Recruitment and regulation of phosphatidylinositol phosphate kinase type 1 gamma by the FERM domain of talin

Membrane phosphoinositides control a variety of cellular processes through the recruitment and/or regulation of cytosolic proteins. One mechanism ensuring spatial specificity in phosphoinositide signalling is the targeting of enzymes that mediate their metabolism to specific subcellular sites. Phosphatidylinositol phosphate kinase type 1 gamma (PtdInsPKI gamma) is a phosphatidylinositol-4-phosphate 5-kinase that is expressed at high levels in brain, and is concentrated at synapses. Here we show that the predominant brain splice variant of PtdInsPKI gamma (PtdInsPKI gamma-90) binds, by means of a short carboxy-terminal peptide, to the FERM domain of talin, and is strongly activated by this interaction. Talin, a principal component of focal adhesion plaques, is also present at synapses. PtdInsPKI gamma-90 is expressed in non-neuronal cells, albeit at much lower levels than in neurons, and is concentrated at focal adhesion plaques, where phosphatidylinositol-4,5-bisphosphate has an important regulatory role. Overexpression of PtdInsPKI gamma-90, or expression of its C-terminal domain, disrupts focal adhesion plaques, probably by local disruption of normal phosphoinositide balance. These findings define an interaction that has a regulatory role in cell adhesion and suggest new similarities between molecular interactions underlying synaptic junctions and general mechanisms of cell adhesion.     

An interaction between vinculin and talin

In cultured fibroblasts, microfilament bundles terminate at adhesion plaques (focal contacts), the specialized regions where the cells adhere most tightly to the underlying substrate. Vinculin is a protein concentrated in adhesion plaques and has been suggested as a possible link between the ends of the bundles of actin filaments and the plasma membrane. If vinculin is one protein in a chain of attachment between the bundles of microfilaments and the plasma membrane, it is important to identify other components which interact with vinculin. We have recently discovered a new protein in adhesion plaques which we refer to as talin. Here we show that talin binds to vinculin, which suggests that talin may be involved with vinculin in the attachment of microfilament bundles to the plasma membrane at the adhesion plaques.     

平均数构造的数列性质

对任意给出的m个正实数,通过连续计算其去掉一个实数后所得数组的算术平均数,得到新的m个无穷数列.讨论了这m个无穷数列的性质,得出这m个无穷数列都收敛于初始数组的算术平均数,并将结论拓展到了通过计算几何平均数、调和平均数、平方平均数所得到的数列的情形.得出结论:对任意一个数组,连续计算去掉一个数据后所得数组的平均数,其数字特征不会发生变化,并且如果从每次计算平均数所得的数组中任取一个数据构成无穷子数列,必定收敛于相应的数字特征.     

平均数构造的数列性质

对任意给出的m个正实数,通过连续计算其去掉一个实数后所得数组的算术平均数,得到新的m个无穷数列。讨论了这m个无穷数列的性质,得出这m个无穷数列都收敛于初始数组的算术平均数,并将结论拓展到了通过计算几何平均数、调和平均数、平方平均数所得到的数列的情形。得出结论:对任意一个数组,连续计算去掉一个数据后所得数组的平均数,其数字特征不会发生变化,并且如果从每次计算平均数所得的数组中任取一个数据构成无穷子数列,必定收敛于相应的数字特征。     

Sequence of reovirus haemagglutinin predicts a coiled-coil structure

The use of modern techniques has led to new insights into the molecular mechanisms of viral pathogenesis. Although the infectious process is quite complex, it is clear that one critical stage, the interaction of viral attachment proteins with cell-surface receptors, often has a major role in determining the pattern of infection. The mammalian reoviruses have served as useful models for understanding the molecular basis of viral pathogenesis. The mammalian reovirus haemagglutinin (sigma 1 protein), which is an outer capsid protein, has been shown to be a major factor in determining virus-host cell interactions. To further our understanding of the structure and function of the haemagglutinin, we have cloned a complementary DNA copy of the reovirus type 3 S1 double-stranded RNA gene which encodes the virus haemagglutinin and have sequenced the DNA complementary to the S1 gene. Analysis of the predicted amino-acid sequence of the virus haemagglutinin has allowed us to determine that the amino-terminal portion contains an alpha-helical coiled-coil structure and that the carboxy-terminal portion contains the receptor-interacting domains. Using this information, we propose here a model of how the reovirus haemagglutinin is attached to the virus particle.     

