramolecular complex between alpha-actinin and actin is formed in vitro. In the electron microscope, this complex displays substructures similar to those of microfilament bundles in vivo. Furthermore, such alpha-actinin/lipid complexes can also be formed in situ during the stimulation of blood platelet aggregation. Thus, alpha-actinin may be one of the proteins directly involved in structures connecting the cytoskeleton to cell membranes.     

A yeast gene (BEM1) necessary for cell polarization whose product contains two SH3 domains.

Cell polarization requires that a cellular axis or cell-surface site be chosen and that the cytoskeleton be organized with respect to it. Details of the link between the cytoskeleton and the chosen axis or site are not clear. Cells of the yeast Saccharomyces cerevisiae exhibit cell polarization in two phases of their life cycle, during vegetative growth and during mating, which reflects responses to intracellular and extracellular signals, respectively. Here we describe the isolation of two mutants defective specifically in cell polarization in response to peptide mating pheromones. The mutants carry special alleles (denoted bem1-s) of the BEM1 gene required for cell polarization during vegetative growth. Unlike other bem1 mutants, the bem1-s mutants are normal for vegetative growth. Complete deletion of BEM1 leads to the defect in polarization of vegetative cells seen in bem1 mutants. The predicted sequence of the BEM1 protein (Bem1p) reveals two copies of a domain (denoted SH3) that is found in many proteins associated with the cortical cytoskeleton and which may mediate binding to actin or some other component of the cell cortex. The sequence of Bem1p and the properties of mutants defective in this protein indicate that it may link the cytoskeleton to morphogenetic determinants on the cell surface.     

影响洋葱表皮细胞骨架制片效果的探讨

以洋葱表皮细胞为材料,探究其经卡诺氏液固定后进行细胞骨架制片观察时,1%Triton X-100抽提非骨架蛋白的最佳时间,并研究洋葱不同表皮细胞及经不同理化因素处理的材料,经卡诺氏液固定后进行细胞骨架制片的步骤及制片效果。结果表明:经卡诺氏液固定后,1%Triton X-100抽提非骨架蛋白内表皮30min较佳,外表皮25min较佳;不同成熟度的鳞片叶细胞骨架形态及稳定性有差异;经紫外线、温度、秋水仙素处理后增加固定步骤再制片对制片观察结果基本无影响。     

Myosin V orientates the mitotic spindle in yeast

Coordination of spindle orientation with the axis of cell division is an essential process in all eukaryotes. In addition to ensuring accurate chromosomal segregation, proper spindle orientation also establishes differential cell fates and proper morphogenesis. In both animal and yeast cells, this process is dependent on cytoplasmic microtubules interacting with the cortical actin-based cytoskeleton, although the motive force was unknown. Here we show that yeast Myo2, a myosin V that translocates along polarized actin cables into the bud, orientates the spindle early in the cell cycle by binding and polarizing the microtubule-associated protein Kar9 (refs 7-9). The tail domain of Myo2 that binds Kar9 also interacts with secretory vesicles and vacuolar elements, making it a pivotal component of yeast cell polarization.     

Structure of the small G protein Cdc42 bound to the GTPase-binding domain of ACK.

The proteins Cdc42 and Rac are members of the Rho family of small GTPases (G proteins), which control signal-transduction pathways that lead to rearrangements of the cell cytoskeleton, cell differentiation and cell proliferation. They do so by binding to downstream effector proteins. Some of these, known as CRIB (for Cdc42/Rac interactive-binding) proteins, bind to both Cdc42 and Rac, such as the PAK1-3 serine/threonine kinases, whereas others are specific for Cdc42, such as the ACK tyrosine kinases and the Wiscott-Aldrich-syndrome proteins (WASPs). The effector loop of Cdc42 and Rac (comprising residues 30-40, also called switch I), is one of two regions which change conformation on exchange of GDP for GTP. This region is almost identical in Cdc42 and Racs, indicating that it does not determine the specificity of these G proteins. Here we report the solution structure of the complex of Cdc42 with the GTPase-binding domain ofACK. Both proteins undergo significant conformational changes on binding, to form a new type of G-protein/effector complex. The interaction extends the beta-sheet in Cdc42 by binding an extended strand from ACK, as seen in Ras/effector interactions, but it also involves other regions of the G protein that are important for determining the specificity of effector binding.     

Matrix proteins can generate the higher order architecture of the Golgi apparatus

The Golgi apparatus in animal cells comprises a reticulum of linked stacks in the pericentriolar and often in the juxtanuclear regions of the cell. The unique architecture of this organelle is thought to depend on the cytoskeleton and cytoplasmic matrix proteins--the best characterized being the golgin family of fibrous, coiled-coil proteins and the GRASP family of stacking proteins. Here we show that these matrix proteins can be separated from oligosaccharide-modifying enzymes in the Golgi stack without affecting their ability to form a ribbon-like reticulum in the correct location near to the nucleus. Our data suggest that the Golgi is a structural scaffold that can exist independently of, but is normally populated by, the enzyme-containing membranes that modify transiting cargo. This new concept of the Golgi further indicates that the Golgi may be an autonomous organelle rather than one that is in simple dynamic equilibrium with the endoplasmic reticulum.     

