Cell-autonomous action of zebrafish spt-1 mutation in specific mesodermal precursors

In zebrafish, as in Xenopus, the well-orchestrated cell movements of gastrulation can be dissected into several components, including epiboly, involution, convergence and extension. Embryos homozygous for the recessive lethal mutation spt-1(b104) or 'spadetail' have a complex set of defects in the trunk of the embryo that may arise secondarily after loss of one of these movements, convergence, from those precursors that would normally have given rise to trunk somitic mesoderm. We have now tested this hypothesis by transplanting cells between wild-type and mutant embryos, to identify the cells that spt-1 affects directly. Our results show that the mutation autonomously affects only those mesodermal precursors located along the lateral margin of the early gastrula blastoderm. Other mesodermal cells and all ectodermal precursors seem not to require function of the wild-type gene. Our findings reveal an unexpectedly delicate genetic control of vertebrate gastrulation.     

Novel form of growth cone motility involving site-directed actin filament assembly.

Regulation of cytoskeletal structure and motility by extracellular signals is essential for all directed forms of cell movement and underlies the developmental process of axonal guidance in neuronal growth cones. Interaction with polycationic microbeads can trigger morphogenic changes in neurons and muscle cells normally associated with formation of pre- and postsynaptic specializations. Furthermore, when various types of microscopic particles are applied to the lamellar surface of a neuronal growth cone or motile cell they often exhibit retrograde movement at rates of 1-6 microns min-1 (refs 3-6). There is strong evidence that this form of particle movement results from translocation of membrane proteins associated with cortical F-actin networks, not from bulk retrograde lipid flow and may be a mechanism behind processes such as cell locomotion, growth cone migration and capping of cell-surface antigens. Here we report a new form of motility stimulated by polycationic bead interactions with the growth-cone membrane surface. Bead binding rapidly induces intracellular actin filament assembly, coincident with a production of force sufficient to drive bead movements. These extracellular bead movements resemble intracellular movements of bacterial parasites known to redirect host cell F-actin assembly for propulsion. Our results suggest that site-directed actin filament assembly may be a widespread cellular mechanism for generating force at membrane-cytoskeletal interfaces.     

Activin- and Nodal-related factors control antero-posterior patterning of the zebrafish embryo

Definition of cell fates along the dorso-ventral axis depends on an antagonistic relationship between ventralizing transforming growth factor-beta superfamily members, the bone morphogenetic proteins and factors secreted from the dorsal organizer, such as Noggin and Chordin. The extracellular binding of the last group to the bone morphogenetic proteins prevents them from activating their receptors, and the relative ventralizer:antagonist ratio is thought to specify different dorso-ventral cell fates. Here, by taking advantage of a non-genetic interference method using a specific competitive inhibitor, the Lefty-related gene product Antivin, we provide evidence that cell fate along the antero-posterior axis of the zebrafish embryo is controlled by the morphogenetic activity of another transforming growth factor-beta superfamily subgroup--the Activin and Nodal-related factors. Increasing antivin doses progressively deleted posterior fates within the ectoderm, eventually resulting in the removal of all fates except forebrain and eyes. In contrast, overexpression of activin or nodal-related factors converted ectoderm that was fated to be forebrain into more posterior ectodermal or mesendodermal fates. We propose that modulation of intercellular signalling by Antivin/Activin and Nodal-related factors provides a mechanism for the graded establishment of cell fates along the antero-posterior axis of the zebrafish embryo.     

