bluuebunny/crossref_metadata_2025 huggingface dataset:
Graded phenotypic response to partial and complete deficiency of a brain-specific transcript variant of the winged helix transcription factor RFX4
Authors: Blackshear Perry J. , Graves Joan P. , Stumpo Deborah J. , Cobos Inma , Rubenstein John L. R. , Zeldin Darryl C.
Published: August 2003

One line of mice harboring a cardiac-specific epoxygenase transgene developed head swelling and rapid neurological decline in young adulthood, and had marked hydrocephalus of the lateral and third ventricles. The transgene was found to be inserted into an intron in the mouse Rfx4 locus. This insertion apparently prevented expression of a novel variant transcript of RFX4 (RFX4_v3), a member of the regulatory factor X family of winged helix transcription factors. Interruption of two alleles resulted in profound failure of dorsal midline brain structure formation and perinatal death,presumably by interfering with expression of downstream genes. Interruption of a single allele prevented formation of the subcommissural organ, a structure important for cerebrospinal fluid flow through the aqueduct of Sylvius, and resulted in congenital hydrocephalus. These data implicate the RFX4_v3 variant transcript as being crucial for early brain development, as well as for the genesis of the subcommissural organ. These findings may be relevant to human congenital hydrocephalus, a birth defect that affects ∼0.6 per 1000 newborns.
Mice humanised for the EGF receptor display hypomorphic phenotypes in skin, bone and heart
Authors: Sibilia Maria , Wagner Bettina , Hoebertz Astrid , Elliott Candace , Marino Silvia , Jochum Wolfram , Wagner Erwin F.
Published: August 2003

Mice lacking the epidermal growth factor receptor (EGFR) develop epithelial defects and a neurodegenerative disease and die within the first month of birth. By employing a conditional knock-in approach using the human EGFR cDNA mice humanised for EGFR (hEGFRKI/KI)were generated. Homozygous hEGFRKI/KI mice are viable and live up to six months. However, these mice are growth retarded and show skin and hair defects similar to Egfr-/- mutants. Interestingly, the neurodegeneration is fully rescued in hEGFRKI/KI mice, however, they develop a severe heart hypertrophy with semilunar valve abnormalities. Moreover, hEGFRKI/KI mice display accelerated chondrocyte and osteoblast differentiation, a phenotype that is also present in Egfr-/- mice and has not been previously described. The severity of the phenotypes correlates with the expression levels of the hEGFRKI allele, which is not efficiently expressed in epithelial and bone cells, but is expressed at similar and even higher levels as the endogenous Egfr in brain and heart. These results demonstrate that mice humanised for EGFR display tissue-specific hypomorphic phenotypes and describe a novel function for EGFR in bone development.
Crucial roles of Brn1 in distal tubule formation and function in mouse kidney
Authors: Nakai Shigeyasu , Sugitani Yoshinobu , Sato Hiroshi , Ito Sadayoshi , Miura Yukio , Ogawa Masaharu , Nishi Miyuki , Jishage Kou-ichi , Minowa Osamu , Noda Tetsuo
Published: August 2003

This study identifies a role for the gene for the POU transcription factor Brn1 in distal tubule formation and function in the mammalian kidney. Normal development of Henle's loop (HL), the distal convoluted tubule and the macula densa was severely retarded in Brn1-deficient mice. In particular,elongation and differentiation of the developing HL was affected. In the adult kidney, Brn1 was detected only in the thick ascending limb (TAL) of HL. In addition, the expression of a number of TAL-specific genes was reduced in the Brn1+/- kidney, including Umod,Nkcc2/Slc12a1, Bsnd, Kcnj1 and Ptger3. These results suggest that Brn1 is essential for both the development and function of the nephron in the kidney.
A defect in a novel ADAMTS family member is the cause of the belted white-spotting mutation
Authors: Rao Cherie , Foernzler Dorothee , Loftus Stacie K. , Liu Shanming , McPherson John D. , Jungers Katherine A. , Apte Suneel S. , Pavan William J. , Beier David R.
Published: August 2003

Several features of the pigment defect in belted (bt) mutant mice suggest that it occurs as a result of a defect in melanocyte development that is unique from those described for other classical white-spotting mutations. We report here that bt mice carry mutations in Adamts20, a novel member of the ADAMTS family of secreted metalloproteases. Adamts20 shows a highly dynamic pattern of expression in the developing embryo that generally precedes the appearance of melanoblasts in the same region, and is not expressed in the migrating cells themselves. Adamts20 shows remarkable homology with GON-1, an ADAMTS family protease required for distal tip cell migration in C. elegans. Our results suggest that the role of ADAMTS proteases in the regulation of cell migration has been conserved in mammalian development.
Spatial and temporal patterns of ERK signaling during mouse embryogenesis
Authors: Corson Laura Beth , Yamanaka Yojiro , Lai Ka-Man Venus , Rossant Janet
Published: August 2003

