bluuebunny/crossref_metadata_2025 huggingface dataset:
Drosophila necroticmutations mirror disease-associated variants of human serpins
Authors: Green Clare , Brown Gemma , Dafforn Timothy R. , Reichhart Jean-Marc , Morley Terri , Lomas David A. , Gubb David
Published: February 2003

Polymerization of members of the serpin superfamily underlies diseases as diverse as cirrhosis, angioedema, thrombosis and dementia. TheDrosophila serpin Necrotic controls the innate immune response and is homologous to human α1-antitrypsin. We show thatnecrotic mutations that are identical to the Z-deficiency variant ofα 1-antitrypsin form urea-stable polymers in vivo. Thesenecrotic mutations are temperature sensitive, which is in keeping with the temperature-dependent polymerization of serpins in vitro and the role of childhood fevers in exacerbating liver disease in Z α-antitrypsin deficiency. In addition, we identify two nec mutations homologous to an antithrombin point mutation that is responsible for neonatal thrombosis. Transgenic flies carrying an S>F amino-acid substitution equivalent to that found in Siiyama-variant antitrypsin (necS>F.UAS) fail to complement nec-null mutations and demonstrate a dominant temperature-dependent inactivation of the wild-type nec allele. Taken together, these data establish Drosophila as a powerful system to study serpin polymerization in vivo.
A role for the mesenchymal T-box geneBrachyuryin AER formation during limb development
Authors: Liu Chunqiao , Nakamura Eiichiro , Knezevic Vladimir , Hunter Sherrie , Thompson Katherine , Mackem Susan
Published: February 2003

During limb development, several signaling centers organize limb pattern. One of these, the apical ectodermal ridge (AER), is critical for proximodistal limb outgrowth mediated by FGFs. Signals from the underlying mesoderm,including WNTs and FGFs, regulate early steps of AER induction. Ectodermal factors, particularly En1, play a critical role in regulating morphogenesis of a mature, compact AER along the distal limb apex, from a broad ventral ectodermal precursor domain. Contribution of mesodermal factors to the morphogenesis of a mature AER is less clear. We previously noted that the chick T gene (Brachyury), the prototypical T-box transcription factor, is expressed in the limb bud as well as axial mesoderm and primitive streak. Here we show that T is expressed in lateral plate mesoderm at the onset of limb bud formation and subsequently in the subridge mesoderm beneath the AER. Retroviral misexpression of T in chick results in anterior extension of the AER and subsequent limb phenotypes consistent with augmented AER extent and function. Analysis of markers for functional AER in mouse T-/- null mutant limb buds reveals disrupted AER morphogenesis. Our data also suggest that FGF and WNT signals may operate both upstream and downstream of T. Taken together, the results show that T plays a role in the regulation of AER formation,particularly maturation, and suggest that T may also be a component of the epithelialmesenchymal regulatory loop involved in maintenance of a mature functioning AER.
Dock and Pak regulate olfactory axon pathfinding inDrosophila
Authors: Ang Lay-Hong , Kim Jenny , Stepensky Vitaly , Hing Huey
Published: February 2003

The convergence of olfactory axons expressing particular odorant receptor(Or) genes on spatially invariant glomeruli in the brain is one of the most dramatic examples of precise axon targeting in developmental neurobiology. The cellular and molecular mechanisms by which olfactory axons pathfind to their targets are poorly understood. We report here that the SH2/SH3 adapter Dock and the serine/threonine kinase Pak are necessary for the precise guidance of olfactory axons. Using antibody localization, mosaic analyses and cell-type specific rescue, we observed that Dock and Pak are expressed in olfactory axons and function autonomously in olfactory neurons to regulate the precise wiring of the olfactory map. Detailed analyses of the mutant phenotypes in whole mutants and in small multicellular clones indicate that Dock and Pak do not control olfactory neuron (ON) differentiation, but specifically regulate multiple aspects of axon trajectories to guide them to their cognate glomeruli. Structure/function studies show that Dock and Pak form a signaling pathway that mediates the response of olfactory axons to guidance cues in the developing antennal lobe (AL). Our findings therefore identify a central signaling module that is used by ONs to project to their cognate glomeruli.
Tumour-endothelium interactions in co-culture: coordinated changes of gene expression profiles and phenotypic properties of endothelial cells
Authors: Khodarev Nikolai N. , Yu Jianqing , Labay Edwardine , Darga Thomas , Brown Charles K. , Mauceri Helena J. , Yassari Reza , Gupta Nalin , Weichselbaum Ralph R.
Published: February 2003

Tumour angiogenesis is a complex process based upon a sequence of interactions between tumour cells and endothelial cells. To model tumour/endothelial-cell interactions, we co-cultured U87 human glioma cells with human umbilical vein endothelial cells (HUVECs). U87 cells induced an`activated' phenotype in HUVECs, including an increase in proliferation,migration and net-like formation. Activation was observed in co-cultures where cells were in direct contact and physically separated, suggesting an important role for soluble factor(s) in the phenotypic and genotypic changes observed. Expressional profiling of tumour-activated endothelial cells was evaluated using cDNA arrays and confirmed by quantitative PCR. Matching pairs of receptors/ligands were found to be coordinately expressed, including TGFβRII with TGFβ3, FGFRII and cysteine-rich fibroblast growth factor receptor (CRF-1) with FGF7 and FGF12, CCR1, CCR3, CCR5 with RANTES and calcitronin receptor-like gene (CALCRL) with adrenomedullin. Consistent with cDNA array data, immunohistochemical staining of expressed proteins revealed the upregulation of Tie-2 receptor in vitro and in vivo. Our data suggest that tumour-induced activation of quiescent endothelial cells involves the expression of angiogenesis-related receptors and the induction of autocrine growth loops. We suggest that tumour cells release growth factors that induce endothelial cells to express specific ligands and their cognate receptors coordinately.
Creep analysis of pressurized circumferential pipe weldments—a review
Authors: Hyde T H , Sun W , Williams J. A
Published: February 2003