Silberblick/Wnt11 mediates convergent extension movements during zebrafish gastrulation

Vertebrate gastrulation involves the specification and coordinated movement of large populations of cells that give rise to the ectodermal, mesodermal and endodermal germ layers. Although many of the genes involved in the specification of cell identity during this process have been identified, little is known of the genes that coordinate cell movement. Here we show that the zebrafish silberblick (slb) locus encodes Wnt11 and that Slb/Wnt11 activity is required for cells to undergo correct convergent extension movements during gastrulation. In the absence of Slb/Wnt11 function, abnormal extension of axial tissue results in cyclopia and other midline defects in the head. The requirement for Slb/Wnt11 is cell non-autonomous, and our results indicate that the correct extension of axial tissue is at least partly dependent on medio-lateral cell intercalation in paraxial tissue. We also show that the slb phenotype is rescued by a truncated form of Dishevelled that does not signal through the canonical Wnt pathway, suggesting that, as in flies, Wnt signalling might mediate morphogenetic events through a divergent signal transduction cascade. Our results provide genetic and experimental evidence that Wnt activity in lateral tissues has a crucial role in driving the convergent extension movements underlying vertebrate gastrulation.     

Molecular recycling within amyloid fibrils

Amyloid fibrils are thread-like protein aggregates with a core region formed from repetitive arrays of beta-sheets oriented parallel to the fibril axis. Such structures were first recognized in clinical disorders, but more recently have also been linked to a variety of non-pathogenic phenomena ranging from the transfer of genetic information to synaptic changes associated with memory. The observation that many proteins can convert into similar structures in vitro has suggested that this ability is a generic feature of polypeptide chains. Here we have probed the nature of the amyloid structure by monitoring hydrogen/deuterium exchange in fibrils formed from an SH3 domain using a combination of nuclear magnetic resonance spectroscopy and electrospray ionization mass spectrometry. The results reveal that under the conditions used in this study, exchange is dominated by a mechanism of dissociation and re-association that results in the recycling of molecules within the fibril population. This insight into the dynamic nature of amyloid fibrils, and the ability to determine the parameters that define this behaviour, have important implications for the design of therapeutic strategies directed against amyloid disease.     

Eomesodermin is required for mouse trophoblast development and mesoderm formation

The earliest cell fate decision in the mammalian embryo separates the extra-embryonic trophoblast lineage, which forms the fetal portion of the placenta, from the embryonic cell lineages. The body plan of the embryo proper is established only later at gastrulation, when the pluripotent epiblast gives rise to the germ layers ectoderm, mesoderm and endoderm. Here we show that the T-box gene Eomesodermin performs essential functions in both trophoblast development and gastrulation. Mouse embryos lacking Eomesodermin arrest at the blastocyst stage. Mutant trophoectoderm does not differentiate into trophoblast, indicating that Eomesodermin may be required for the development of trophoblast stem cells. In the embryo proper, Eomesodermin is essential for mesoderm formation. Although the specification of the anterior-posterior axis and the initial response to mesoderm-inducing signals is intact in mutant epiblasts, the prospective mesodermal cells are not recruited into the primitive streak. Our results indicate that Eomesodermin defines a conserved molecular pathway controlling the morphogenetic movements of germ layer formation and has acquired a new function in mammals in the differentiation of trophoblast.     

金黄滴虫的纤维系统

用含TritonX-100和EGTA的PIPES缓冲液处理金黄滴虫（Ochromonasdanica,原生动物鞭毛虫纲）,溶去细胞中可溶性蛋白,可获得完整的、在透射电镜下清晰可辨的细胞纤维系统。细胞前端有一横跨长鞭毛基粒的,由微管束构成的三叉形纤维,其上有约25条纵行微管束,沿细胞外周向后延伸直达细胞后端,从而形成一个完整的细胞骨架。细胞前端还有一对较细的环纤维,围绕在短鞭毛的基部,其中一个缠绕成“8”字形。根质体位于长鞭毛基粒和细胞核之间,有横纹,呈带状或扇状。对于纤维的化学组成,也作了一些初步的探查。     

Peptides containing the cell-attachment recognition signal Arg-Gly-Asp prevent gastrulation in Drosophila embryos