Signaling between tissues is essential to form the complex,three-dimensional organization of an embryo. Because many receptor tyrosine kinases signal through the RAS-MAPK pathway, phosphorylated ERK can be used as an indicator of when and where signaling is active during development. Using whole-mount immunohistochemistry with antibodies specific to phosphorylated ERK1 and ERK2, we analyzed the location, timing, distribution, duration and intensity of ERK signaling during mouse embryogenesis (5-10.5 days postcoitum). Spatial and temporal domains of ERK activation were discrete with well-defined boundaries, indicating specific regulation of signaling in vivo. Prominent, sustained domains of ERK activation were seen in the ectoplacental cone, extra-embryonic ectoderm, limb buds, branchial arches, frontonasal process, forebrain, midbrain-hindbrain boundary, tailbud, foregut and liver. Transient activation was seen in neural crest, peripheral nervous system,nascent blood vessels, and anlagen of the eye, ear and heart. In the contiguous domains of ERK signaling, phospho-ERK staining was cytoplasmic with no sign of nuclear translocation. With few exceptions, the strongest domains of ERK activation correlated with regions of known or suspected fibroblast growth factor (FGF) signaling, and brief incubation with an inhibitor of the fibroblast growth factor receptor (FGFR) specifically diminished the phospho-ERK staining in these regions. Although many domains of ERK activation were FGFR-dependent, not all domains of FGF signaling were phospho-ERK positive. These studies identify key domains of sustained ERK signaling in the intact mouse embryo, give significant insight into the regulation of this signaling in vivo and pinpoint regions where downstream target genes can be sought.
Modulation of EGF receptor-mediated vulva development by the heterotrimeric G-protein Gαq and excitable cells inC.elegans
Authors: Moghal Nadeem , Garcia L. Rene , Khan Liakot A. , Iwasaki Kouichi , Sternberg Paul W.
Published: August 2003

The extent to which excitable cells and behavior modulate animal development has not been examined in detail. Here, we demonstrate the existence of a novel pathway for promoting vulval fates in C. elegansthat involves activation of the heterotrimeric Gαq protein, EGL-30. EGL-30 acts with muscle-expressed EGL-19 L-type voltage-gated calcium channels to promote vulva development, and acts downstream or parallel to LET-60 (RAS). This pathway is not essential for vulval induction on standard Petri plates,but can be stimulated by expression of activated EGL-30 in neurons, or by an EGL-30-dependent change in behavior that occurs in a liquid environment. Our results indicate that excitable cells and animal behavior can provide modulatory inputs into the effects of growth factor signaling on cell fates,and suggest that communication between these cell populations is important for normal development to occur under certain environmental conditions.
Zebrafish fgf24functions withfgf8to promote posterior mesodermal development
Authors: Draper Bruce W. , Stock David W. , Kimmel Charles B.
Published: August 2003

Fibroblast growth factor (Fgf) signaling plays an important role during development of posterior mesoderm in vertebrate embryos. Blocking Fgf signaling by expressing a dominant-negative Fgf receptor inhibits posterior mesoderm development. In mice, Fgf8 appears to be the principal ligand required for mesodermal development, as mouse Fgf8 mutants do not form mesoderm. In zebrafish, Fgf8 is encoded by the acerebellarlocus, and, similar to its mouse otholog, is expressed in early mesodermal precursors during gastrulation. However, zebrafish fgf8 mutants have only mild defects in posterior mesodermal development, suggesting that it is not the only Fgf ligand involved in the development of this tissue. We report here the identification of an fgf8-related gene in zebrafish, fgf24, that is co-expressed with fgf8 in mesodermal precursors during gastrulation. Using morpholino-based gene inactivation, we have analyzed the function of fgf24 during development. We found that inhibiting fgf24 function alone has no affect on the formation of posterior mesoderm. Conversely, inhibiting fgf24 function in embryos mutant for fgf8 blocks the formation of most posterior mesoderm. Thus, fgf8 and fgf24 are together required to promote posterior mesodermal development. We provide both phenotypic and genetic evidence that these Fgf signaling components interact with no tailand spadetail, two zebrafish T-box transcription factors that are required for the development of all posterior mesoderm. Last, we show that fgf24 is expressed in early fin bud mesenchyme and that inhibiting fgf24 function results in viable fish that lack pectoral fins.
Dissimilar regulation of cell differentiation in mesencephalic (cranial)and sacral (trunk) neural crest cells in vitro
Authors: Abzhanov Arhat , Tzahor Eldad , Lassar Andrew B. , Tabin Clifford J.
Published: August 2003

During development neural crest cells give rise to a wide variety of specialized cell types in response to cytokines from surrounding tissues. Depending on the cranial-caudal level of their origin, different populations of neural crest cells exhibit differential competence to respond to these signals as exemplified by the unique ability of cranial neural crest to form skeletal cell types. We show that in addition to differences in whether they respond to particular signals, cranial neural crest cells differ dramatically from the trunk neural crest cells in how they respond to specific extracellular signals, such that under identical conditions the same signal induces dissimilar cell fate decisions in the two populations in vitro. Conversely, the same differentiated cell types are induced by different signals in the two populations. These in vitro differences in neural crest response are consistent with in vivo manipulations. We also provide evidence that these differences in responsiveness are modulated, at least in part, by differential expression of Hox genes within the neural crest.
Oligodendrocyte ablation impairs cerebellum development
Authors: Mathis Carole , Collin Ludovic , Borrelli Emiliana
Published: August 2003

