bluuebunny/crossref_metadata_2025 huggingface dataset:
Induction of heat shock protein (HSP)‐70 in posterior cingulate and retrosplenial cortex of rat brain by dizocilpine and phencyclidine: lack of protective effects of s receptor ligands
Authors: HASHIMOTO KENJI , TOMITAKA SHIN‐ICHIRO , NARITA NATSUKO , MINABE YOSHIO , IYO MASAOMI , FUKUI SUSUMU
Published: August 2003

The role of σ receptors in the induction of heat shock protein (HSP)‐70 by non‐competitive N‐methyl‐Daspartate (NMDA) receptor antagonists (+)‐MK‐801 (dizocilpine) and phencyclidine (PCP) was studied. HSP‐70 is induced in the posterior cingulate and retrosplenial cortex of rat brain 24 hours after a single administration of dizocilpine (1 mg/kg) or PCP (50 mg/kg). The induction of heat shock protein HSP‐70 by dizocilpine or PCP was attenuated partially by pre‐treatment with the antipsychotic drug haloperidol (3 mg/kg, i.p., 15 minutes previously). However, pre‐treatment with high potent and selective σ receptor ligands, 4‐phenyl‐4‐(1‐phenylbutyl)piperidine (4‐PPBP, 3 mg/kg, i.p., 15 minutes previously) and N,N‐dipropyl‐2‐[4‐methoxy‐3‐(2‐phenylethoxy)phenyl]‐ethylamine monohydrochloride) (NE‐100, 3 mg/kg, i.p., 15 minutes previously) did not alter the induction of HSP‐70 by dizocilpine or PCP. These findings suggest that σ receptors may not play a significant role in the induction of HSP‐70 by non‐competitive NMDA receptor antagonists dizocilpine and PCP, and that protective effects of haloperidol on induction of HSP‐70 protein by dizocilpine or PCP may be due to other effect(s) except σ receptors.
The PUF Family of RNA‐binding Proteins: Does Evolutionarily Conserved Structure Equal Conserved Function?
Authors: Spassov Danislav , Jurecic Roland
Published: August 2003

Drosophila Pumilio ( Pum ) protein is a founder member of a novel family of RNA‐binding proteins, known as the PUF family. The PUF proteins constitute an evolutionarily highly conserved family of proteins present from yeast to humans and plants, and are characterized by a highly conserved C‐terminal RNA‐binding domain, composed of eight tandem repeats. The conserved biochemical features and genetic function of PUF family members have emerged from studies of model organisms. PUF proteins bind to related sequence motifs in the 3' untranslated region (3'UTR) of specific target mRNAs and repress their translation. Frequently, PUF proteins function asymmetrically to create protein gradients, thus causing asymmetric cell division and regulating cell fate specification. Thus, it was recently proposed that the primordial role of PUF proteins is to sustain mitotic proliferation of stem cells. Here we review the evolution, conserved genetic and biochemical properties of PUF family of proteins, and discuss protein interactions, upstream regulators and downstream targets of PUF proteins. We also suggest that a conserved mechanism of PUF function extends to the newly described mammalian members of the PUF family (human PUM1 and PUM2, and mouse Pum1 and Pum2 ), that show extensive homology to Drosophila Pum, and could have an important role in cell development, fate specification and differentiation. IUBMB Life, 55: 359‐366, 2003
U4 snRNA nucleolar localization requires the NHPX/15.5-kD protein binding site but not Sm protein or U6 snRNA association
Authors: Gerbi Susan A. , Borovjagin Anton V. , Odreman Federico E. , Lange Thilo Sascha
Published: August 2003

All small nuclear RNAs (snRNAs) of the [U4/U6.U5] tri-snRNP localize transiently to nucleoli, as visualized by microscopy after injection of fluorescein-labeled transcripts into Xenopus laevis oocyte nuclei. Here, we demonstrate that these RNAs traffic to nucleoli independently of one another, because U4 snRNA deleted in the U6 base-pairing region still localizes to nucleoli. Furthermore, depletion of endogenous U6 snRNA does not affect nucleolar localization of injected U4 or U5. The wild-type U4 transcripts used here are functional: they exhibit normal nucleocytoplasmic traffic, associate with Sm proteins, form the [U4/U6] di-snRNP, and localize to nucleoli and Cajal bodies. The nucleolar localization element (NoLE) of U4 snRNA was mapped by mutagenesis. Neither the 5′-cap nor the 3′-region of U4, which includes the Sm protein binding site, are essential for nucleolar localization. The only region in U4 snRNA required for nucleolar localization is the 5′-proximal stem loop, which contains the binding site for the NHPX/15.5-kD protein. Even mutation of just five nucleotides, essential for binding this protein, impaired U4 nucleolar localization. Intriguingly, the NHPX/15.5-kD protein also binds the nucleolar localization element of box C/D small nucleolar RNAs, suggesting that this protein might mediate nucleolar localization of several small RNAs.
The PCH family protein, Cdc15p, recruits two F-actin nucleation pathways to coordinate cytokinetic actin ring formation in Schizosaccharomyces pombe
Authors: Carnahan Robert H. , Gould Kathleen L.
Published: August 2003