Vinculin activation by talin through helical bundle conversion

Vinculin is a conserved component and an essential regulator of both cell-cell (cadherin-mediated) and cell-matrix (integrin-talin-mediated focal adhesions) junctions, and it anchors these adhesion complexes to the actin cytoskeleton by binding to talin in integrin complexes or to alpha-actinin in cadherin junctions. In its resting state, vinculin is held in a closed conformation through interactions between its head (Vh) and tail (Vt) domains. The binding of vinculin to focal adhesions requires its association with talin. Here we report the crystal structures of human vinculin in its inactive and talin-activated states. Talin binding induces marked conformational changes in Vh, creating a novel helical bundle structure, and this alteration actively displaces Vt from Vh. These results, as well as the ability of alpha-actinin to also bind to Vh and displace Vt from pre-existing Vh-Vt complexes, support a model whereby Vh functions as a domain that undergoes marked structural changes that allow vinculin to direct cytoskeletal assembly in focal adhesions and adherens junctions. Notably, talin's effects on Vh structure establish helical bundle conversion as a signalling mechanism by which proteins direct cellular responses.     

The cargo-binding domain regulates structure and activity of myosin 5

Myosin 5 is a two-headed motor protein that moves cargoes along actin filaments. Its tail ends in paired globular tail domains (GTDs) thought to bind cargo. At nanomolar calcium levels, actin-activated ATPase is low and the molecule is folded. Micromolar calcium concentrations activate ATPase and the molecule unfolds. Here we describe the structure of folded myosin and the GTD's role in regulating activity. Electron microscopy shows that the two heads lie either side of the tail, contacting the GTDs at a lobe of the motor domain (approximately Pro 117-Pro 137) that contains conserved acidic side chains, suggesting ionic interactions between motor domain and GTD. Myosin 5 heavy meromyosin, a constitutively active fragment lacking the GTDs, is inhibited and folded by a dimeric GST-GTD fusion protein. Motility assays reveal that at nanomolar calcium levels heavy meromyosin moves robustly on actin filaments whereas few myosins bind or move. These results combine to show that with no cargo, the GTDs bind in an intramolecular manner to the motor domains, producing an inhibited and compact structure that binds weakly to actin and allows the molecule to recycle towards new cargoes.     

抽象论域到数据结构的转换

给出了函数式语言到过程式语言转换的关键技术,即抽象论域到具体数据结构的转换技术.抽象论域的转换是抽象表达式到具体表达式转换、模式匹配处理的前提.函数式语言到过程式语言的转换,解决了函数式语言在具体实现和时间方面存在的大量动态函数复制的问题.这种转换技术可用于开发指称语义到解释器、指称语义到编译器、属性文法到过程式语言、属性文法到YACC(Yet Another Computer Compilier)输入文件的各种自动生成器.     

Nir2碳端结构域的晶体结构

Nir2蛋白具有多个结构域，在生物体中可能发挥多种生理功能。本实验用MAD软件对大肠杆菌BL21（plys）原核表达的Nir2蛋白碳端进行了晶体结构分析。结果显示，硒蛋氨酸取代蛋白晶体及未取代蛋白晶体同属空间群P212121,晶胞参数a=109.468 Å, b=120.621 Å, c=70.315 Å，分辨率分别为2.2 Å和2.7 Å。一个不对称单位含有三个分子，含水量为48%。Nir2蛋白碳端由C片层和N片层构成，其中C片层与钙ATPase的催化区域P相似，N片层为许多不相关的功能性蛋白所共有。研究证明二者之间的静电作用是影响Nir2蛋白碳端与Pyk2 FERM区域结合的主要因素。该晶体结构的分析结果为进一步研究Nir2蛋白碳端的潜在功能提供了结构基础。     