Oligodendrocytes (OLs) are the glial cells of the central nervous system and are classically known to form myelin sheaths around most axons of higher vertebrates. Whether these cells might have other roles, in particular during development, has not been studied. Taking advantage of a transgenic mouse model in which OLs can be selectively killed in a desired time-frame, we have investigated the impact of OL ablation on cerebellar development. OL ablation was induced during the first 3 postnatal weeks, a time at which cerebellum development is ongoing. Strikingly, OL ablation triggers a profound perturbation of the known cerebellum developmental program, characterized by the disorganization of the cortical layers, abnormal foliation and a complete alteration of Purkinje cell dendritic arborization and axonal fasciculation. This phenotype is accompained by decreased granule cell density, a disorganized Bergmann glia network and impaired migration of interneurons in the molecular layer. These results demonstrate a previously ignored role of OLs in the formation of the cerebellar cytoarchitecture.
Lack of Bdnf and TrkB signalling in the postnatal cochlea leads to a spatial reshaping of innervation along the tonotopic axis and hearing loss
Authors: Schimmang Thomas , Tan Justin , Müller Marcus , Zimmermann Ulrike , Rohbock Karin , Köpschall Iris , Limberger Annette , Minichiello Liliana , Knipper Marlies
Published: August 2003

Members of the neurotrophin gene family and their high-affinity Trk receptors control innervation of the cochlea during embryonic development. Lack of neurotrophin signalling in the cochlea has been well documented for early postnatal animals, resulting in a loss of cochlear sensory neurones and a region-specific reduction of target innervation along the tonotopic axis. However, how reduced neurotrophin signalling affects the innervation of the mature cochlea is currently unknown. Here, we have analysed the consequences of a lack of the TrkB receptor and its ligand, the neurotrophin brain-derived neurotrophic factor (Bdnf), in the late postnatal or adult cochlea using mouse mutants. During early postnatal development, mutant animals show a lack of afferent innervation of outer hair cells in the apical part of the cochlea,whereas nerve fibres in the basal part are maintained. Strikingly, this phenotype is reversed during subsequent maturation of the cochlea, which results in a normal pattern of outer hair cell innervation in the apex and loss of nerve fibres at the base in adult mutants. Measurements of auditory brain stem responses of these mice revealed a significant hearing loss. The observed innervation patterns correlate with opposing gradients of Bdnf and Nt3 expression in cochlear neurones along the tonotopic axis. Thus, the reshaping of innervation may be controlled by autocrine signalling between neurotrophins and their receptors in cochlear neurones. Our results indicate a substantial potential for re-innervation processes in the mature cochlea,which may also be of relevance for treatment of hearing loss in humans.
senselessis necessary for the survival of embryonic salivary glands inDrosophila
Authors: Chandrasekaran Vidya , Beckendorf Steven K.
Published: August 2003

Apoptosis in developing Drosophila embryos is rare and confined to specific groups of cells. We explain how one organ, salivary glands, of Drosophila embryos avoids apoptosis. senseless(sens), a Zn-finger transcription factor, is expressed in the salivary primordium and later in the differentiated salivary glands. The regulation of sens expression in the salivary placodes is more complex than observed in the embryonic PNS. We have shown that sensexpression is initiated in the salivary placodes by fork head(fkh), a winged helix transcription factor. The expression of sens is maintained in the salivary glands by fkh and by daughterless (da), a bHLH family member. In this study, we have identified sage, a salivary-specific bHLH protein as a new heterodimeric partner for da protein in the salivary glands. In addition, our data suggest that sage RNAi embryos have a phenotype similar to sens and that sage is necessary to maintain expression of sens in the embryonic salivary glands. Furthermore, we show that in the salivary glands, sens acts as an anti-apoptotic protein by repressing reaper and possibly hid.
TheDrosophilaproneural geneamospromotes olfactory sensillum formation and suppresses bristle formation
Authors: Lage Petra I. zur , Prentice David R. A. , Holohan Eimear E. , Jarman Andrew P.
Published: August 2003

Proneural genes encode basic-helix-loop-helix (bHLH) transcription factors required for neural precursor specification. Recently amos was identified as a new candidate Drosophila proneural gene related to atonal. Having isolated the first specific amosloss-of-function mutations, we show definitively that amos is required to specify the precursors of two classes of olfactory sensilla. Unlike other known proneural mutations, a novel characteristic of amos loss of function is the appearance of ectopic sensory bristles in addition to loss of olfactory sensilla, owing to the inappropriate function of scute. This supports a model of inhibitory interactions between proneural genes, whereby ato-like genes (amos and ato) must suppress sensory bristle fate as well as promote alternative sense organ subtypes.
Physical properties of Tld, Sog, Tsg and Dpp protein interactions are predicted to help create a sharp boundary in Bmp signals during dorsoventral patterning of theDrosophilaembryo
Authors: Shimmi Osamu , O'Connor Michael B.
Published: August 2003