Cytokinetic actin ring (CAR) formation in Schizosaccharomyces pombe requires two independent actin nucleation pathways, one dependent on the Arp2/3 complex and another involving the formin Cdc12p. Here we investigate the role of the S. pombe Cdc15 homology family protein, Cdc15p, in CAR assembly and find that it interacts with proteins from both of these nucleation pathways. Cdc15p binds directly to the Arp2/3 complex activator Myo1p, which likely explains why actin patches and the Arp2/3 complex fail to be medially recruited during mitosis in cdc15 mutants. Cdc15p also binds directly to Cdc12p. Cdc15p and Cdc12p not only display mutual dependence for CAR localization, but also exist together in a ring-nucleating structure before CAR formation. The disruption of these interactions in cdc15 null cells is likely to be the reason for their complete lack of CARs. We propose a model in which Cdc15p plays a critical role in recruiting and coordinating the pathways essential for the assembly of medially located F-actin filaments and construction of the CAR.
Interaction of Aurora-A and centrosomin at the microtubule-nucleating site in Drosophila and mammalian cells
Authors: Terada Yasuhiko , Uetake Yumi , Kuriyama Ryoko
Published: August 2003

A mitosis-specific Aurora-A kinase has been implicated in microtubule organization and spindle assembly in diverse organisms. However, exactly how Aurora-A controls the microtubule nucleation onto centrosomes is unknown. Here, we show that Aurora-A specifically binds to the COOH-terminal domain of a Drosophila centrosomal protein, centrosomin (CNN), which has been shown to be important for assembly of mitotic spindles and spindle poles. Aurora-A and CNN are mutually dependent for localization at spindle poles, which is required for proper targeting of γ-tubulin and other centrosomal components to the centrosome. The NH2-terminal half of CNN interacts with γ-tubulin, and induces cytoplasmic foci that can initiate microtubule nucleation in vivo and in vitro in both Drosophila and mammalian cells. These results suggest that Aurora-A regulates centrosome assembly by controlling the CNN's ability to targeting and/or anchoring γ-tubulin to the centrosome and organizing microtubule-nucleating sites via its interaction with the COOH-terminal sequence of CNN.
Phosphorylation of mitotic kinesin-like protein 2 by polo-like kinase 1 is required for cytokinesis
Authors: Neef Rüdiger , Preisinger Christian , Sutcliffe Josephine , Kopajtich Robert , Nigg Erich A. , Mayer Thomas U. , Barr Francis A.
Published: August 2003

We have investigated the function of mitotic kinesin-like protein (MKlp) 2, a kinesin localized to the central spindle, and demonstrate that its depletion results in a failure of cleavage furrow ingression and cytokinesis, and disrupts localization of polo-like kinase 1 (Plk1). MKlp2 is a target for Plk1, and phosphorylated MKlp2 binds to the polo box domain of Plk1. Plk1 also binds directly to microtubules and targets to the central spindle via its polo box domain, and this interaction controls the activity of Plk1 toward MKlp2. An antibody to the neck region of MKlp2 that prevents phosphorylation of MKlp2 by Plk1 causes a cytokinesis defect when introduced into cells. We propose that phosphorylation of MKlp2 by Plk1 is necessary for the spatial restriction of Plk1 to the central spindle during anaphase and telophase, and the complex of these two proteins is required for cytokinesis.
Stroke-induced Immunodeficiency Promotes Spontaneous Bacterial Infections and Is Mediated by Sympathetic Activation Reversal by Poststroke T Helper Cell Type 1–like Immunostimulation
Authors: Prass Konstantin , Meisel Christian , Höflich Conny , Braun Johann , Halle Elke , Wolf Tilo , Ruscher Karsten , Victorov Ilya V. , Priller Josef , Dirnagl Ulrich , Volk Hans-Dieter , Meisel Andreas
Published: August 2003

Infections are a leading cause of death in stroke patients. In a mouse model of focal cerebral ischemia, we tested the hypothesis that a stroke-induced immunodeficiency increases the susceptibility to bacterial infections. 3 d after ischemia, all animals developed spontaneous septicemia and pneumonia. Stroke induced an extensive apoptotic loss of lymphocytes and a shift from T helper cell (Th)1 to Th2 cytokine production. Adoptive transfer of T and natural killer cells from wild-type mice, but not from interferon (IFN)-γ–deficient mice, or administration of IFN-γ at day 1 after stroke greatly decreased the bacterial burden. Importantly, the defective IFN-γ response and the occurrence of bacterial infections were prevented by blocking the sympathetic nervous system but not the hypothalamo-pituitary-adrenal axis. Furthermore, administration of the β-adrenoreceptor blocker propranolol drastically reduced mortality after stroke. These data suggest that a catecholamine-mediated defect in early lymphocyte activation is the key factor in the impaired antibacterial immune response after stroke.
A Naturally Selected Dimorphism within the HLA-B44 Supertype Alters Class I Structure, Peptide Repertoire, and T Cell Recognition
Authors: Macdonald Whitney A. , Purcell Anthony W. , Mifsud Nicole A. , Ely Lauren K. , Williams David S. , Chang Linus , Gorman Jeffrey J. , Clements Craig S. , Kjer-Nielsen Lars , Koelle David M. , Burrows Scott R. , Tait Brian D. , Holdsworth Rhonda , Brooks Andrew G. , Lovrecz George O. , Lu Louis , Rossjohn Jamie , McCluskey James
Published: August 2003