It has recently been suggested that the Arg-Gly-Asp sequence (RGD) forms part of a widespread cell-extracellular matrix recognition system. Analysis of the cell binding sites of vertebrate fibronectin and other extracellular proteins that interact with cell surfaces implicate the same amino acid triplet. Peptides containing this sequence inhibit certain developmental events such as cell-matrix adhesion or cellular migration in vitro and in vivo. The RGD-sequence is also part of the cellular recognition site of the aggregation protein discoidin I in Dictyostelium suggesting that the RGD-recognition system could be universally used. In Drosophila, despite its advanced genetics, very little is known about the extracellular components that are involved in cell movements and morphogenesis. We report here that peptides containing the RGD-sequence prevent gastrulation of Drosophila embryos. The phenotypic effect is similar to that observed in the dorsal-group mutants: no ventral furrow is formed and the embryos lack dorsal-ventral polarity. The specificity of the inhibiting action suggests that the RGD-sequence may also be used by invertebrates to mediate cell-attachment phenomena.     

斑节对虾白斑综合症杆状病毒（WSSV）感染的组织特异性

应用光镜和电镜研究斑节对虾白斑综合症杆状病毒（ＷＳＳＶ）感染的组织特异性,结果证实ＷＳＳＶ组织特异性差,感染外胚层和中胚层来源的各种组织细胞的细胞核,如甲壳表皮细胞、鳃上皮细胞、神经细胞、造血组织细胞、胃上皮细胞、心肌细胞及其随体细胞、触角腺上皮细胞、胃和消化管的肌细胞和结缔组织细胞等,以甲壳表皮细胞、胃上皮细胞、结缔组织细胞和造血组织细胞最为显,细胞解体坏死。电镜观察对虾中肠和肝胰细胞未发现病     

Structure-based design of non-natural amino-acid inhibitors of amyloid fibril formation

Many globular and natively disordered proteins can convert into amyloid fibrils. These fibrils are associated with numerous pathologies as well as with normal cellular functions, and frequently form during protein denaturation. Inhibitors of pathological amyloid fibril formation could be useful in the development of therapeutics, provided that the inhibitors were specific enough to avoid interfering with normal processes. Here we show that computer-aided, structure-based design can yield highly specific peptide inhibitors of amyloid formation. Using known atomic structures of segments of amyloid fibrils as templates, we have designed and characterized an all-D-amino-acid inhibitor of the fibril formation of the tau protein associated with Alzheimer's disease, and a non-natural L-amino-acid inhibitor of an amyloid fibril that enhances sexual transmission of human immunodeficiency virus. Our results indicate that peptides from structure-based designs can disrupt the fibril formation of full-length proteins, including those, such as tau protein, that lack fully ordered native structures. Because the inhibiting peptides have been designed on structures of dual-β-sheet 'steric zippers', the successful inhibition of amyloid fibril formation strengthens the hypothesis that amyloid spines contain steric zippers.     

芳香族聚酰胺（PPTA）纤维弯曲疲劳的形变和断裂机理

普通PPTA纤维和改性PPTA纤维具有近似的应力应变的曲线,但后者的弯曲疲劳寿命比前者要长的多。在SEM下观察了弯曲疲劳过程的形态变化,发现该过程主要是结构沿纤维长度分裂即原纤化过程。改性PPTA纤维对原纤化的阻抗显著地大于普通PPTA纤维。粉碎降解和低温断裂的结果也得出类似结果。表明大分子间结合力在PPTA纤维弯曲疲劳中起很重要的作用。弯曲疲劳作用被归结为对材料拉伸和压缩的复合作用。拉伸PPTA纤维的动态观察得出,拉伸或压缩纤维将出现剪切带或滑移带,它们引发纤维原纤间的分离。剪切屈服是其主要形变机制。PPTA纤维弯曲疲劳过程如下：重复弯曲引起材料局部区域的剪切屈服,出现剪切带,并随之发生结构分裂,纤维广泛地原纤化,最后原纤和原纤束相继破坏,纤维最终断裂。     