Dorsal cell fate in Drosophila embryos is specified by an activity gradient of Decapentaplegic (Dpp), a homologue of bone morphogenetic proteins(Bmps) 2/4. Previous genetic and biochemical studies have revealed that the Sog, Tsg and Tld proteins modify Dpp activity at the post-transcriptional level. The predominant view is that Sog and Tsg form a strong ternary complex with Dpp that prevents it from binding to its cognate receptors in lateral regions of the embryo, while in the dorsalmost cells Tld is proposed to process Sog and thereby liberate Dpp for signaling. In this model, it is not readily apparent how Tld activity is restricted to the dorsal-most cells as it is expressed throughout the entire dorsal domain. In this study, additional genetic and biochemical assays were developed to further probe the relationships between the Sog, Tsg, Tld and Dpp proteins. Using cell based assays, we find that the dynamic range over which Dpp functions for signaling is the same range in which Dpp stimulates the cleavage of Sog by Tld. In addition, our data supports a role for Tsg in sensitizing the patterning mechanism to low levels of Dpp. We propose that the strong Dpp concentration dependence exhibited by the processing reaction, together with movement of Dpp by Sog and Tsg protein can help explain how Tld activity is confined to the dorsal-most region of the embryo through formation of a spatially dependent positive and negative reinforcement loop. Such a mechanism also explains how a sharp rather than smooth signaling boundary is formed.
The concerted action of Meox homeobox genes is required upstream of genetic pathways essential for the formation, patterning and differentiation of somites
Authors: Mankoo Baljinder S. , Skuntz Susan , Harrigan Ian , Grigorieva Elena , Candia Al , Wright Christopher V. E. , Arnheiter Heinz , Pachnis Vassilis
Published: August 2003

The paraxial mesoderm of the somites of the vertebrate embryo contains the precursors of the axial skeleton, skeletal muscles and dermis. The Meox1 and Meox2 homeobox genes are expressed in the somites and their derivatives during embryogenesis. Mice homozygous for a null mutation in Meox1 display relatively mild defects in sclerotome derived vertebral and rib bones, whereas absence of Meox2 function leads to defective differentiation and morphogenesis of the limb muscles. By contrast, mice carrying null mutations for both Meox genes display a dramatic and wide-ranging synthetic phenotype associated with extremely disrupted somite morphogenesis, patterning and differentiation. Mutant animals lack an axial skeleton and skeletal muscles are severely deficient. Our results demonstrate that Meox1 and Meox2 genes function together and upstream of several genetic hierarchies that are required for the development of somites. In particular, our studies place Meox gene function upstream of Pax genes in the regulation of chondrogenic and myogenic differentiation of paraxial mesoderm.
The FORKED genes are essential for distal vein meeting inArabidopsis
Authors: Steynen Quintin J. , Schultz Elizabeth A.
Published: August 2003

As in most dicotyledonous plants, the leaves and cotyledons of Arabidopsis have a closed, reticulate venation pattern. This pattern is proposed to be generated through canalization of the hormone auxin. We have identified two genes, FORKED 1 (FKD1) and FORKED 2 (FKD2),that are necessary for the closed venation pattern: mutations in either gene result in an open venation pattern that lacks distal meeting. In fkd1leaves and cotyledons, the defect is first evident in the provascular tissue,such that the distal end of the newly forming vein does not connect to the previously formed, more distal vein. Plants doubly mutant for both genes have widespread defects in leaf venation, suggesting that the genes function in an overlapping manner at the distal junctions, but act redundantly throughout leaf veins. Expression of an auxin responsive reporter gene is reduced in fkd1 leaves, suggesting that FKD1 is necessary for the auxin reponse that directs vascular tissue development. The reduction in reporter gene expression and the fkd1 phenotype are relieved in the presence of auxin transport inhibition. The restoration of vein junctions in situations where auxin concentrations are increased indicates that distal vein junctions are sites of low auxin concentration and are particularly sensitive to reduced FKD1 and FKD2 activity.
Frizzled signalling and cell polarisation inDrosophilaand vertebrates
Authors: Strutt David
Published: August 2003

A key aspect of animal development is the appropriate polarisation of different cell types in the right place at the right time. Such polarisation is often precisely coordinated relative to the axes of a tissue or organ, but the mechanisms underlying this coordination are still poorly understood. Nevertheless, genetic analysis of animal development has revealed some of the pathways involved. For example, a non-canonical Frizzled signalling pathway has been found to coordinate cell polarity throughout the insect cuticle, and recent work has implicated an analogous pathway in coordinated polarisation of cells during vertebrate development. This review discusses recent findings regarding non-canonical Frizzled signalling and cell polarisation.
Thermal elastic-plastic stress analysis of an anisotropic structure
Authors: Wang X , Dong M-Ch , Lu G
Published: August 2003

In this paper, a polynomial stress function is utilized to satisfy both the governing differential equation for an anisotropic plane stress problem and the corresponding boundary conditions for plastic deformation. A theoretical solution for the thermal elastic-plastic problem of composite structure is obtained by means of the Tsai-Hill strength theory of anisotropic material. The composite structure is composed of a steel fibre-reinforced aluminium metal-matrix with a linear hardening material property. On the other hand, an elastic-plastic finite element analysis for the same problem is also carried out by using ABAQUS. The theoretical solution is in good agreement with the results from the finite element analysis. Finally, some examples are given and the corresponding results are discussed.
Making the eye blink—modelling the operation of the Gateshead Millennium Bridge
Authors: Tilley D G , Keogh P S , Mutch K , Robotham M , Curran P , Mehrkar-Asl S
Published: August 2003