Two-piconewton slip bond between fibronectin and the cytoskeleton depends on talin

Mechanical forces on matrix-integrin-cytoskeleton linkages are crucial for cell viability, morphology and organ function. The production of force depends on the molecular connections from extracellular-matrix-integrin complexes to the cytoskeleton. The minimal matrix complex causing integrin-cytoskeleton connections is a trimer of fibronectin's integrin-binding domain FNIII7-10 (ref. 4). Here we report a specific, molecular slip bond that was broken repeatedly by a force of 2 pN at the cellular loading rate of 60 nm x s(-1); this occurred with single trimer beads but not with monomer. Talin1, which binds to both integrins and actin filaments in vitro, is required for the 2-pN slip bond and rapid cytoskeleton binding. Further, inhibition of fibronectin binding to alpha(v)beta3 and deletion of beta3 markedly decreases the 2-pN force peak. We suggest that talin1 initially forms a molecular slip bond between closely packed fibronectin-integrin complexes and the actin cytoskeleton, which can apply a low level of force to fibronectin until many bonds form or a signal is received to activate a force response.     

Crystal structure of the specificity domain of ribonuclease P

RNase P is the only endonuclease responsible for processing the 5' end of transfer RNA by cleaving a precursor and leading to tRNA maturation. It contains an RNA component and a protein component and has been identified in all organisms. It was one of the first catalytic RNAs identified and the first that acts as a multiple-turnover enzyme in vivo. RNase P and the ribosome are so far the only two ribozymes known to be conserved in all kingdoms of life. The RNA component of bacterial RNase P can catalyse pre-tRNA cleavage in the absence of the RNase P protein in vitro and consists of two domains: a specificity domain and a catalytic domain. Here we report a 3.15-A resolution crystal structure of the 154-nucleotide specificity domain of Bacillus subtilis RNase P. The structure reveals the architecture of this domain, the interactions that maintain the overall fold of the molecule, a large non-helical but well-structured module that is conserved in all RNase P RNA, and the regions that are involved in interactions with the substrate.     

软件过程领域本体的构造

针对目前软件过程领域知识缺乏明确统一的表示、不同组织构造的过程模型缺少互操作性而难以共享和重用的情况,划分并描述了软件过程的顶层本体;在此基础上,对其中包含的任务描述、过程模型和过程实施与改进三个核心本体分别进行了展开和细化,给出了涉及到的关键概念的形式化定义.传统的本体系统并不适宜描述动态过程,因此使用谓词逻辑定义了过程本体中概念间的基本关系,以此来表示和描述软件过程模型的“柔性”和动态知识,使不同的参与者易于交流而达成共识,为构造可共享、易重用的过程模型元模型提供坚实、统一的基础.     

焦耳热调制CoFe基微丝的GMI与畴结构相关性分析

采用常规焦耳热退火调制熔体抽拉Co68.15Fe4.35Si12.25B13.75Nb1Cu0.5非晶微丝的巨磁阻抗效应,并观测不同电流幅值(0 mA～100 mA)退火后微丝表面磁畴结构,分析其磁阻抗性能与畴结构的相关性,以此提出表征微丝GMI效应的一种方法.研究结果表明:随着退火电流幅值的增大,磁阻抗效应比值呈现先增大后减少的变化规律.制备态时,微丝内部残余较大径向应力,表面周向畴较弱,磁阻抗效应较低.80 mA焦耳热退火后,其表面周向畴规则分布,阻抗比值[Z/Z0]max(％)值达到114.0％,对应的场响应灵敏度为127.7％/0e;100 mA焦耳热退火后,大量焦耳热促使微丝内部局域晶化较明显,其磁畴壁出现钉扎现象,周向畴被破坏,阻抗性能降低.     