Quasi-hexagonal molecular packing in collagen fibrils

Collagen molecules in native 66.8 nm (D) periodic fibrils are widely believed to be assembled into discrete, rope-like substructures, or microfibrils. Several types of microfibril have been proposed (2, 4, 5, 7- and 8-stranded) mainly on the basis of information contained in the medium angle X-ray diffraction patterns of native tendon fibres. These patterns show a series of equatorial and near-equatorial Bragg reflections which indicate that the collagen molecules are arranged on a three-dimensional crystalline lattice. The 4-stranded, 5-stranded and 8-stranded microfibrils are D-periodic with approximate diameter 3.8 nm, and these and the 2-stranded model are supposed to be packed on a three-dimensional lattice whose basal unit cell, (approximately) perpendicular to the fibril axis, is tetragonal (or quasi-tetragonal)with side a, a square root 2 or 2a, where a is approximately 3.8 nm. In this paper we describe a re-interpretation of the X-ray data which leads to a new model for the crystalline regions of the fibril, based on quasi-hexagonal molecular packing without microfibrillar sub-structures, and hence having the character of a molecular crystal.     

Homeobox gene expression correlated with the bifurcation process of limb cartilage development.

The complex architecture of the limb cartilage pattern probably develops by the sequential segmentation and branching process of precartilaginous cell condensation under the control of positional signalling provided by the zone of polarizing activity (anteroposterior) and the apical ectodermal ridge (proximodistal). This signalling is monitored and interpreted in the mesenchymal cells and induces the position-specific response of subsets of genes. Homeobox genes may be responsible for the interpretation of signalling. A correlation between limb pattern and expression domains of the homeobox genes in the upstream region of Hox/Chox-4 has been proposed. We have analysed the spatial expression pattern of the Chox-1 genes during development of chick limb buds. In contrast to genes in Hox/Chox-4 expressed coordinately along the anteroposterior axis, homeobox genes in Chox-1 have unique and mutually exclusive expression domains along the proximodistal axis. We report here that the expression domains of the Chox-1 genes are closely related to the segmental structure of cartilage along the proximodistal axis, whereas the expression domains of the Chox-4 genes are related to the cartilage branching pattern.     

The amniote primitive streak is defined by epithelial cell intercalation before gastrulation

During gastrulation, a single epithelial cell layer, the ectoderm, generates two others: the mesoderm and the endoderm. In amniotes (birds and mammals), mesendoderm formation occurs through an axial midline structure, the primitive streak, the formation of which is preceded by massive 'polonaise' movements of ectoderm cells. The mechanisms controlling these processes are unknown. Here, using multi-photon time-lapse microscopy of chick (Gallus gallus) embryos, we reveal a medio-lateral cell intercalation confined to the ectodermal subdomain where the streak will later form. This intercalation event differs from the convergent extension movements of the mesoderm described in fish and amphibians (anamniotes): it occurs before gastrulation and within a tight columnar epithelium. Fibroblast growth factor from the extraembryonic endoderm (hypoblast, a cell layer unique to amniotes) directs the expression of Wnt planar-cell-polarity pathway components to the intercalation domain. Disruption of this Wnt pathway causes the mesendoderm to form peripherally, as in anamniotes. We propose that the amniote primitive streak evolved from the ancestral blastopore by acquisition of an additional medio-lateral intercalation event, preceding gastrulation and acting independently of mesendoderm formation to position the primitive streak at the midline.     

Effects of contact guidance and gravity on L929 cell orientation

Contact guidance and external force field are two important factors that influence cell orientation. Microgroove-like patterns on PDMS substrates were fabricated using soft-lithography, and the effects of gravitational field direction relative to the pattern, on L929 cell orientation were investigated. The majority of cells were aligned along the grooves when gravity was parallel to the groove direction, because of contact guidance. When gravity was perpendicular to the groove direction, cell alignment decreased noticeably. Although contact guidance induced by the pattern played a dominant role in determining cell orientation, gravity also had an important influence when the field direction was perpendicular to the groove direction. No synergistic influence on cell alignment was observed when the field direction was parallel to the groove direction.     