This paper considers the system dynamics associated with a hydraulic system moving a large flexible structure, namely the Gateshead Millennium Bridge. The hydraulic components possess a range of non-linear characteristics, which become coupled to the bridge structural dynamics. The bridge structure is lightly damped and may be represented by a linear model, provided that the amplitudes of vibrational modes are small enough to maintain positive cable tensions. The limiting amplitudes were known at the design stage and the hydraulic circuit had to ensure that these were not exceeded during operation, which may include various conditions of wind loading, emergency stops and variability of components. System design was achieved through an extensive iterative process of simulation that included all operational procedures to eliminate problematic circuits that would have caused instability or excessive bridge oscillations. The actual bridge and coupled hydraulic circuits underwent a series of commissioning trials and no undue problems were experienced. This indicated that the simulation procedures developed greatly reduced the risks associated with a unique structure actuated by a bespoke hydraulic circuit.
Preliminary investigations on non-linear fluid-structure interaction of an offshore structure
Authors: Sudharsan N M , Kantharaj M , Kumar K
Published: August 2003

Fluid-structure interaction (FSI) of offshore structures is an important area of research. It is also of significant industrial value, where such capabilities could be used for simulating the structural response in numerical wave tanks. The current research focuses on the development of a numerical wave tank with FSI capabilities. This requires solving the non-linear fluid and structure behaviour in the time domain considering a non-linear velocity potential formulation for the wave flow and a linear elastic formulation for the structure. The fluid and structural behaviour are coupled in time through a mixed Lagrangian-Eulerian procedure and the differential equations are discretized using the finite element method. Unstructured triangular meshes are used to discretize the fluid and structural domain, augmented by mesh smoothing algorithms. Based on the studie, it is shown that the transient force developed by the fluid on the structure varies due to the elastic nature of the structure. The perturbations in the free-surface profile due to the structural response are also captured. It is also shown that transient non-linear studies are important as they can help in identifying critical design limitations of traditional guides. It is thus demonstrated that the present work is capable of solving such FSI problems in order to obtain a reliable and complete analysis leading to a better understanding and safer design of compliant structures. The effect on the structure due to prolonged interaction with the fluid needs to be further studied for a better understanding.
Elastohydrodynamic lubrication analysis of hip implants with ultra high molecular weight polyethylene cups under transient conditions
Authors: Jalali-Vahid D , Jin Z M , Dowson D
Published: August 2003

The transient variation of both the load and speed experienced during walking in an elastohydrodynamic lubrication (EHL) analysis for artificial hip joints employing an ultra high molecular weight polyethylene (UHMWPE) acetabular cup against either a metallic or ceramic femoral head was considered in this study. A general numerical procedure to solve the transient EHL problem in spherical ball-in-socket coordinates, developed in a previous study by Jalali-Vahid and Jin in 2002, was applied under three specific conditions experienced during typical gait cycles, including speed reversal, a sudden load increase and a sudden load decrease. The predicted minimum film thickness was found to stay remarkably constant and similar to that prior to the change in either the load or the angular velocity, despite a large change in these operating conditions. This was attributed to the remarkably effective squeeze-film action of preserving and maintaining the lubricating film developed before the transient variations in either the load or speed. It is therefore possible to neglect the effect of these specific transient variations of load and speed under physiological walking conditions considered in the present study on the predicted film thickness in hip implants with UHMWPE cups.
Configuration analysis of the Schatz linkage
Authors: Lee C-C , Dai J S
Published: August 2003

This paper investigates different configurations of the Schatz linkage based on the analysis of a reciprocal screw and of the relationship between the reciprocal screw and its stem-screw system, which consists of twists of freedom located at six revolute joints of the linkage. A new method of using cofactors of an augmenting screw is used to obtain the reciprocal screw. The stem-screw system of order 5 of the linkage is developed from the special geometry of the six revolute joints and closed-form displacement solutions are provided based on the stem-screw system. The screw surface representing the trajectory of the reciprocal screw with a chosen pitch is established. This surface is then used to characterize zero-pitch reciprocal screw configurations. The special relationship between the reciprocal screw and the stem-screw system is analysed and used to characterize the constraint wrench and configurations created by the changes of the constraint wrench with a non-zero pitch. Consequently, a set of configurations are presented in conjunction with a ruled surface produced by the progression of the constraint wrench when the linkage drive joint rotates from 5° to 85°.
Feature investigation of a cracked rotor with initial deflection
Authors: Zou J , Chen J , Dong G M
Published: August 2003

The dynamic equation of a cracked rotor with initial deflection is modelled, which is based on the simple hinge crack model; the numerical simulation solutions are obtained from the model. The subharmonic resonance of the cracked rotor is analysed and the frequency components of the cracked rotor with initial deflection and without initial deflection are investigated. By simulation research, the influence of the stiffness variation, the initial deflection, the unbalance, the unbalance angle and the orientation of the initial deflection on the vibrational characteristics is studied, which can be used as the criterion for identification of a cracked rotor.
Laser shortpulse heating of a gold-chromium-gold multilayer assembly
Authors: Yilbas B S , Arif A F M
Published: August 2003