Sequence and structure of D-glyceraldehyde 3-phosphate dehydrogenase from Bacillus stearothermophilus.

The glyceraldehyde 3-phosphate dehydogenase holoenzyme of Bacillus stearothermophilus possesses precise 222 symmetry: in this respect it differs from the reported structure of the lobster muscle enzyme. Pairs of active sites are linked through a flexible polypeptide loop which probably mediates the structural changes giving rise to cooperative effects. Three additional salt bridges made by each subunit to others would make a major contribution to thermostability of the tetramer.     

High-resolution structure of a retroviral capsid hexameric amino-terminal domain

Retroviruses are the aetiological agents of a range of human diseases including AIDS and T-cell leukaemias. They follow complex life cycles, which are still only partly understood at the molecular level. Maturation of newly formed retroviral particles is an essential step in production of infectious virions, and requires proteolytic cleavage of Gag polyproteins in the immature particle to form the matrix, capsid and nucleocapsid proteins present in the mature virion. Capsid proteins associate to form a dense viral core that may be spherical, cylindrical or conical depending on the genus of the virus. Nonetheless, these assemblies all appear to be composed of a lattice formed from hexagonal rings, each containing six capsid monomers. Here, we describe the X-ray structure of an individual hexagonal assembly from N-tropic murine leukaemia virus (N-MLV). The interface between capsid monomers is generally polar, consistent with weak interactions within the hexamer. Similar architectures are probably crucial for the regulation of capsid assembly and disassembly in all retroviruses. Together, these observations provide new insights into retroviral uncoating and how cellular restriction factors may interfere with viral replication.     

挠性结构模型的频域极大似然法辨识

在构建的以压电陶瓷为执行器、电阻式应变片为传感器的挠性悬臂梁物理实验系统基础上,优化设计多正弦辨识输入信号,利用频域极大似然法辨识得到了整个系统的数学模型,并根据该模型进行了数字和物理控制仿真,验证了模型的有效性.良好的控制效果表明:对于挠性结构,频域极大似然法是一种有效的模型辨识方法.     

Identification of talin as a major cytoplasmic protein implicated in platelet activation

During platelet activation there is a major reorganization in the platelet cytoskeleton that accompanies a rapid change in platelet shape. Many of the events associated with activation are attributed to a rise in calcium concentration within the platelet cytoplasm. One direct consequence of the elevated calcium is the activation of a calcium-dependent protease that cleaves a major platelet protein of relative molecular mass (Mr) approximately 235,000 (235K) to 200K. This protein, P235, has been purified and reported to interact with actin, but the significance of the proteolytic cleavage is unknown. Talin, a cytoskeletal protein in smooth muscle and fibroblasts, binds vinculin and, together with vinculin, is localized in fibroblasts at sites of actin-membrane attachment. Talin and P235 have similar purification procedures, sedimentation coefficients and Stokes' radii (ref. 6 and Molony et al., unpublished observations). Of particular significance, talin is readily cleaved by proteases from approximately 215K to a fragment of approximately 190K. Given these similarities we have investigated the possible relationship between these proteins. Here we demonstrate that platelet P235 is recognized by anti-talin antibody and that it binds vinculin. Both proteins are cleaved in vitro by the calcium-activated protease to yield similar fragments. We conclude that P235 corresponds to the platelet form of talin.     

Cloning and domain structure of the mammalian splicing factor U2AF.

A complementary DNA clone encoding the large subunit of the essential mammalian pre-messenger RNA splicing component U2 snRNP auxiliary factor (U2AF65) has been isolated and expressed in vitro. It contains two functional domains: a sequence-specific RNA-binding region composed of three ribonucleoprotein-consensus sequence domains, and an arginine/serine-rich motif necessary for splicing but not for binding to pre-mRNA.