This paper reviews work related to the high-temperature creep analysis of pressurized circumferential pipe weldments. It is important to define the problem and thus metallurgical features correctly; the identification of material microstructural-property variations within the heat-affected zone (HAZ) and the failure modes of welds are briefly included, as well as in-service experience of pipe welds. Experimental methods, including model and full-size component testing, are summarized and examples of typical tests results are described. Material constitutive equations, which can be used in describing the creep deformation of and failure mechanisms in welds, are briefly described. Numerical modelling using finite element (FE) methods, covering a range of approaches and analyses, taking account of the effects of material properties, pipe geometry, weld dimensions and system loading, on the stresses and failure behaviour of pipe weldments, are summarized. Typical results are presented to illustrate the potential uses and limitations of the FE methods.
Structural deflection measurement
Authors: Djordjevich A
Published: February 2003

The concept of measuring deflection curvature of structures under bending loading with a device called a ‘curvature gauge’ is described. The application domain is specified in which this concept offers advantages over the alternative of measuring strain in terms of the measurement sensitivity, flexibility to choose the location for the sensor placement and immunity to the effects of microstructural interfacing between dissimilar materials that the sensor presence introduces. Details of the curvature gauge sensor are also given.
Neuber prediction of elastic-plastic strain concentration in notched tensile specimens under large-scale yielding
Authors: Härkegård G , Mann T
Published: February 2003

A general version of Neuber's rule, applicable to large-scale yielding, is presented. It is applied to a variety of notched tensile specimens made of power-hardening elastic-plastic material. Neuber's rule predictions of notch stress and strain are compared with finite element results. The best agreement is obtained for shallow mild notches. Models of more limited applicability, based on the invariance of the total strain energy density and the strain energy density at the root of the notch, have been applied to some of the specimens.
Organised Mesoporous Silica Synthesised by Nanoscale Duplication of an Ordered Mesoporous Carbon Material Using a Gas Phase Process
Authors: Parmentier J , Vix-Guterl C , Saadallah S , Reda M , Illescu M , Werckmann J , Patarin J
Published: February 2003

Organised mesoporous silica was synthesised by nanoscale duplication of an ordered mesoporous carbon material using a gas phase process.
Recovery of Original Organic Parameters of the the Outcropping Source Rocks from South China
Authors: Sun Yuzhuang , Sun J. P. , Zhang H. J. , Liu L. F.
Published: February 2003

Two samples from outcrops and six core samples of the clay source rocks from the Shiwanshan and Baise Basins were analysed in order to study the variation of selected organic parameters. A comparison of the samples taken from the outcrops with core samples was then performed. The results indicate a significant alteration of the organic matter at and near the surface. A depletion of about 70% TOC and extract yields was seen. The depth influenced by oxidation was found to be about 50 m. The most significant depletion of the organic matter occurred within 16 m.
Energy Utilization Systems
Authors: Demirbaş Ayhan
Published: February 2003

Energy affects all aspects of modern life. The demand for energy is increasing at an exponential rate due to the exponential growth of world population. Advanced energy-efficiency technologies reduce the energy needed to provide energy services, thereby reducing environmental and national security costs of using energy and potentially increasing its reliability. Energy resources are not used for their own sake, but for the services they provide. These energy services are fundamental to our modern economy; they heat, cool, and light our buildings; power our industrial processes; process our food; fuel our transport; and energize our communications and information technologies.
Wind Turbine Structural Dynamics – A Review of the Principles for Modern Power Generation, Onshore and Offshore
Authors: van der Tempel Jan , Molenaar David-Pieter
Published: February 2003

Wind turbines for electricity production have two seemingly opposing constraints; they need to be structural secure yet of low cost. To meet the first constraint, it would be an obvious choice to design a stiff structure of consequently large mass but this would drive up the cost. By reducing the mass a more cost effective turbine can be realized. However, such lightweight structures are by definition more flexible. To design a cost effective flexible system, thorough understanding of the dynamics is essential. This paper reviews the theoretical basics of the dynamic design options and applies these to realistic situations, including offshore machines under wave action. The wind energy converter and the support structure form an integrated dynamic system that must be developed in mutual interdependency and close co-operation. This paper provides a contribution to this integration process by extending the design approach initiated in the Opti-OWECS study [1] and the work of Kühn [2].
Isolated cytoskeletons of human blood platelets: dark‐field imaging of coiled and uncoiled microtubules
Authors: Kowit Joel D. , Linck Richard W. , Kenney Dianne M.
Published: February 2003