Laser shortpulse heating of metallic substrates results in non-equilibrium energy transport in the region irradiated by a laser beam. Although the heating duration is short, thermal strain developed due to high-temperature gradients results in high stress levels in this region. In the present study, laser shortpulse heating of a gold-chromium-gold (Au-Cr-Au) multilayer assembly is considered. An electron kinetic theory is introduced when modelling the non-equilibrium energy transport while elastoplastic analysis is carried out to obtain thermal stresses inside the multilayer assembly. A numerical scheme is introduced to discretize the governing equations while the finite element method is employed for the solution of the stress field. It is found that temperature in the Cr layer is higher than the Au layer in the surface region. This is because of a high electron-phonon coupling factor of Cr. The stress levels below the elastic limit of the substrate material are obtained. The displacement of the surface at the irradiated spot centre is in the order of 10−9-10−7m.
Numerical study on the bubbly flow around a hydrofoil in pitching and heaving motions
Authors: Uchiyama T
Published: August 2003

The air-water bubbly flow around a hydrofoil of NACA65-010, undergoing the heaving and pitching motions in the uniform flow on a horizontal plane, is simulated by an incompressible two-fluid model. The finite element method proposed in a prior paper is applied to solve the model. The Reynolds number defined by the volumetric velocity of the water is 10000 and the volumetric fraction of the air upstream of the hydrofoil, αg0, ranges from 0 to 0.06. The simulation reveals the effects of the αg0 value, the phase difference between the heaving and pitching motions, and the oscillating frequency. The propulsive performance is also discussed in relation to the time variation of the flow properties around the hydrofoil.
Stress in a flat section band clamp
Authors: Shoghi K , Rao H V , Barrans S M
Published: August 2003

This paper analyses the stress in a flat section band clamp and validates by experimental data the predictions based on the developed theory. In the experimental work flat section band clamps were positioned around the rigid cylinder and strain and corresponding displacement were measured; the clamp nut was tightened to gradually increasing torque until failure occurred. The error in stress and corresponding displacement predicted by the theory and that obtained by testing was found to be 1–2 per cent for the initial stage where the band is slack on the rigid cylinder, 3–5 per cent where the band makes contact with the cylinder and 5–7 per cent for post-yielding of the band material. This error could mainly be attributed to an uncertainty in the properties for the material and the accuracy of instrumentation. The coefficient of friction between the relevant components and the elastic modulus of the band material are the important properties, which influence the performance of the clamp. In the current experimental work failure occurred in the T-bolt.
Prediction of drop size distribution in a spray from a pressure swirl atomizer using maximum entropy formalism
Authors: Mondal D , Datta A , Sarkar A
Published: August 2003

Drop size distribution is an important characterizing parameter of a spray. In the present work a theoretical model has been described, based on the maximum entropy formalism principle, for the determination of the drop size distribution in a spray issued from a pressure swirl atomizer. The atomization efficiency is also derived from the model, assuming the velocities of all the drop sizes to be uniform. The results show that the drop size distribution, described from the present model, resembles the Rosin-Rammler type distribution very well, with a dispersion parameter of 3.47. The atomization efficiency is found to decrease with the increase in liquid mass flowrates, when the pressure differential across the atomizer remains the same. On the other hand, an increase in the orifice diameter increases the atomization efficiency, when the liquid mass flow rate and pressure differential are the same. The ratio of the surface energy to the kinetic energy at the atomizer exit is seen to influence the atomization efficiency.
Determining the surface form of polystyrene through the coordinate measurement machine
Authors: Aitchison D , Sulaiman R
Published: August 2003

The market for foam materials has been growing rapidly throughout the world as they have a variety of uses. Some examples are in the automotives industries, food packaging industries, medical application, sports gears, home insulations and floatation in offshore drilling rigs, buoys and small boats. Since the uses of foam affects greatly the daily lives of humans, the need to have foams in different shapes requires speed in cutting and manufacture. This can only be done through computer aided cutting machines or automated cutting of foams. However, the speed of cutting will affect the surface finish of the cut. Therefore, it is necessary to determine the surface form of the polystyrene to achieve quality results. This is an on-going research to produce a rapid-prototyping machine that cuts foam models. The first phase of this research is to determine the surface form of polystyrene through the use of a coordinate measuring machine (CMM), after being cut with different types of wires, at different temperatures and cutting feedrates.
Rapidly Increasing Concentrations of CFC Alternatives (HFC-134a, HCFC-141b, and HCFC-142b) in the Atmosphere as Observed in Hokkaido and Antarctica
Authors: Shirai Tomoko , Makide Yoshihiro
Published: September 2003

Sharply increasing trends were observed for recent atmospheric concentrations of CFC alternatives by GC/MS analysis of samples collected in Hokkido, Japan and Syowa Station, Antarctica. These trends were consistent with those calculated based on the world production/release statistics for HFC-134a and HCFC-141b, but inconsistent for HCFC-142b, suggestive of some inaccurate release estimations.
siRNA function in RNAi: A chemical modification analysis
Authors: CHIU YA-LIN , RANA TARIQ M.
Published: August 2003