HLA-B*4402 and B*4403 are naturally occurring MHC class I alleles that are both found at a high frequency in all human populations, and yet they only differ by one residue on the α2 helix (B*4402 Asp156→B*4403 Leu156). CTLs discriminate between HLA-B*4402 and B*4403, and these allotypes stimulate strong mutual allogeneic responses reflecting their known barrier to hemopoeitic stem cell transplantation. Although HLA-B*4402 and B*4403 share >95% of their peptide repertoire, B*4403 presents more unique peptides than B*4402, consistent with the stronger T cell alloreactivity observed toward B*4403 compared with B*4402. Crystal structures of B*4402 and B*4403 show how the polymorphism at position 156 is completely buried and yet alters both the peptide and the heavy chain conformation, relaxing ligand selection by B*4403 compared with B*4402. Thus, the polymorphism between HLA-B*4402 and B*4403 modifies both peptide repertoire and T cell recognition, and is reflected in the paradoxically powerful alloreactivity that occurs across this “minimal” mismatch. The findings suggest that these closely related class I genes are maintained in diverse human populations through their differential impact on the selection of peptide ligands and the T cell repertoire.
The IκB Function of NF-κB2 p100 Controls Stimulated Osteoclastogenesis
Authors: Novack Deborah Veis , Yin Li , Hagen-Stapleton Amanda , Schreiber Robert D. , Goeddel David V. , Ross F. Patrick , Teitelbaum Steven L.
Published: August 2003

The prototranscription factor p100 represents an intersection of the NF-κB and IκB families, potentially serving as both the precursor for the active NF-κB subunit p52 and as an IκB capable of retaining NF-κB in the cytoplasm. NF-κB–inducing kinase (NIK) controls processing of p100 to generate p52, and thus NIK-deficient mice can be used to examine the biological effects of a failure in such processing. We demonstrate that treatment of wild-type osteoclast precursors with the osteoclastogenic cytokine receptor activator of NF-κB ligand (RANKL) increases both expression of p100 and its conversion to p52, resulting in unchanged net levels of p100. In the absence of NIK, p100 expression is increased by RANKL, but its conversion to p52 is blocked, leading to cytosolic accumulation of p100, which, acting as an IκB protein, binds NF-κB complexes and prevents their nuclear translocation. High levels of unprocessed p100 in osteoclast precursors from NIK−/− mice or a nonprocessable form of the protein in wild-type cells impair RANKL-mediated osteoclastogenesis. Conversely, p100-deficient osteoclast precursors show enhanced sensitivity to RANKL. These data demonstrate a novel, biologically relevant means of regulating NF-κB signaling, with upstream control and kinetics distinct from the classical IκBα pathway.
Truncation of C-mip (Tc-mip), a New Proximal Signaling Protein, Induces c-maf Th2 Transcription Factor and Cytoskeleton Reorganization
Authors: Grimbert Philippe , Valanciute Asta , Audard Vincent , Pawlak André , Le gouvelo Sabine , Lang Philippe , Niaudet Patrick , Bensman Albert , Guellaën Georges , Sahali Djillali
Published: August 2003

Several arguments suggest that minimal change nephrotic syndrome (MCNS) results from yet unknown systemic disorder of T cell function. By screening a cDNA library from T cell relapse, we identified a new pleckstrin homology (PH) domain-containing protein encoded by a gene located on chromosome 16q24. Two alternative transcripts were identified. The first species (c-mip) was expressed in fetal liver, kidney, and peripheral blood mononuclear cells (PBMCs), but weakly detected in PBMCs from MCNS patients. The second form (Tc-mip, standing for truncated c-maf inducing protein), corresponds to subtracted transcript and lacks the NH2-terminal PH domain. The expression of Tc-mip was restricted to fetal liver, thymus, and MCNS PBMCs where it was specifically recruited in CD4+ T cells subset. Overexpression of Tc-mip in T cell Jurkat induced c-maf, transactivated the interleukin 4 gene and down-regulated the interferon γ expression, characteristic of a Th2 commitment. Moreover, the overexpression of Tc-mip induced Src phosphorylation, T cell clustering, and a cellular redistribution of the cytoskeleton-associated L-plastin, by a PI3 kinase independent pathway. Tc-mip represents therefore the first identified protein, which links proximal signaling to c-maf induction.
Continuous Activation of Autoreactive CD4+ CD25+ Regulatory T Cells in the Steady State
Authors: Fisson Sylvain , Darrasse-Jèze Guillaume , Litvinova Elena , Septier Franck , Klatzmann David , Liblau Roland , Salomon Benoît L.
Published: August 2003

Despite a growing interest in CD4+ CD25+ regulatory T cells (Treg) that play a major role in self-tolerance and immunoregulation, fundamental parameters of the biology and homeostasis of these cells are poorly known. Here, we show that this population is composed of two Treg subsets that have distinct phenotypes and homeostasis in normal unmanipulated mice. In the steady state, some Treg remain quiescent and have a long lifespan, in the order of months, whereas the other Treg are dividing extensively and express multiple activation markers. After adoptive transfer, tissue-specific Treg rapidly divide and expand preferentially in lymph nodes draining their target self-antigens. These results reveal the existence of a cycling Treg subset composed of autoreactive Treg that are continuously activated by tissue self-antigens.
Coordinated Adenine Nucleotide Phosphohydrolysis and Nucleoside Signaling in Posthypoxic Endothelium
Authors: Eltzschig Holger K. , Ibla Juan C. , Furuta Glenn T. , Leonard Martin O. , Jacobson Kenneth A. , Enjyoji Keiichi , Robson Simon C. , Colgan Sean P.
Published: August 2003