Detergent extraction of human blood platelets pre‐treated with Taxol to stabilize microtubules allows isolation of marginal band (MB) cytoskeletons. We studied MB cytoskeleton structure using dark‐field light microscopy and negative stain electron microscopy (EM). Dark‐field illumination clearly demonstrated the “hoop” shape of MB cytoskeletons in unfixed suspension where the microtubule coils had a mean diameter of 2.87 μm (± 0.18 μm, SD). Microtubules were uncoiled by brief exposure to trypsin (2 mg/μg protein) or by NaCl (154–600 mM) but not by DNase I, which removed ∼40% of total actin, but had no effect on dark‐field images of microtubule coils. As microtubules uncoiled, a single fiber emerged from the hoop and gradually lengthened as the brightness of the hoop diminished; these fibers correspond to the single microtubules seen by EM. Polypeptides of coiled and uncoiled MB cytoskeletons were analyzed by SDS‐PAGE. When microtubules became uncoiled, no changes in the major components (α‐ and β‐tubulin, IEF‐51K, or actin) were found. However, a number (> 10) of minor polypeptides, each <5% of total cytoskeletal protein and with an Mr ranging from 80,000≽ 260,000, were decreased in “uncoiled” MB cytoskeletons. These results implicate one or more of these minor polypeptides in maintenance of hoop integrity. Dark‐field light microscopy thus provides an approach toward investigating the mechanisms(s) involved in maintaining the microtubule coil of the platelet marginal band.
Autoimmune serum containing an antibody against a 94 kDa nucleolar protein
Authors: HERNANDEZ‐VERDUN Daniele , PRÉVOT Sophie , ANDRÉ Chantal , GUILLY Marie‐Noëlle , MASSON Claude , WOLFE Jason
Published: February 2003

Little information exists on how various nucleolar proteins function in ribosome biogenesis. Of special interest is that group of nucleolar proteins which are not incorporated into mature ribosomes because they are candidates for a role in the regulation of ribosome construction. Non‐ribosomal nucleolar proteins can be analyzed using autoimmune sera from scleroderma patients which often contain antinuclear antibodies. One such serum, designated ScBr, is shown by indirect immunofluorescence to react specifically with nucleoli in cells of 3 different mammalian species, indicating that the antigen is at least partly conserved evolutionarily. It is not RNase‐sensitive, but is completely eliminated after incubation with pronase and 2 M NaCl. Immuno‐electron microscopy was carried out on Lowicryl ultrathin sections to localize the antigen. The labeling was observed over both the granular and the dense fibrillar component but not the fibrillar centers, indicating that the antigen is associated with ribosomal RNA transcription sites and ribosome assembly into precursor particles. In addition, the antibody was localized to small nucleoplasmic entities, termed dense nuclear bodies. This could indicate a relationship between nucleoli and dense nuclear bodies. By immunoprecipitation, only a single protein of 94 kDa molecular weight was revealed. By immunoblotting, the band at 94 kDa was found to be the only positive band for high ScBr dilutions. Observation of the behavior of the antigen during mitosis revealed that it became dispersed into the cytoplasm after breakdown of the nuclear envelope, lining most of the chromosomes rather that remaining associated with the NOR‐chromosomes. The antigen appeared to be restored to nucleoli only in late telophase; phase‐dense prenucleolar bodies of early telophase cells did not show positive staining for the antigen. During actinomycin‐D RNA synthesis inhibition as well as in non‐stimulated lymphocytes the positive staining is greatly decreased. These results were consistent with a role for the 94 kDa nucleolar protein in the process of preribosome assembly.
Immunolocalization of laminin during estrogen‐induced differentiation of quail oviduct epithelial cells
Authors: PERCHE Olivier , SANDOZ Daniel
Published: February 2003

During estrogen‐induced development of the quail oviduct, tubular glands are formed by evagination of epithelial cells into the stroma. The distribution of laminin was studied during the early stages by means of immunofluorescence and immunoperoxidase techniques. Ultrastructural changes in the basal lamina were studied by electron miscrocopy.

Basement membranes at all stages of development were delineated with 3 polyclonal antilaminin antisera. However, in ovariectomized birds, laminin could not be detected by one of the polyclonal antilaminin antisera. Subsequently, this antibody detected laminin as epithellal cell evaginations were induced by estradiol benzoate. The heavy and light chains of Engelbreth Holm sarcoma (EHS) laminin were revealed in immunoblotting by all antibodies.

By electron microscopy after the immunoperoxidase technique with antilaminin antisera laminin appears to be accumulated mainly in the lamina densa. Furthermore, the thickness of the basal lamina increases during oviduct development. These data indicate that basal lamina organization is modified during oviduct cell differentiation and that immunoreactivity of epithelial basement membrane laminin changes during development.
Ultrastructure of the flagellar apparatus of Oxyrrhis marina
Authors: Cachon Monique , Cosson Jacky , Cosson Marie‐Paule , Huitorel Philippe , Cachon Jean
Published: February 2003

Summary— Oxyrrhis marina, like all dinoflagellates, possesses one transverse and one longitudinal flagellum, which show structural differences.