Various chemical modifications were created in short-interfering RNAs (siRNAs) to determine the biochemical properties required for RNA interference (RNAi). Remarkably, modifications at the 2′-position of pentose sugars in siRNAs showed the 2′-OHs were not required for RNAi, indicating that RNAi machinery does not require the 2′-OH for recognition of siRNAs and catalytic ribonuclease activity of RNA-induced silencing complexes (RISCs) does not involve the 2′-OH of guide antisense RNA. In addition, 2′ modifications predicted to stabilize siRNA increased the persistence of RNAi as compared with wild-type siRNAs. RNAi was also induced with chemical modifications that stabilized interactions between A–U base pairs, demonstrating that these types of modifications may enhance mRNA targeting efficiency in allele-specific RNAi. Modifications altering the structure of the A-form major groove of antisense siRNA–mRNA duplexes abolished RNAi, suggesting that the major groove of these duplexes was required for recognition by activated RISC*. Comparative analysis of the stability and RNAi activities of chemically modified single-stranded antisense RNA and duplex siRNA suggested that some catalytic mechanism(s) other than siRNA stability were linked to RNAi efficiency. Modified or mismatched ribonucleotides incorporated at internal positions in the 5′ or 3′ half of the siRNA duplex, as defined by the antisense strand, indicated that the integrity of the 5′ and not the 3′ half of the siRNA structure was important for RNAi, highlighting the asymmetric nature of siRNA recognition for initiation of unwinding. Collectively, this study defines the mechanisms of RNAi in human cells and provides new rules for designing effective and stable siRNAs for RNAi-mediated gene-silencing applications.
Localization of nuclear retained mRNAs in Saccharomyces cerevisiae
Authors: THOMSEN RUNE , LIBRI DOMENICO , BOULAY JOCELYNE , ROSBASH MICHAEL , JENSEN TORBEN HEICK
Published: August 2003

In the yeast Saccharomyces cerevisiae, a common conditional phenotype associated with deletion or mutation of genes encoding mRNA export factors is the rapid accumulation of mRNAs in intranuclear foci, suggested to be near transcription sites. The nuclear RNA exosome has been implicated in retaining RNAs in these foci; on deletion of the exosome component Rrp6p, the RNA is released. To determine the exact nuclear location of retained as well as released mRNAs, we have used mRNA export mutant strains to analyze the spatial relationship between newly synthesized heat shock mRNA, the chromosomal site of transcription, and known S. cerevisiae nuclear structures such as the nucleolus and the nucleolar body. Our results show that retained SSA4 RNA localizes to an area in close proximity to the SSA4 locus. On deletion of Rrp6p and release from the genomic locus, heat shock mRNAs produced in the rat7–1 strain colocalize predominantly with nucleolar antigens. Bulk poly(A)+ RNA, on the other hand, is localized primarily to the nuclear rim. Interestingly, the RNA binding nucleocytoplasmic shuttle protein Npl3p shows strong colocalization with bulk poly(A)+ RNA, regardless of its nuclear location. Taken together, our data show that retention occurs close to the gene and indicate distinct nuclear fates of different mRNAs.
Novel “anti-reverse” cap analogs with superior translational properties
Authors: JEMIELITY JACEK , FOWLER TOLVERT , ZUBEREK JOANNA , STEPINSKI JANUSZ , LEWDOROWICZ MAGDALENA , NIEDZWIECKA ANNA , STOLARSKI RYSZARD , DARZYNKIEWICZ EDWARD , RHOADS ROBERT E.
Published: August 2003

Synthetic analogs of the 5′-terminal caps of eukaryotic mRNAs and snRNAs are used in elucidating such physiological processes as mRNA translation, pre-mRNA splicing, intracellular transport of mRNA and snRNAs, and mRNA turnover. Particularly useful are RNAs capped with synthetic analogs, which are produced by in vitro transcription of a DNA template using a bacteriophage RNA polymerase in the presence of ribonucleoside triphosphates and a cap dinucleotide such as m7Gp3G. Unfortunately, because of the presence of a 3′-OH on both the m7Guo and Guo moieties, up to half of the mRNAs contain caps incorporated in the reverse orientation. Previously we designed and synthesized two “anti-reverse” cap analogs (ARCAs), m73′dGp3G and m27,3′-OGp3G, that cannot be incorporated in the reverse orientation because of modifications at the C3′ position of m7Guo. In the present study, we have synthesized seven new cap analogs modified in the C2′ and C3′ positions of m7Guo and in the number of phosphate residues, m27,2′-OGp3G, m72′dGp3G, m72′dGp4G, m27,2′-OGp4G, m27,3′-OGp4G, m7Gp5G, and m27,3′-OGp5G. These were analyzed for conformation in solution, binding affinity to eIF4E, inhibition of in vitro translation, degree of reverse capping during in vitro transcription, capping efficiency, and the ability to stimulate cap-dependent translation in vitro when incorporated into mRNA. The results indicate that modifications at C2′, like those at C3′, prevent reverse incorporation, that tetra- and pentaphosphate cap analogs bind eIF4E and inhibit translation more strongly than their triphosphate counterparts, and that tetraphosphate ARCAs promote cap-dependent translation more effectively than previous cap analogs.
tRNA requirements for glyQS antitermination: A new twist on tRNA
Authors: YOUSEF MARY R. , GRUNDY FRANK J. , HENKIN TINA M.
Published: August 2003