Limited oxygen delivery to tissues (hypoxia) is common in a variety of disease states. A number of parallels exist between hypoxia and acute inflammation, including the observation that both influence vascular permeability. As such, we compared the functional influence of activated polymorphonuclear leukocytes (PMN) on normoxic and posthypoxic endothelial cells. Initial studies indicated that activated PMN preferentially promote endothelial barrier function in posthypoxic endothelial cells (>60% increase over normoxia). Extension of these findings identified at least one soluble mediator as extracellular adenosine triphosphate (ATP). Subsequent studies revealed that ATP is coordinately hydrolyzed to adenosine at the endothelial cell surface by hypoxia-induced CD39 and CD73 (>20-and >12-fold increase in mRNA, respectively). Studies in vitro and in cd39-null mice identified these surface ecto-enzymes as critical control points for posthypoxia-associated protection of vascular permeability. Furthermore, insight gained through microarray analysis revealed that the adenosine A2B receptor (AdoRA2B) is selectively up-regulated by hypoxia (>5-fold increase in mRNA), and that AdoRA2B antagonists effectively neutralize ATP-mediated changes in posthypoxic endothelial permeability. Taken together, these results demonstrate transcription coordination of adenine nucleotide and nucleoside signaling at the vascular interface during hypoxia.
Matrix Metalloproteinase 19 Regulates Insulin-like Growth Factor-mediated Proliferation, Migration, and Adhesion in Human Keratinocytes through Proteolysis of Insulin-like Growth Factor Binding Protein-3
Authors: Sadowski Thorsten , Dietrich Sebastian , Koschinsky Felix , Sedlacek Radislav
Published: August 2003

Unlike most other matrix metalloproteinases (MMPs) MMP-19 is expressed in undifferentiated basal keratinocytes of healthy human skin. The human keratinocyte cell line HaCaT, which like basal keratinocytes constitutively expresses MMP-19, down-regulated the expression of MMP-19 at high calcium concentrations. Calcium-regulation occurred through E-cadherin mediated cell-cell contacts because neutralizing anti-E-cadherin antibodies restored MMP-19 expression in high calcium. Overexpression of MMP-19 in HaCaT cells (HaCaT-WT) increased cellular proliferation, as well as migration and adhesion on type I collagen. This was due to proteolysis of the insulin-like growth factor (IGF) binding protein-3 by MMP-19, which augmented signaling through the IGF-I receptor, as evidenced by its increased autophosphorylation. Conversely, these effects were not observed in cells transfected with MMP-2 or a catalytically inactive MMP-19 mutant. As further proof that increased IGF-signaling promoted adhesion and migration in HaCaT-WT cells, we reproduced these effects by treating parental HaCaT with IGF-I. We observed dephosphorylation of the focal adhesion kinase in HaCaT-WT as well as IGF-I–treated HaCaT cells, suggesting that inactivating focal adhesion kinase is a mechanism by which IGF-I enhances adhesion. Furthermore, IGF-I-triggered motility on type I collagen was mediated by MMP activity, which, however, was distinct from MMP-19. Considering the coexpression of IGFBP-3 and MMP-19 in the skin, we conclude that MMP-19 is a likely candidate to be the major IGFBP-3 degrading MMP in the quiescent epidermis. This activity might have widespread consequences for the behavior of epidermal keratinocytes.
Mutations in a β-Tubulin Disrupt Spindle Orientation and Microtubule Dynamics in the EarlyCaenorhabditis elegansEmbryo
Authors: Wright Amanda J. , Hunter Craig P.
Published: August 2003

The early Caenorhabditis elegans embryo contains abundant transcripts for two α- and two β-tubulins, raising the question of whether each isoform performs specialized functions or simply contributes to total tubulin levels. Our identification of two recessive, complementing alleles of a β-tubulin that disrupt nuclear-centrosome centration and rotation in the early embryo originally suggested that this tubulin, tbb-2, has specialized functions. However, embryos from tbb-2 deletion worms do not have defects in nuclear-centrosome centration and rotation suggesting that the complementing alleles are not null mutations. Both complementing alleles have distinct effects on microtubule dynamics and show allele-specific interactions with the two embryonically expressed α-tubulins: One of the alleles causes microtubules to be cold stable and resistant to the microtubule-depolymerizing drug benomyl, whereas the other causes cell cycle-specific defects in microtubule polymerization. Gene-specific RNA interference targeting all four embryonically expressed tubulin genes singly and in all double combinations showed that the tubulin isoforms in the early embryo are largely functionally redundant with the exception of tbb-2. tbb-2 is required for centrosome stabilization during anaphase of the first cell division, suggesting that tbb-2 may be specialized for interactions with the cell cortex.
Mechanism of Constitutive Export from the Golgi: Bulk Flow via the Formation, Protrusion, and En Bloc Cleavage of largetrans-Golgi Network Tubular Domains
Authors: Polishchuk Elena V. , Di Pentima Alessio , Luini Alberto , Polishchuk Roman S.
Published: August 2003

Transport of constitutive cargo proteins from the Golgi complex to the plasma membrane (PM) is known to be mediated by large tubular-saccular carriers moving along microtubules. However, the process by which these large structures emerge from the trans-Golgi network (TGN) remains unclear. Here, we address the question of the formation of Golgi-to-PM carriers (GPCs) by using a suitable cluster of morphological techniques, providing an integrated view of their dynamics and three-dimensional structure. Our results indicate that exit from the TGN of a constitutive traffic marker, the VSVG protein, occurs by bulk flow and is a three-step process. First, the formation of a tubular-reticular TGN domain (GPC precursor) that includes PM-directed proteins and excludes other cargo and Golgi-resident proteins. Notably, this step does not require membrane fusion. Second, the docking of this preformed domain on microtubules and its kinesin-mediated extrusion. Finally, the detachment of the extruded domain by membrane fission. The formation of GPCs does not involve cargo concentration and is not associated with the presence of known coat proteins on GPC precursors. In summary, export from the Golgi occurs via the formation, protrusion and en bloc cleavage of specialized TGN tubular-saccular domains.
Inhibition of Proteasomal Activity Causes Inclusion Formation in Neuronal and Non-Neuronal Cells Overexpressing Parkin
Authors: Ardley Helen C. , Scott Gina B. , Rose Stephen A. , Tan Nancy G. S. , Markham Alexander F. , Robinson Philip A.
Published: August 2003