Polarity controls forces governing asymmetric spindle positioning in the Caenorhabditis elegans embryo

Cell divisions that create daughter cells of different sizes are crucial for the generation of cell diversity during animal development. In such asymmetric divisions, the mitotic spindle must be asymmetrically positioned at the end of anaphase. The mechanisms by which cell polarity translates to asymmetric spindle positioning remain unclear. Here we examine the nature of the forces governing asymmetric spindle positioning in the single-cell-stage Caenorhabditis elegans embryo. To reveal the forces that act on each spindle pole, we removed the central spindle in living embryos either physically with an ultraviolet laser microbeam, or genetically by RNA-mediated interference of a kinesin. We show that pulling forces external to the spindle act on the two spindle poles. A stronger net force acts on the posterior pole, thereby explaining the overall posterior displacement seen in wild-type embryos. We also show that the net force acting on each spindle pole is under control of the par genes that are required for cell polarity along the anterior-posterior embryonic axis. Finally, we discuss simple mathematical models that describe the main features of spindle pole behaviour. Our work suggests a mechanism for generating asymmetry in spindle positioning by varying the net pulling force that acts on each spindle pole, thus allowing for the generation of daughter cells with different sizes.     

有限厚度的Marangoni对流边界层问题的解析近似解

讨论了由不同温度介质的界面张力梯度而诱导的有限厚度Marangoni对流的边界层问题.假设界面张力是温度的平方函数,下表面保持恒温,上表面的温度是水平距离的线性函数.通过坐标变换和巧妙引入小参数对速度和温度边界层控制方程组摄动渐进展开,得到了问题的解析近似解.研究了Marangoni数和Prandtl数对速度、温度边界层的影响,对相应的流动特性进行了探讨.     

巴楚-麦盖提地区油气动态成藏的运移通道

巴楚-麦盖提地区油气运移输导网络由断裂、不整合面、储层孔、洞、缝系统组成;通过对断裂封堵和开启性进行分析,烃类沿断裂面作s形路径运移;八期主要不整合面中T60和T55不整合面在全区为开启,T70和T54不整合面在巴楚隆起南界断裂构造带为开启.在海西和喜马拉雅两大成烃期中,该区具多期动态成藏的特点,主运移通道也存在时空上的差别:加里东中期-海西晚期之前不整合面、下寒武统白云岩孔、洞、缝系统为主通道;海西晚期-喜马拉雅期,断裂的多期活动促使早期古油气藏调整和喜马拉雅期生成的油气聚集,不整合面和断裂通道占主导,巴楚隆起西段边界断裂带为喜马拉雅晚期油气聚集调整分配区.     

Electron microscope evidence for an 80 A unit in collagen fibrils.

Connective tissues are composite structures containing collagen, elastin, glycosaminoglycans, minerals, water and other minor components. In all cases the collagen exists predominantly in fibrillar form. The size distribution of the fibrils does, however, vary markedly with both age and the mechanical requirements of the tissue. Little is known about the mechanism of fibril formation in vivo, although some information is now available form in vitro polymerisation studies. We have now collected new and extensive electron microscope data on the size of collagen fibrils from tendon, skin, cornea and other tissues from both fetal and immature animals. The results reported here show that the diameters of the collagen fibrils thus measured lie close to a multiple of 80 A, a result which may be simply and readily interpreted in terms of the collagen microfibril and its mode of packing.     

Forward transport of glycoproteins on leading lamellipodia in locomoting cells

In several types of locomoting cells, active rearward transport of particles on the cell surface has been observed and correlated with motility. No forward transport of particles has previously been reported, however. Here we report rapid forward transport of concanavalin A-coated gold particles on the dorsal surfaces of lamellipodia of fish epidermal keratocytes. These movements are active, not diffusive, and more rapid than either rearward particle transport or the rate of cell locomotion. We observed forward transport in migrating, but not in stationary cells, and could block the movement by treatment with cytochalasin D. These studies demonstrate for the first time that small numbers of glycoproteins can be actively transported on the surface of the cell to the front of the lamellipodium. We suggest that this mechanism transports proteins involved in cell locomotion, such as proteins necessary for adhesion, and could also produce an extensile force.