The transverse flagellum contains a small fibre, 20 nm in diameter, associated with doublet no.7, whereas the longitudinal flagellum is substantially by a large (200–300 nm) hollows structure closely resembling the paraflagellar rod described by several authors in kinetoplastidae and in euglenoids. This structure is made up of a hemicylindrical network of filaments which are often linked on one side to the outer doublet no. 4, and on the other side to a dense plate. Another thinner filamentous network closes this hemicyclinder. In cross‐section, the wall of this structure is made up of 8 filaments 2–4 nm in diameter that show a thicker periodic structure. In longitudinal section the same filaments appear arranged in periodic rhombus meshes or a helicoidal pattern, depending on the orientation of the section relative to the axoneme.
Ultrastructures and evolutionary modalities of flagellar and ciliary systems in protists
Authors: Grain Jean , Mignot Jean‐Pierre , de Puytorac Pierre
Published: February 2003

Summary— As both the ultrastructure and function of flagella and cilia have been for the main part remarkably conserved during Eukaryote, evolution, the question arises as to whether the variations observed at the organite ultrastural level, or at the level of the development of a flagellar or ciliary cellular system could be considered as systematic or phylogenetic criteria.

With regard to the fundamental structure, the known variations concern: 1) the kinetosome (length, position and number of cartwheels, number of triplets, and respective lengths of the microtubules); 2) the transition zone (various structural types); 3) the axoneme (number of doublets, central tubules, arms); 4) the paraxonemal formations (presence, position, structure); 5) the membrane (intramembranous particles; intramembranous particles; addition of components, mastigonemes, scales); 6) the fibres associated with the kinetosomes. Some of these variations are characteristic of taxa, and are considered as phylogenetic markers.

Regarding the variations in the number of ciliary or flagellar units per cell, the following can be distinguished: cells with only one kinetosome (Ks), carrying one flagellum; cells with 2 neighbouring Ks (primary Ks = 2A), only one Ks, or the 2 Ks, bearing a flagellum; cells with numerous flagella or cilia. We consider that this configuration can result from: a) addition of new Ks around the primary Ks, forming a primary group (PG): either by replication of the primary couple (2 × 2A in Karotomorpha; 2 × 2A in Phaeopolykrikos; 4(2 × 2A) in Polkrikos), or by addition of new Ks (N) without copy or replication of the primary Ks (2A + 2N in Polytomella; 2A +3N in Tetratrichomonas; 2A + 3N in Tetratrichomonas; 2A + 4N in Hexamastix; 2A + 6N in Pyramimonas octopus). b) amplification of each constituent of the primordial couple in opalinids. c) amplification of the primary group [yx(AA + 2N) with y = 2 in diplomonadida and y > 2 in calonymphidae. d) appearance in morphogenetic fields situated outside the primary group (but possibly related to it) of new Ks which then form secondary groups (SG) where they are arranged in polarized rows composed of juxtaposed cortical units (monokinetids). In this case, PG can be of type 2A+2N (1PG+1SG in lophomonadida; 1PG+2SG in some trichonymphina), or can lose some Ks, resulting in reduced PG (4 reduced PG+4GS in Staurojoenina, n reduced PG + nGS in spirotrichonymphina). The PG may even disappear; either partially leaving remnants in the form of MTOCs in Stephanopogon, or completely, as in cilliates where the general ciliature would represent on the SG.

From the above, it is seen that: a) the pluriflagellar state in protists depends on only a small number of factors; b) the same factors are used in different groups; c) different states have been obtained within different groups. An analysis of the respective evolution of kinetosomes and centrioles is proposed.
Tubulin expression in trypanosomes
Authors: Gallo Jean‐Marc , Precigout Eric
Published: February 2003

Microtubules in trypanosomes are the main component of the flagellar axoneme and of the subpellicular microtubule corset, whose relative positions determine the morphology of each cell stage of the life cycle of these parasites. Microtubules are polymers of tubulin, a protein dimer of two 55‐kDa subunits, α‐ and β‐tubulin; in Trypanosoma brucei, the tubulin‐coding sequences are clustered in a 40‐kb fragment of tandemly repeated α‐ and β‐tubulin genes separated by a 170‐bp intergenic zone. This cluster is transcribed in a unique RNA which is rapidly processed into mature mRNAs carrying the 5′ 35‐nucleotide leader sequence found in all trypanosome mRNAs. Although no heterogeneity has been found at the gene level, tubulin can be post‐translationally modified in 2 ways: the C‐terminal tyrosine of α‐tubulin can be selectively cleaved and added again with 2 enzymes, tubulin carboxypeptidase and tubulin‐tyrosine ligase; α‐tubulin can also be acetylated on a lysine residue. Some molecular domains of tubulin are restricted to subpopulations of microtubules; for instance, the β‐tubulin form defined by the monoclonal antibody 1B41 is sequested into a part of the subpellicular cytoskeleton limited to the flagellar adhesion zone, which might correspond to the group of 4 microtubules associated with a cisterna of the endoplasmic reticulum, forming the so‐called “subpellicular microtubule quartet” (SFMQ). The early assembly of this zone in each daughter cell during the cell division of T. brucei, together with the alterations undergone by the domain defined by the monoclonal antitubulin 24E3 during the differentiation of Trypanosoma cruzi, suggest that specific tubulin forms are responsible for dynamic properties of SFMQ possibly involved in trypanosome morphogenesis.
Compartmentation of glycolysis in trypanosomes: a potential target for new trypanocidal drugs
Authors: Michels Paul A.M.
Published: February 2003