Association between protein inclusions and neurodegenerative diseases, including Parkinson's and Alzheimer's diseases, and polyglutamine disorders, has been widely documented. Although ubiquitin is conjugated to many of these aggregated proteins, the 26S proteasome does not efficiently degrade them. Mutations in the ubiquitin-protein ligase Parkin are associated with autosomal recessive juvenile Parkinsonism. Although Parkin-positive inclusions are not detected in brains of autosomal recessive juvenile Parkinsonism patients, Parkin is found in Lewy bodies in sporadic disease. This suggests that loss of Parkin ligase activity via mutation, or sequestration to Lewy bodies, is a contributory factor to sporadic disease onset. We now demonstrate that decreased proteasomal activity causes formation of large, noncytotoxic inclusions within the cytoplasm of both neuronal and nonneuronal cells overexpressing Parkin. This is not a general phenomenon as there is an absence of similar inclusions when HHARI, a structural homolog of Parkin, is overexpressed. The inclusions colocalize with ubiquitin and with proteasomes. Furthermore, Parkin inclusions colocalize with γ-tubulin, acetylated α-tubulin, and cause redistribution of vimentin, suggesting aggresome-like properties. Our data imply that lower proteasomal activity, previously observed in brain tissue of Parkinson's disease patients, leads to Parkin accumulation and a concomitant reduction in ligase activity, thereby promoting Lewy body formation.
Villin Enhances Hepatocyte Growth Factor-induced Actin Cytoskeleton Remodeling in Epithelial Cells
Authors: Athman Rafika , Louvard Daniel , Robine Sylvie
Published: August 2003

Villin is an actin-binding protein localized to intestinal and kidney brush borders. In vitro, villin has been demonstrated to bundle and sever F-actin in a calcium-dependent manner. Although villin is not necessary for the bundling of F-actin in vivo, it is important for the reorganization of the actin cytoskeleton elicited by stress during both physiological and pathological conditions ( Ferrary et al., 1999 ). These data suggest that villin may be involved in actin cytoskeleton remodeling necessary for many processes requiring cellular plasticity. Here, we study the role of villin in hepatocyte growth factor (HGF)-induced epithelial cell motility and morphogenesis. For this purpose, we used primary cultures of enterocytes derived from wild-type and villin knock-out mice and Madin-Darby canine kidney cells, expressing villin in an inducible manner. In vitro, we show that epithelial cell lysates from villin-expressing cells induced dramatic, calcium-dependent severing of actin filaments. In cell culture, we found that villin-expressing cells exhibit enhanced cell motility and morphogenesis upon HGF stimulation. In addition, we show that the ability of villin to potentiate HGF-induced actin reorganization occurs through the HGF-activated phospholipase Cγ signaling pathway. Collectively, these data demonstrate that villin acts as a regulator of HGF-induced actin dynamics.
DNA Damage Modulates Nucleolar Interaction of the Werner Protein with the AAA ATPase p97/VCP
Authors: Partridge Juneth Joaquin , Lopreiato Joseph Onofrio , Latterich Martin , Indig Fred Eliezer
Published: August 2003

We report a novel nucleolar interaction between the AAA ATPase p97/VCP and the Werner protein (WRNp), a member of the RecQ helicase family. p97/VCP mediates several important cellular functions in eucaryotic cells, including membrane fusion of the endoplasmic reticulum and Golgi and ubiquitin-dependent protein degradation. Mutations in the WRN gene cause Werner syndrome, a genetic disorder characterized by premature onset of aging symptoms, a higher incidence of cancer, and a high susceptibility to DNA damage caused by topoisomerase inhibitors. We observed that both WRNp and valosin-containing protein (VCP) were present in the nucleoplasm and in nucleolar foci in mammalian cells and that WRNp and p97/VCP physically interacted in the nucleoli. Importantly, the nucleolar WRNp/VCP complex was dissociated by treatment with camptothecin, an inhibitor of topoisomerase I, whereas other WRNp-associated protein complexes, such as WRNp/Ku 80, were not dissociated by this drug. Because WRN syndrome cells are sensitive to topoisomerase inhibitors, these observations suggest that the VCP/WRNp interaction plays an important role in WRN biology. We propose a novel role for VCP in the DNA damage response pathway through modulation of WRNp availability.
Maximal Polar Growth Potential Depends on the Polarisome Component AgSpa2 in the Filamentous FungusAshbya gossypii
Authors: Knechtle Philipp , Dietrich Fred , Philippsen Peter
Published: August 2003

We used actin staining and videomicroscopy to analyze the development from a spore to a young mycelium in the filamentous ascomycete Ashbya gossypii. The development starts with an initial isotropic growth phase followed by the emergence of germ tubes. The initial tip growth speed of 6–10 μm/h increases during early stages of development. This increase is transiently interrupted in response to the establishment of lateral branches or septa. The hyphal tip growth speed finally reaches a maximum of up to 200 μm/h, and the tips of these mature hyphae have the ability to split into two equally fast-growing hyphae. A search for A. gossypii homologs of polarisome components of the yeast Saccharomyces cerevisiae revealed a remarkable size difference between Spa2p of both organisms, with AgSpa2p being double as long as ScSpa2p due to an extended internal domain. AgSpa2 colocalizes with sites of polarized actin. Using time-lapse videomicroscopy, we show that AgSpa2p-GFP polarization is established at sites of branch initiation and then permanently maintained at hyphal tips. Polarization at sites of septation is transient. During apical branching the existing AgSpa2p-GFP polarization is symmetrically divided. To investigate the function of AgSpa2p, we generated two AgSPA2 mutants, a partial deletion of the internal domain alone, and a complete deletion. The mutations had an impact on the maximal hyphal tip growth speed, on the hyphal diameter, and on the branching pattern. We suggest that AgSpa2p is required for the determination of the area of growth at the hyphal tip and that the extended internal domain plays an important role in this process.
Caenorhabditis elegansNucleoporins Nup93 and Nup205 Determine the Limit of Nuclear Pore Complex Size Exclusion In Vivo
Authors: Galy Vincent , Mattaj Iain W. , Askjaer Peter
Published: August 2003