A kinase-regulated PDZ-domain interaction controls endocytic sorting of the beta2-adrenergic receptor.

A fundamental question in cell biology is how membrane proteins are sorted in the endocytic pathway. The sorting of internalized beta2-adrenergic receptors between recycling endosomes and lysosomes is responsible for opposite effects on signal transduction and is regulated by physiological stimuli. Here we describe a mechanism that controls this sorting operation, which is mediated by a family of conserved protein-interaction modules called PDZ domains. The phosphoprotein EBP50 (for ezrinradixin-moesin(ERM)-binding phosphoprotein-50) binds to the cytoplasmic tail of the beta2-adrenergic receptor through a PDZ domain and to the cortical actin cytoskeleton through an ERM-binding domain. Disrupting the interaction of EBP50 with either domain or depolymerization of the actin cytoskeleton itself causes missorting of endocytosed beta2-adrenergic receptors but does not affect the recycling of transferrin receptors. A serine residue at position 411 in the tail of the beta2-adrenergic receptor is a substrate for phosphorylation by GRK-5 (for G-protein-coupled-receptor kinase-5) and is required for interaction with EBP50 and for proper recycling of the receptor. Our results identify a new role for PDZ-domain-mediated protein interactions and for the actin cytoskeleton in endocytic sorting, and suggest a mechanism by which GRK-mediated phosphorylation could regulate membrane trafficking of G-protein-coupled receptors after endocytosis.     

人肝癌细胞骨架网络系统的初步研究

利用ＴｒｉｔｏｎＸ－１００及其联合（ＮＨ４）２ＳＯ４的抽提技术与ＣｏｏｍａｓｉｅｂｌｕｅＲ２５０染色、免疫酶标技术相结合，体外实验比较研究人肝癌细胞系（ＨｅｐＧ２）细胞骨架网络的分布构像及其中间纤维蛋白构型。结果：人肝癌细胞系（ＨｅｐＧ２）显示其细胞骨架网络的分布构像及其中间纤维对Ｖｉｍｅｎｔｉｎ、Ｋｅｒａｔｉｎ两种抗体均呈现阳性反应。结论：恶性肿瘤细胞的中间纤维蛋白构型可能具有异质性的蛋白分子共表达，这对仅以ＴｒｉｔｏｎＸ－１００加（ＮＨ４）２ＳＯ４的抽提技术进行恶性肿瘤细胞中间纤维蛋白构型分析，是一个必需考虑的问题。     

c-myc oncogene protein synthesis is independent of the cell cycle in human and avian cells

Several lines of evidence suggest a role for the myc oncogene in cell proliferation. Most recently, mitogenic stimulation of quiescent lymphoid, fibroblast and epithelial cells has been demonstrated to lead to a sharp increase in c-myc RNA levels. To determine how c-myc expression is linked to the cell proliferative cycle, we have used centrifugal elutriation to enrich for populations of avian and human cells at different stages of the cell cycle. Centrifugal elutriation is a counterflow centrifugation method that separates cells on the basis of volume, a parameter correlating well with progression through the cell cycle. Using myc-specific anti-peptide antibodies, we show here that the synthesis, half-life and modification of c-myc proteins are constant throughout the cell cycle of normal and transformed cells.     

The Par complex directs asymmetric cell division by phosphorylating the cytoskeletal protein Lgl

To generate different cell types, some cells can segregate protein determinants into one of their two daughter cells during mitosis. In Drosophila neuroblasts, the Par protein complex localizes apically and directs localization of the cell fate determinants Prospero and Numb and the adaptor proteins Miranda and Pon to the basal cell cortex, to ensure their segregation into the basal daughter cell. The Par protein complex has a conserved function in establishing cell polarity but how it directs proteins to the opposite side is unknown. We show here that a principal function of this complex is to phosphorylate the cytoskeletal protein Lethal (2) giant larvae (Lgl; also known as L(2)gl). Phosphorylation by Drosophila atypical protein kinase C (aPKC), a member of the Par protein complex, releases Lgl from its association with membranes and the actin cytoskeleton. Genetic and biochemical experiments show that Lgl phosphorylation prevents the localization of cell fate determinants to the apical cell cortex. Lgl promotes cortical localization of Miranda, and we propose that phosphorylation of Lgl by aPKC at the apical neuroblast cortex restricts Lgl activity and Miranda localization to the opposite, basal side of the cell.     

天然家蚕抗菌肽对人白细胞及巨噬细胞淋巴瘤细胞骨架系统作用的比较

报道了天然家蚕抗菌肽ＣＭ４对离体Ｕ９３７癌细胞骨架及核骨架损伤作用的扫描电镜观察。随着时间的延长，经天然家蚕抗菌肽ＣＭ４作用后的癌细胞骨架断裂，固缩成团状；癌细胞核骨架断裂，部分凝聚成团，结构不完整。相同剂量的天然家蚕抗菌肽ＣＭ４与正常人白细胞作用后细胞骨架及核骨架未见损伤现象。说明天然抗菌肽与重组抗菌肽的抗癌作用相同。