In African trypanosomes most enzymes of the glycolytic pathway are found in a microbody‐like organelle, called the glycosome. The analysis of their structural and functional properties has shown that these glycosomal enzymes possess some specific features which are absent from the cytosolic proteins of trypanosomes and from the glycolytic enzymes of other organisms, where glycolysis is not compartmentalized within an organelle. The specific properties of the glycosomal enzymes may be responsible for the routing of the proteins from their site of synthesis, the cytosol, into the glycosome, or they may be involved in the proper functioning of the enzymes within the organelle. Whatever the role of the unique features, they are potential targets for compounds that could specifically interfere with glycolysis in trypanosomes. Therefore, a detailed study of the glycolytic enzymes of trypanosomes may lead to the development of therapeutically useful drugs against these harmful parasites.
Morphological changes in erythrocytes induced by malarial parasites
Authors: Aikawa Masamichi
Published: February 2003

Host cell alterations induced by Plasmodium falciparum, P. brasillanum, P. vivax and P. malariae were described by electron microscopy and post‐embedding immunoelectron microscopy. P. falciparum infection induces knobs, electron‐dense material and clefts in the erythrocyte. Clefts are involved in exporting P. falciparum antigen from the parasite to the erythrocyte membrane. P. falciparum antigen is present in knobs which adhere to endothelial cells causing the blockage of cerebral capillaries and ensuing pathological changes in cerebral tissues. P. brasilianum infection induces knobs, short and long clefts and electron‐dense material. These structures appear to contain different P. brasilianum antigens. This indicates that each structure functions independently in trafficking P. brasilianum protein to the erythrocyte surface. P. vivax infection induces caveola‐vesicle complexes and clefts in the erythrocyte. These structures are also involved in trafficking P. vivax protein from the parasite to the erythrocyte membrane. P. malariae induces caveolae, electron‐dense material, vesicles, clefts and knobs in the erythrocyte. Although vesicles and caveolae are seen in the erythrocyte cytoplasm, they do not form caveola‐vesicle complexes as seen in P. vivax‐infected erythrocytes. They also appear to be involved in trafficking of malaria antigens. These studies, therefore, indicate that host cell changes occur in order to facilitate the transport of malarial antigens to the host cell membrane. The significance of these phenomena is still not clear.
Isolation and functional characterization of Plasmodium falciparum merozoite antigens
Authors: Heidrich Hans‐G.
Published: February 2003

Merozoite surface proteins are thought to play an important role during the invasion of red blood cells by merozoites. In this article the strategies for the chromatographic isolation and for the functional and molecular characterisation of isolated antigens from freshly harvested Plasmodium falciparum merozoites from cultures are described.
Biology and biochemistry of sexual and sporogenic stages of Plasmodium falciparum: a review
Authors: Meuwissen Joseph H.E.T. , Ponnudurai Thivi
Published: February 2003

Gametocyte development has been studied in synchronous cultures of P. falciparum. These studies have confirmed that maturation of this stage occurs over 7 days and doubling of the DNA content takes place early in the development stage. There appears to be no direct relationship between the number of gametocytes ingested by a mosquito and the oocyst load, nor between the latter and the number of salivary gland sporozoites. The variability in oocyst load could in part be explained by the individual differences in digestion speed of mosquitoes. The number of salivary gland sporozoites and thus also that of sporozoites inoculated per bite of a mosquito is influenced by the number of blood meals which the mosquito has ingested after the infectious blood meal. Moreover, the longer the mosquito survives the more infective it becomes due to increased density of sporozoites in the glands. Anti‐sporozoite antibodies present in the blood meal of an already infected mosquito do not hinder migration of sporozoites to the salivary glands. It has been demonstrated that monoclonal antibodies directed against epitopes of gamete surface of molecular weight of 230, 45/48 and 25 kDa could block sporogony. Transmission blocking antibodies are also present in the sera of individuals exposed to natural transmission. These epitopes of the target proteins are non‐repititive and conformational and thus rendered non‐reactive by reduction. Moreover, some of these surface proteins are glycosylated and/or acylated, and may depend on the presence of the native disulphide bonds for the preservation of immune reactivity. These post‐translational modifications have made screening of gene libraries with immunoproteins difficult. Alternative strategies have been sought to overcome this problem. With regard to the 25 kDa target protein, they have recently been overcome.
Comparison between the molecular characteristics and the potential activity of X and Y nucleolar organizers from various Drosophila melanogaster laboratory lines
Authors: ROSENBERG‐BOURGIN Myriam , PAUMARD Solange , BRACONE Annie , CONTESSE Gérard
Published: February 2003

The molecular characteristics of nucleolar organizers from X and Y chromosomes of different Drosophila melanogaster lines have previously been studied (17).

By analysis of appropriate genetic crosses we show in the present study that the X and Y chromosomes of these lines can confer different degrees of resistance on an inhibitor of ribosomal RNA synthesis (beta exotoxin or thuringiensin) present in the thermostable supernatant of Bacillus thuringiensis cultures. None of the lines studied gives rise to any particular phenotype under normal culture conditions; variations in the degree of supernatant resistance of these lines provide a relative measure of what can be called the potential activity of the nucleolar organizers of the different X and/or Y chromosomes. The potential activity of the Y nucleolar organizers is found to be generally higher than that of the X organizers.