Nuclear pore complexes (NPCs) span the nuclear envelope and mediate communication between the nucleus and the cytoplasm. To obtain insight into the structure and function of NPCs of multicellular organisms, we have initiated an extensive analysis of Caenorhabditis elegans nucleoporins. Of 20 assigned C. elegans nucleoporin genes, 17 were found to be essential for embryonic development either alone or in combination. In several cases, depletion of nucleoporins by RNAi caused severe defects in nuclear appearance. More specifically, the C. elegans homologs of vertebrate Nup93 and Nup205 were each found to be required for normal NPC distribution in the nuclear envelope in vivo. Depletion of Nup93 or Nup205 caused a failure in nuclear exclusion of nonnuclear macromolecules of ∼70 kDa without preventing active nuclear protein import or the assembly of the nuclear envelope. The defects in NPC exclusion were accompanied by abnormal chromatin condensation and early embryonic arrest. Thus, the contribution to NPC structure of Nup93 and Nup205 is essential for establishment of normal NPC function and for cell viability.
Inactivation of Mitotic Kinase Triggers Translocation of MEN Components to Mother-Daughter Neck in Yeast
Authors: Hwa Lim Hong , Yeong Foong May , Surana Uttam
Published: August 2003

Chromosome segregation, mitotic exit, and cytokinesis are executed in this order during mitosis. Although a scheme coordinating sister chromatid separation and initiation of mitotic exit has been proposed, the mechanism that temporally links the onset of cytokinesis to mitotic exit is not known. Exit from mitosis is regulated by the mitotic exit network (MEN), which includes a GTPase (Tem1) and various kinases (Cdc15, Cdc5, Dbf2, and Dbf20). Here, we show that Dbf2 and Dbf20 functions are necessary for the execution of cytokinesis. Relocalization of these proteins from spindle pole bodies to mother daughter neck seems to be necessary for this role because cdc15-2 mutant cells, though capable of exiting mitosis at semipermissive temperature, are unable to localize Dbf2 (and Dbf20) to the “neck” and fail to undergo cytokinesis. These cells can assemble and constrict the actomyosin ring normally but are incapable of forming a septum, suggesting that MEN components are critical for the initiation of septum formation. Interestingly, the spindle pole body to neck translocation of Dbf2 and Dbf20 is triggered by the inactivation of mitotic kinase. The requirement of kinase inactivation for translocation of MEN components to the division site thus provides a mechanism that renders mitotic exit a prerequisite for cytokinesis.
KLP-18, a Klp2 Kinesin, Is Required for Assembly of Acentrosomal Meiotic Spindles inCaenorhabditis elegans
Authors: Segbert Christoph , Barkus Rosemarie , Powers Jim , Strome Susan , Saxton William M. , Bossinger Olaf
Published: August 2003

The proper segregation of chromosomes during meiosis or mitosis requires the assembly of well organized spindles. In many organisms, meiotic spindles lack centrosomes. The formation of such acentrosomal spindles seems to involve first assembly or capture of microtubules (MTs) in a random pattern around the meiotic chromosomes and then parallel bundling and bipolar organization by the action of MT motors and other proteins. Here, we describe the structure, distribution, and function of KLP-18, a Caenorhabditis elegans Klp2 kinesin. Previous reports of Klp2 kinesins agree that it concentrates in spindles, but do not provide a clear view of its function. During prometaphase, metaphase, and anaphase, KLP-18 concentrates toward the poles in both meiotic and mitotic spindles. Depletion of KLP-18 by RNA-mediated interference prevents parallel bundling/bipolar organization of the MTs that accumulate around female meiotic chromosomes. Hence, meiotic chromosome segregation fails, leading to haploid or aneuploid embryos. Subsequent assembly and function of centrosomal mitotic spindles is normal except when aberrant maternal chromatin is present. This suggests that although KLP-18 is critical for organizing chromosome-derived MTs into a parallel bipolar spindle, the order inherent in centrosome-derived astral MT arrays greatly reduces or eliminates the need for KLP-18 organizing activity in mitotic spindles.
Impairing Actin Filament or Syndapin Functions Promotes Accumulation of Clathrin-coated Vesicles at the Apical Plasma Membrane of Acinar Epithelial Cells
Authors: da Costa Silvia R. , Sou Eunbyul , Xie Jiansong , Yarber Francie A. , Okamoto Curtis T. , Pidgeon Michael , Kessels Michael M. , Mircheff Austin K. , Schechter Joel E. , Qualmann Britta , Hamm-Alvarez Sarah F.
Published: August 2003