Transcription antitermination of the Bacillus subtilis glyQS gene, a member of the T box gene regulation family, can be induced during in vitro transcription in a minimal system using purified B. subtilis RNA polymerase by the addition of unmodified T7 RNA polymerase-transcribed tRNAGly. Antitermination was previously shown to depend on base-pairing between the glyQS leader and the tRNA at the anticodon and acceptor ends. In this study, variants of tRNAGly were generated to identify additional tRNA elements required for antitermination activity, and to determine the effect of structural changes in the tRNA. We find that additions to the 3′ end of the tRNA blocked antitermination, in agreement with the prediction that uncharged tRNA is the effector in vivo, whereas insertion of 1 nucleotide between the acceptor stem and the 3′ UCCA residues had no effect. Disruption of the D-loop/T-loop tertiary interaction inhibited antitermination function, as was previously demonstrated for tRNATyr-directed antitermination of the B. subtilis tyrS gene in vivo. Insertion of a single base pair in the anticodon stem was tolerated, whereas further insertions abolished antitermination. However, we find that major alterations in the length of the acceptor stem are tolerated, and the insertions exhibited a pattern of periodicity suggesting that there is face-of-the-helix dependence in the positioning of the unpaired UCCA residues at the 3′ end of the tRNA for interaction with the antiterminator bulge and antitermination.
Inhibition of HIV-1 reverse transcriptase by RNA aptamers in Escherichia coli
Authors: NICKENS DAVID G. , PATTERSON JAMES T. , BURKE DONALD H.
Published: August 2003

A better understanding of aptamer function in bacteria would help to establish simple model systems for screening RNA–protein interactions within an intracellular context. Escherichia coli DNA polymerase I mutants (Pol Its) fail to grow at 37°C unless an exogenous DNA polymerase such as HIV-1 reverse transcriptase (RT) is expressed within the cell. Here, we show that four RNA aptamers that inhibit HIV-1 RT in vitro block complementation by HIV-1 RT when expressed in vivo. No other essential functions are impaired by aptamer expression at either temperature. Intracellular aptamer RNA concentrations from induced cultures were measured to range from 76 to 180 nM, which is comparable with exogenously expressed HIV-1 RT levels in these cells. RT polymerase activity was reduced to background levels in cell-free extracts prepared from cultures expressing both HIV-1 RT and the 70.28 aptamer, compared with extracts from cultures expressing HIV-1 RT alone. Intracellularly expressed RNA aptamers can thus be used to generate conditional null mutants in bacteria by titrating an essential protein.
Contribution of domain structure to the RNA 3′ end processing and degradation functions of the nuclear exosome subunit Rrp6p
Authors: PHILLIPS SEASSON , BUTLER J. SCOTT
Published: August 2003

The 3′–5′ riboexonuclease Rrp6p, a nuclear component of the exosome, functions with other exosome components to produce the mature 3′ ends of 5.8S rRNA, sno- and snRNAs, and to destroy improperly processed precursor (pre)-rRNAs and pre-mRNAs. Rrp6p is a member of the RNase D family of riboexonucleases and displays a high degree of homology with the active site of the deoxyriboexonuclease domain of Escherichia coli DNA polymerase I, the crystal structure of which indicates a two-metal ion mechanism for phosphodiester bond hydrolysis. Mutation of each of the conserved residues predicted to coordinate metal ions in the active site of Rrp6p abolished activity of the enzyme in vitro and in vivo. Complete loss of Rrp6p activity caused by the Y361F and Y361A mutations supports the critical role proposed for the phenolic hydroxyl of Tyr361 in the reaction mechanism. Rrp6p also contains an helicase RNase D C-terminal (HRDC) domain of unknown function that is similar to domains in the Werner’s and Bloom’s Syndrome proteins. A point mutation in this domain results in Rrp6p that localizes to the nucleus, but fails to efficiently process the 3′ ends of 5.8S pre-rRNA and some pre-snoRNAs. In contrast, this mutant retains the ability to degrade rRNA processing intermediates and 3′-extended, poly(A)+ snoRNAs. These findings indicate the potential for independent control of the processing and degradation functions of Rrp6p.
Partitioning and translation of mRNAs encoding soluble proteins on membrane-bound ribosomes
Authors: LERNER RACHEL S. , SEISER ROBERT M. , ZHENG TIANLI , LAGER PATRICK J. , REEDY MARY C. , KEENE JACK D. , NICCHITTA CHRISTOPHER V.
Published: August 2003

In eukaryotic cells, it is generally accepted that protein synthesis is compartmentalized; soluble proteins are synthesized on free ribosomes, whereas secretory and membrane proteins are synthesized on endoplasmic reticulum (ER)-bound ribosomes. The partitioning of mRNAs that accompanies such compartmentalization arises early in protein synthesis, when ribosomes engaged in the translation of mRNAs encoding signal-sequence-bearing proteins are targeted to the ER. In this report, we use multiple cell fractionation protocols, in combination with cDNA microarray, nuclease protection, and Northern blot analyses, to assess the distribution of mRNAs between free and ER-bound ribosomes. We find a broad representation of mRNAs encoding soluble proteins in the ER fraction, with a subset of such mRNAs displaying substantial ER partitioning. In addition, we present evidence that membrane-bound ribosomes engage in the translation of mRNAs encoding soluble proteins. Single-cell in situ hybridization analysis of the subcellular distribution of mRNAs encoding ER-localized and soluble proteins identify two overall patterns of mRNA distribution in the cell—endoplasmic reticular and cytosolic. However, both partitioning patterns include a distinct perinuclear component. These results identify previously unappreciated roles for membrane-bound ribosomes in the subcellular compartmentalization of protein synthesis and indicate possible functions for the perinuclear membrane domain in mRNA sorting in the cell.