This result can be correlated with the fact that the number of uninterrupted ribosomal units is much greater on the Y chromosomes than on the Xs. Significant variations in potential activity have been shown to occur among the X as also among the Y nucleolar organizers.

Comparison between the molecular characteristics of the nucleolar organizers and their level of activity shows that among the different ribosomal units, only those devoid of insertion interface with the level of activity. However, some of our results could also indicate that not all the uninterrupted units have the same level of activity; this level could be related to the size of the nontranscribed spacer of the ribosomal units.

The peculiarly low level of activity of the Y F71 nucleolar organizer can be explained by genetic rehandlings which could interfere with the degree of activity of the ribosomal genes. Certain results obtained with the X chromosomes do not seem explainable by the molecular characteristics of their nucleolar organizers; conceivably, other genetic factors present on these chromosomes could interfere with the degree of activity of their nucleolar organizers.
Nicotinic and muscarinic responses of embryonic chick ventricular myocardium to acetylcholine: chronotropic and electrophysiological effects
Authors: SUIGNARD‐KHASKIYE Gisèle , RENAUD Didier , LE DOUARIN Georges
Published: February 2003

Sensitivity of 7‐day‐old chick embryo ventricular heart fragments to acetylcholine was investigated. Low doses mainly produced a positive chronotropic effect, whereas high doses of acetylcholine provoked a decrease in the heart beat rhythm. The positive chronotropic effect of acetylcholine was related to the presence of nicotinic receptors that were evidenced within ventricular myocardium by autoradiography. Membrane potential recording showed that acetylcholine hyperpolarizes the diastolic membrane potential when the drug had a negative chronotropic effect. This effect of acetylcholine on the membrane potential was not observed when the drug had a positive chronotropic effect. In many cases, the diastolic membrane potential exhibited spontaneous small depolarizing potentials. Their amplitude was low and their frequency was irregular. These potentials were suppressed by treatment with α‐bungarotoxin, suggesting that they are triggered by nicotinic receptor activation.
Process of re‐establishment of β‐adrenergic induced protein secretion after cytochalasin D inhibition in rat parotid gland. Effect of cholinergic agonist, phorbol ester and calcium
Authors: Huleux Claire , Dreux Catherine , Lemullois Michel , Rossignol Bernard
Published: February 2003

Summary— In rat parotid gland, 3H‐protein secretion is stimulated by β‐adrenergic receptor activation (via cAMP) and also by cholinergic receptor activation (via IP3, calcium and diacylglycerol). The disorganization of microfilament system by cytochalasin D induced an inhibition of β‐adrenergic induced 3H‐protein secretion whereas it did not modify the cholinergic muscarinic one. Cytochalasin D induced the formation of vacuoles in the parotid cell. In this work we show that the activation of muscarinic receptors (with carbachol) partially abolished the inhibitory effect of cytochalasin D on β‐adrenergic induced secretion. Since carbachol induced both intracellular calcium increase and protein kinase C activation, we decided to test separately the effect of calcium (using the calcium ionophore A23187) and protein kinase C activation (using phorbol ester) on the inhibitory effect of cytochalasin D on β‐adrenergic induced secretion. A23187, in the presence of calcium in the external medium was able to partially abolish cytochalasin D effect (ie re‐establishing protein secretion) whereas activation of protein kinase C by phorbol 12–13 di‐butyrate had no effect. These results suggest that protein kinase C is not involved in re‐establishing a ‘normal’ secretion phenomenom whereas calcium does interfere. Furthermore, our fluorescence study shows that, when cytochalasin D is present in the incubation medium, the actin network is disturbed even in the presence of arbachol. This indicates that a calcium entry in the cell is not sufficient to restore a ‘normal’ actin network. Taken altogether, these results show that the total reorganization of the previously disorganized actin network is not necessary to restore β‐adrenergic secretion since carbachol is able to partially abolish the inhibitory effect of cytochalasin D on protein secretion whereas it has no effect on the effect of cytochalasin D on the actin network. However, this study confirms that, in rat parotid gland, the actin network plays an important role in β‐adrenergic transduction mechanisms.
Affinodetection of the sites of formation and of the further distribution of polygalacturonans and native cellulose in growing plant cells
Authors: Vian Brigitte , Roland Jean‐Claude
Published: February 2003

Summary— The object of the present paper is to complement the cytochemical detection of the polysacharides of the plant cell wall and of its precursors, taking benefit of two kinds of affinity methods: the enzyme‐gold technique and immunocytochemistry. Cellobiohydrolase (CBH 1, EC 3.2.1.91) was used to target native crystalline cellulose and two monoclonal antibodies, JIM 5 and JIM 7, were used to target homogalacturonan sequences with various degrees of esterification. Observations were performed at the light microscope level (UV epifluorescence, enzyme‐gold silver staining) and at the electron microscope level. Two types of biological specimens, both in steady state of growth, were chosen: in vitro cultures of melon cells (thin, unidirectional primary walls, loosely associated cells), and elongating zone of mung bean hypocotyl (thick walled and tightly associated cells). The following points were examined successively: the labelling at the histological level, the detection of cellulose and of polygalacturonan components in muro, the visualization of the emerging sites of the polymers along the endomembrane flow and their post‐synthetic modifications (crystallization and methylation respectively). JIM antibodies showed the early labelling of homogalacturonans on the bulging margins of the dictyosomes. The labelled vesicles appeared as sites of polymerization, cytoplasmic transport and beginning of molecular maturation with likely an early action of methyl transferases. The first labelling of cellulose occurred only on the outer face of the plasma‐membrane. Later on, CBH 1‐gold complexes remained distributed throughout the width of the growing wall, despite the surface expansion and the dispersion of the ordered framework. No significant change of the cellulose crystallinity was noticed. A co‐localization of polygalacturonan and cellulose markers was seen from the assembly to the deassembly of the cell wall. In complement, subtractive cytochemistry was performed using PATAg in association with an endopolygalacuronase to split the pectic chains or chelators (EDTA, EGTA, oxalate) to solubilize the calcium‐connected polyuronic acid chains. All the attacks exposed the individual microfibrils of the cellulose framework revealing uniformly the helicoidal organization and confirming that cellulose and polygalacturonans remain closely associated spatially during growth.
Cold‐stress response in the amphibian oocyte: changes in synthesis and nucleocytoplasmic distribution of some proteins
Authors: Moreau Nicole , Lautredou Nicole , N'Da Éléonore , Angelier Nicole
Published: February 2003