In this article, we investigate the contributions of actin filaments and accessory proteins to apical clathrin-mediated endocytosis in primary rabbit lacrimal acini. Confocal fluorescence and electron microscopy revealed that cytochalasin D promoted apical accumulation of clathrin, α-adaptin, dynamin, and F-actin and increased the amounts of coated pits and vesicles at the apical plasma membrane. Sorbitol density gradient analysis of membrane compartments showed that cytochalasin D increased [14C]dextran association with apical membranes from stimulated acini, consistent with functional inhibition of apical endocytosis. Recombinant syndapin SH3 domains interacted with lacrimal acinar dynamin, neuronal Wiskott-Aldrich Syndrome protein (N-WASP), and synaptojanin; their introduction by electroporation elicited remarkable accumulation of clathrin, accessory proteins, and coated pits at the apical plasma membrane. These SH3 domains also significantly (p ≤ 0.05) increased F-actin, with substantial colocalization of dynamin and N-WASP with the additional filaments. Coelectroporation with the VCA domain of N-WASP blocked the increase in F-actin and reversed the morphological changes indicative of impaired apical endocytosis. We suggest that transient modulation of actin polymerization by syndapins through activation of the Arp2/3 complex via N-WASP coordinates dynamin-mediated vesicle fission at the apical plasma membrane of acinar epithelia. Trapping of assembled F-actin intermediates during this process by cytochalasin D or syndapin SH3 domains impairs endocytosis.
Unintentional Drownings among New York State Residents, 1988–1994
Authors: Browne Marilyn L. , Lewis-Michl Elizabeth L. , Stark Alice D.
Published: August 2003

Objective.

This study examines situations in which drownings occur (environmental risk factors) and the victims' personal risk factors (age, gender, use of personal flotation device, medical condition, alcohol or drug use) to provide guidance for future drowning prevention efforts.

Methods.

The authors investigated 883 non-bathtub drownings among New York State residents for the years 1988 to 1994 using medical examiner, coroner, police, and/or hospital records in addition to death certificate data.

Results.

Males, children ages 0–4 years, and African American males ages 5–14 years residing in New York State outside New York City experienced the highest rates of drowning. The majority of drownings occurred in a natural body of water for all age groups, with the exception of children ages 0–4 years. Most drownings among children ages 0–4 years occurred in residential swimming pools. The child usually gained access to the pool via inadequate fencing, an open or ineffective gate, or a ladder (to an above-ground pool) left in the “down” position. Less than 10% of victims of watercraft-related drownings were wearing personal flotation devices. Blood alcohol concentration (BAC) tests were positive for 44% of 250 persons 15 years of age and older for whom valid toxicology results were provided; 30% had BACs of 100 mg/dl or more.

Conclusions.

Suggested prevention efforts include stricter enforcement of fencing requirements for residential swimming pools and drowning prevention education stressing personal flotation device use while boating and the danger of mixing alcohol and water-related activities.
Watercraft-Related Drownings among New York State Residents, 1988–1994
Authors: Browne Marilyn L. , Lewis-Michl Elizabeth L. , Stark Alice D.
Published: August 2003

Objective.

To describe environmental and personal risk factors associated with watercraft-related drownings. This information may be useful in developing boating safety programs, regulations, and enforcement priorities.

Methods.

A companion article in this issue (Unintentional Drownings Among New York State Residents, page 448) summarizes an investigation of 883 non-bathtub drownings among New York State residents for the years 1988 to 1994 using medical examiner, coroner, police, and hospital records in addition to death certificate data. This report details the environmental and personal risk factors associated with 216 watercraft-related drownings.

Results.

Ninety-three percent of watercraft-related drowning victims were male, with the highest rate of drowning observed among males ages 15–44 years. Most commonly, the victim entered the water when the watercraft capsized (36%), the victim fell overboard (24%), or the victim intended to swim (11%). Personal flotation devices (PFDs) were known to be worn by only 9% of drowning victims, and in these cases other risks overwhelmed the effectiveness of the PFD. Of 73 individuals 15 years of age or older for whom adequate blood alcohol concentration analyses were provided, 44% were positive for blood alcohol.

Conclusions.

Based on this study, increased use of PFDs, avoidance of dangerous currents, and less alcohol use by operators and passengers of all types of watercraft would result in a reduction in watercraft-related drownings. In addition to continued education efforts, boating safety measures that deserve consideration include enforcement of current PFD and boating while intoxicated (BWI) regulations and expansion of BWI laws to apply to all boaters.
Investigation of the chemical and electrochemical behaviour of mass transfer at an archaeological bronze/soil interface
Authors: Chunchun Xu , Julin Wang
Published: August 2003

This paper describes a study on the corrosion behaviour of archaeological bronze in simulated groundwater by means of a simulated occluded cell. The study revealed migration, enrichment and deposition patterns of ions in the environment and the cations produced by anodic dissolution during localised corrosion of bronze. The cycle voltametry technique was utilised for the first time on the study of electrochemical behaviour of the bronze/simulated occluded solution interface. The potential and current peaks of oxidation‐reduction at the cathode and anode in the occluded solutions at various time intervals facilitated a study of the corrosion products formation processes and their formation speeds.
Electrochemical evaluation of defect growth in primers applied on aluminium alloy 2024T3
Authors: Wrubl C. , Fassin M. , Buffoli A. , Mollica A.
Published: August 2003

In the present work, the protective properties of inhibitive pigments in two epoxy‐primers against corrosion of the aluminium alloy 2024T3 in marine atmosphere were investigated, the first containing SrCrO4 and the second Zn3(PO4)2. Potentiostatic polarisation and impedance measurement methods were utilised to evaluate, both the spontaneous onset of defects on coated samples and the propagation of a small artificial defect of known dimension applied since the beginning of the test on each sample, during 24 months of exposure to the marine atmosphere. These techniques enabled a quantitative evaluation of the protective efficiency of the two primers to be made, and for the effects of the surface pre‐treatments of the metallic substrate to be investigated.
Performance and mechanism of a water stabiliser for low hardness cooling water
Authors: Baiqing Zhou , Xiaowei Wang , Qin Li , Yisheng Peng
Published: August 2003