Summary— In vivo cold stress was found to induce characteristic changes in the protein synthesis pattern of Pleurodeles waltl oocytes, as analyzed by two‐dimensional gel electrophoresis. The nature of the response varied with the duration and intensity of stress. After a short period of cold stress (12 h at 8°C), synthesis and intracellular distribution of polypeptides were dramatically disturbed. There occured: 1) a reduction in synthesis of several polypeptides, including two major polypeptides (54‐kDa and 47‐kDa); 2) changes in distribution of polypeptides in oocyte, ie some polypeptides (185‐ and 96‐kDa) were blocked in the cytoplasm, while other polypeptides (82‐, 74‐, 72‐ and 68‐kDa, actin and nucleoplasmin) continued to enter the nucleus, but were quantitatively reduced; 3) no changes in the distribution of two nuclear polypeptides (53‐ and 43‐kDa); 4) changes in the relative quantities of β‐ and γ‐actin and preferential migration of γ‐actin towards the nucleus. After a long period of in vivo cold stress (5 days at 8°C), we noted a partial recuperation of synthesis and nuclear migration (except for a 96‐kDa polypeptide), but a persistent perturbation at the level of actin. For more drastic stress conditions (4°C), such a recuperation of protein synthesis was never observed.
Cholesteric liquid crystalline DNA; a comparative analysis of cryofixation methods
Authors: Leforestier Amélie , Livolant Françoise
Published: February 2003

Summary— The ultrastructure of liquid crystalline phases of DNA raises numerous problems because of the structure itself which is fluid and which nature depends on the relative amount of DNA, water and ions. Different cryofixation methods were tested and compared after freeze‐fracture of the specimen. A good ultrastructural preservation of the samples can be achieved without addition of any cryoprotectant by quick‐freezing against a copper block cooled down to liquid helium temperature. Then, molecular orientations can be followed very accurately and the local disorder around a mean direction which exists in the liquid state is kept in the frozen structure.
Sites of transcription of adenovirus type 5 genomes in relation to early viral DNA replication in infected HeLa cells. A high resolution in situ hybridization and autoradiographical study
Authors: Puvion‐Dutilleul Francine , Puvion Edmond
Published: February 2003

Summary— The distribution of viral RNA molecules in HeLa cells infected with adenovirus type 5 (Ad5) was determined by in situ hybridization at the ultrastructural level at an intermediate stage of nuclear transformation, when viral DNA synthetic activities were maximal but progeny viruses were still sparse. Transcription sites of the viral DNA were localized by short pulse, high resolution autoradiography. Nascent viral RNA was found mainly within the nuclear compartment identified at the peripheral replicative zone, which is known to be the main replicative site of Ad5 viral genomes. Viral RNA molecules also were present, but to a markedly lesser extent, within the contiguous single‐stranded (ss) DNA accumulation site, another intranuclear virus‐induced structure in which some replication of viral genomes also occurs. Two other virus‐induced nuclear structures contained viral RNA, the occasional exceptionally enlarged clusters of interchromatin granules and the compact rings, both DNA‐free structures of unknown significance but which might play a role in the process of maturation of the Ad5 primary transcripts. Viral messenger RNA molecules were localized over the large areas of the cytoplasm which contain numerous ribosomes. Our analysis of the effects of various enzymatic pretreatments of the sections of infected cells on the revelation of nascent RNA by in situ hybridization is reviewed.
New data on the microtubule surface lattice
Authors: Chrétien Denis , Wade Richard H.
Published: February 2003

Summary— The in vitro polymerisation of tubulin is a remarkable example of protein self‐assembly in thet several closely related microtubule structures coexist on the polymerisation plateau. Unfixed and unstained in vitro assembled microtubules were observed in vitreous ice by cryo‐electron microscopy. New results are reported that considerably extend previous observations [47]. In ice, microtubule images have a distinctive contrast related to the number and skew of the photofilaments. The microtubules observed have from twelve to seventeen protofilaments. Comparison with thin sections of pelleted material allows a direct identification of images from microtubules with thirteen, fourteen and fifteen protofilaments. A surface lattice accommodation mechanism, previously proposed to explain how variable numbers of protofilaments can be incorporated into the basic thirteen protofilament structure, is described in detail. Our new experimental results are shown to be in overall agreement with the theoretical predictions. Only thirteen protofilament microtubules have unskewed protofilaments, this was confirmed by observations on axoneme fragments. The results imply that the microtubule surface lattice is based on a mixed packing which combines features of the standard A and B lattices.
Dinoflagellate flagella adopt various conformations in response to different needs
Authors: Cachon Monique , Cachon Jean , Cosson Jacky , Greuet Claude , Huitorel Philippe
Published: February 2003