Corrosion of carbon steel and copper is a troublesome problem in low hardness cooling water systems. A new kind of water stabiliser containing hydroxy phosphonocarboxylic acid, zinc salts and molybdate has been developed. Its performance has been proved by means of weight loss tests and a static state scale‐inhibiting test method. The mechanism was also studied using polarisation tests, scanning electron microscope examination and XPS analysis. The test results showed that the corrosion rate of carbon steel and copper could be reduced to 0.0136 and 0.0010 mm/a, respectively. A compact film containing P, Mo and Zn was formed on the surface of carbon steel, by means of which the steel was protected.
Work team trust and effectiveness
Authors: Costa Ana Cristina
Published: August 2003

This article aims to explore the nature and functioning of trust in work teams. Trust is defined as a multi‐component variable with distinct but related dimensions. These include propensity to trust, perceived trustworthiness, co‐operative and lack of monitoring behaviours. A model was tested relating trust with perceived task performance, team satisfaction, and two dimensions of organisational commitment, i.e. attitudinal and continuance. Survey data from 112 teams(n=395) was collected in three social care institutions in The Netherlands. The results are supportive of a multi‐component structure for trust and of its importance to the functioning of teams and organisations. Work team trust appeared strongly related with team member's attitudes towards the organisation. Trust between team members was positively associated with attitudinal commitment and negatively with continuance commitment. Trust was also positively related with perceived task performance and with team satisfaction. In addition, perceived task performance appeared strongly related to team satisfaction.
Misplaced marketing Colas big and little: anti‐trust laws, non‐regulation and the disabled marketing of small brands
Authors: Wolburg Joyce M.
Published: August 2003

Shelf access fees and cooperative merchandising agreements are tactics that clearly give power to big business, and nowhere is this more evident than in the soft drink wars. The lack of legal protection against these tactics by US regulators can squeeze small brands out of the retail environment and effectively disable the marketing strategy of small soft drink companies and other small businesses. Thus, marketing becomes not only misplaced, but irrelevant.
Children’s visual memory of packaging
Authors: McNeal James U. , Ji Mindy F.
Published: August 2003

To elicit the visual memory of packaging that facilitates consumers’ identification and selection of products from store displays, children were asked to draw a cereal box and the results were compared with actual cereal boxes. Over 97 percent spontaneously drew a cereal box with a brand name and other brand related symbols. This may be the first time to have a glimpse of the consumer’s evoked set as it really exists. The results suggest that one’s evoked set is not just a list of brand names in the mind, but an elaborate symbolic environment made up of visual and verbal codes in which the brand name is nested. Major implications for brand and package management are discussed.
Customer‐minded growth through services
Authors: Vandermerwe Sandra
Published: August 2003

How corporations can achieve growth, especially in these uncertain economic times, is a preoccupying concern for both academics and practitioners alike today. Many of the old models and templates have fallen by the wayside, as competition has intensified, and innovators – both new entrants and renewed enterprises – have changed the rules of the game, the market, and the industry. Here, by looking at some of the interesting players, the article shows an approach to growth. The view is that using “customer minded” principles and tools to drive strategy and resources creates value and wealth through services.
Service quality: a fulfilment‐oriented and interactions‐centred approach
Authors: Dedeke Adenekan
Published: August 2003

This paper proposes a different approach to, and definition of, service quality. Service quality is framed as being dependent on composite results that a service provider and its systems offer a customer. In contrast to the approach that depicts service quality as a discrepancy construct, this paper frames the concept as a fulfilment‐oriented construct. The premise of the paper is that each services sector should have service quality criteria that specifically fit its features and characteristics. To implement a context‐dependent services quality instrument, it is argued that managers could use a service quality grid to classify firms according to their outcomes and dominant service‐encounter interactions. Three kinds of dominant interactions are introduced: customer‐to‐staff, customer‐to‐technology, and customer‐to‐product/services. Three central recommendations are proposed. First, it is important for managers to define their services in terms of the dominant service interactions. Second, managers should develop their service‐quality instruments around the dominant interactions of their particular sector. Third, managers should develop service‐quality question items using the paired criteria approach to capture customer experiences during service interactions.
Comparing the quality of private and public hospitals
Authors: Jabnoun Naceur , Chaker Mohammed
Published: August 2003

Healthcare is a highly competitive global industry. People accept to travel to remote parts of the world in order to receive the service quality they hope for. Patients usually prefer to go to private hospitals, hoping to receive high service quality. On the other hand, healthcare organizations operating in the public sector are undergoing pressure from governments and the general public to improve quality and compete effectively with their counterparts in the private sector. This paper compares the service quality rendered by private and public hospitals. A questionnaire based on SERVQUAL is developed and tested for this purpose. This questionnaire is found to have five dimensions; namely, empathy, tangibles, reliability, administrative responsiveness and supporting skills. These dimensions, as well as overall service quality, are compared between private and public hospitals. Finally the implications of the results are highlighted for healthcare managers.
Social networks as a source of competitive advantage for the firm
Authors: Van Laere Kristien , Heene Aime´
Published: August 2003

Globalisation is transforming the competitive environment of small and medium‐sized firms. Because these firms are competing with their larger counterparts in an economy where collaboration is increasingly central to organisational effectiveness, one must pay more attention to the social networks that organisations rely on. This article focuses on the relational perspective and describes the characteristics of embedded relationships that firms have to pay attention to in order to survive.