Summary— The two flagella of Dinoflagellates have, up to now, been poorly described. They display different structures and different patterns of behaviour compared with other organisms. In addition, the two flagella are different from each other: the transverse flagellum is ribbon‐shaped and beats with a spiral undulation inside a furrow located around the cell body while the longitudinal flagellum has a larger diameter than simple flagella because it contains structures in addition to the axoneme and propagates essentially sinusoidal waves to push the cell. Ceratium flagella are particularly interesting to study because they both show different types of movements and have complex structures in addition to the axoneme. We propose that the additional structures are responsible for the particular movements of Dinoflagellate flagella. The presence of food particles in vacuoles in the vicinity of the flagella pocket suggests that their flagellar apparatus may not only be a propulsive organelle but could also be involved in prey capture.
Evolutionary conservation of an epitope associated with striated rootlets in different epithelial ciliated cells
Authors: Lemullois Michel , Klotz Catherine , Sandoz Daniel
Published: February 2003

Summary— In ciliated cells of metazoa, striated rootlets associated with basal bodies anchor the ciliary apparatus to the cytoskeleton. We have used here a monoclonal antibody against a 175 kDa protein associated with the striated rootlets of quail ciliated cells [14], to study ciliated cells of different species. In mussel gill epithelium the antibody recognized a protein of 92 kDa which shows a periodic distribution along the striated rootlets. In frog ciliated palate epithelium, two different rootlets are associated with basal bodies, both are decorated and only one protein of 48 kDa is recognized on immunoblot. The antigen is arranged in a helix around the striated rootlets. In rabbit ciliated oviduct epithelium, we detected the presence of very small and thin rootlets which are weakly labeled. We have shown that an epitope associated with the striated rootlets is preserved through evolution although the molecular weight of the peptide varies. We have also observed the appearance of this epitope on protein associated with junctional complexes in rabbit and cytoskeleton component in quail oviduct.
Scaffold‐associated regions and repeated or cross‐hybridizing sequences on an 800 kilobase DNA stretch of the Drosophila× chromosome
Authors: Surdej Patrick , Brandli Dominique , Miassod Raymond
Published: February 2003

Summry— DNA fragments binding to the scaffold (SARs) have been previously mapped on an 800‐kb DNA fragment from the Drosophila × chromosome. Here we have examined whether these 86 DNA fragments bear sequences repeated in the Drosophila genome and/or cross‐hybridizing sequences. Twenty‐two middle‐repeated sequences were localized by hybridizing Southern transfers of representative cloned DNA with total genomic DNA and, reciprocally, by hybridizing Southern transfers of total genomic DNA with this cloned DNA. Seventy‐nine out of the 86 SAR‐containing fragments appeared to be distinct from these 22 repeated motifs. Therefore, SARs, in their vast majority, are not members of middle‐repeated sequence families. However, the 22 middle‐repeated sequences were shown to be significantly located in the near vicinity of SARs. Hybridizations were also performed between SAR‐containing DNA fragments, either at a high or at a low stringency. At a high stringency, the 86 SAR‐containing DNA fragments did not cross‐hybridize. However, at a low stringency, a complex hybridization network was evidenced. These nucleotide sequence similitudes support the idea that the various SARs may play common roles.
Effects of low temperature on the formation of secretion granules in the Golgi apparatus of granular cells of the frog urinary bladder
Authors: Rambourg Alain , Clermont Yves , Pisam Monique , Ripoche Pierre
Published: February 2003

Summry— The formation of secretion granules has been studied in the Golgi apparatus of granular epithelial cells of frog urinary bladders maintained at room temperature or cooled at 4°C for various lengths of time. In control animals, the Golgi apparatus was composed of the following stacked elements: subjacent to the cis‐rmelement mad up of anastomosed tubules, two elements in the mid‐compartment consisted of flattened saccules interconnected by tubules. On the trans‐face, two or three sacculo‐tubular elements were slightly dilated by an electron dense granular material. In the trans‐Golgi elements, this material was segregated into dilatations of various sizes and shapes which are continuous with flattened portions devoid of stained material. In the trans‐Golgi region, free irregular progranules, seemingly formed by rupture of the trans‐most Golgi elements. In granular cells examined after 4 h at 4°C, all Golgi compartments were affected by the low temperature. The cis‐half portion of the Golgi apparatus consisted mainly of anastomosed membranous tubules and the cis‐element was no longer recognizable. The trans‐compartment was reduced to a few flattened saccules with progranules hardly visible on their trans‐aspect. At later time intervals, there was a progressive reconstitution of the cis‐zone while saccular elements started to pile up in the trans‐compartment. At 24 h, the trans‐compartment comprised six to eight saccular elements which showed irregular dilatations filled with granular material separated by large flattened portions. These various observations were interpreted as indicating that the trans‐compartment was a dynamic structure undergoing continuous renewal.