bluuebunny/crossref_metadata_2025 huggingface dataset:
Tunable wavelength-selective waveguiding of photoluminescence in Si-rich silica optical wedges
Authors: Khriachtchev Leonid , Räsänen Markku , Novikov Sergei , Lahtinen Jouko
Published: May 2004

We report accurate tuning of spectral peaks built up upon guiding of broadband light by annealed Si-rich silica layers with varying optical thickness. The optical wedges are characterized with various methods (photoluminescence, Raman spectroscopy, visible absorption, and x-ray photoelectron spectroscopy) showing that their properties change smoothly from point to point along the sample surface. As measured in the conventional transverse detection geometry, the annealed (1100 °C) Si-rich silica material exhibits well-known broad photoluminescence (PL) with maximum at ∼800 nm conventionally connected with Si nanocrystals. In the waveguiding detection geometry, the guiding of PL light by the silica layer containing Si nanocrystals is wavelength-dependent, and the detected spectrum exhibits narrow (down to 100 cm−1) and linearly polarized spectral peaks. As the central observation of the present work, these spectral peaks can be accurately tuned over the whole PL spectral region by moving the optical wedges across the laser beam. For our samples, the tuning coefficient is ∼150 cm−1/mm. It is shown that tuning of the transmission peaks is caused by change of both refractive index (from 1.6 to 1.9) and thickness (from 1.4 to 2.0 μm) of the prepared optical wedges. The propagation loss for the guided light is found to increase for shorter wavelengths and for materials with larger Si concentration. The propagation losses reliably below 0.5 cm−1 are observed in the broad spectral interval in sample areas with smaller Si concentrations.
Interaction of ultraviolet laser with a silicon carbide plume produced by laser ablation
Authors: Nam Sang Hwan , Park Seung Min
Published: May 2004

Formation mechanisms and expansion dynamics of silicon-carbon mixed cluster ions (SinCm+) in a laser-produced plasma plume produced by laser (λ=1064 nm) ablation of a SiC target in high vacuum were investigated by time-of-flight quadrupole mass spectrometry. Space- and time-resolved laser-plume interaction was examined by illuminating the plume with a time-delayed and line-focused. Ultraviolet laser pulse. Si2C was the dominant photodissociation product of silicon carbide cluster ions. Silicon-carbon mixed clusters turned out to be formed via association of atomic species in a region away from, not in front of, the target surface after a certain delay time.
Inverse ray-tracing method for nondestructive mapping of three-dimensional surfaces
Authors: Arazi L. , Kelson I. , Ducommun Y. , Kapon E.
Published: May 2004

A general method for nondestructive mapping of three-dimensional surfaces, particularly useful in cases where the surface is composed of a large array of identical depressions, is presented. The sample is first marked with a uniform flux of particles, which then act as isotropic sources of some detectable radiation. By measuring the emitted radiation from several directions, it is possible to reconstruct the surface using a parametrization coupled with a least squares fit procedure. As an example, we report on the experimental implementation of the algorithm to arrays of micron-scale inverted pyramids etched in GaAs substrates, using a particular sequence of alpha emitting atoms.
Molecular-dynamics analysis of morphological evolution of softly deposited Au nanoclusters
Authors: Shintani K. , Taniguchi Y. , Kameoka S.
Published: May 2004

The initial period following deposition of soft-landing Au clusters is investigated by classical molecular-dynamics simulation. The embedded-atom method potential is adopted for the interaction between Au atoms. Clusters of specified sizes are cut out of the bulk crystal structure. Whether a cluster equilibrated at a given temperature is in a solid state or in a liquid state is judged by tracking the trajectory of an atom in the cluster and by examining the radial distribution function. The deposition simulation reveals that there is an energy barrier in the morphological accommodation of a cluster to the substrate if the cluster is crystalline before deposition, and is equilibrated at a temperature different from that of the substrate. On the other hand, there is no energy barrier in the morphological accommodation of a cluster that is in a liquid state before deposition. Exceptionally, a crystalline cluster that is nearly at a melting temperature can accommodate itself smoothly to the substrate maintained at the same temperature without an energy barrier.
Analysis of the finite difference time domain technique to solve the Schrödinger equation for quantum devices
Authors: Soriano Antonio , Navarro Enrique A. , Portı́ Jorge A. , Such Vicente
Published: May 2004

An extension of the finite difference time domain is applied to solve the Schrödinger equation. A systematic analysis of stability and convergence of this technique is carried out in this article. The numerical scheme used to solve the Schrödinger equation differs from the scheme found in electromagnetics. Also, the unit cell employed to model quantum devices is different from the Yee cell used by the electrical engineering community. A bound for the time step is derived to ensure stability. Several numerical experiments in quantum structures demonstrate the accuracy of a second order, comparable to the analysis of electromagnetic devices with the Yee cell.
Down-regulation of Smad7 expression by ubiquitin-dependent degradation contributes to renal fibrosis in obstructive nephropathy in mice
Authors: Fukasawa Hirotaka , Yamamoto Tatsuo , Togawa Akashi , Ohashi Naro , Fujigaki Yoshihide , Oda Toshiaki , Uchida Chiharu , Kitagawa Kyoko , Hattori Takayuki , Suzuki Sayuri , Kitagawa Masatoshi , Hishida Akira
Published: June 2004

Overexpression of transforming growth factor β (TGF-β) has been shown to play pathogenic roles in progression of renal fibrosis, and the severity of tubulointerstitial fibrosis correlates better with renal function than the severity of glomerulosclerosis. Smad proteins are signaling transducers downstream from TGF-β receptors. Three families of Smad proteins have been identified: receptor-regulated Smad2 and Smad3, common partner Smad4, and inhibitory Smad7 (part of a negative-feedback loop). We investigated Smad-mediated TGF-β signaling pathway and regulatory mechanisms of inhibitory Smad7 in unilateral ureteral obstruction (UUO) kidneys in mice, a model of progressive tubulointerstitial fibrosis. Compared with sham-operated kidneys, the level of Smad7 protein, but not mRNA, decreased progressively in UUO kidneys, whereas immunoreactivity for nuclear phosphorylated Smad2 and Smad3 and renal fibrosis were inversely increased. Furthermore, we demonstrated that both the degradation and ubiquitination activity of Smad7 protein were increased markedly in UUO kidneys compared with sham-operated ones. We also found that both Smurf1 and Smurf2 (Smad ubiquitination regulatory factors), which are E3 ubiquitin ligases for Smad7, were increased and that they interacted with Smad7 in UUO kidneys. Our results suggest that the reduction of Smad7 protein resulting from enhanced ubiquitin-dependent degradation plays a pathogenic role in progression of tubulointerstitial fibrosis.
Total synthesis of (–)-spinosyn A
Authors: Mergott Dustin J. , Frank Scott A. , Roush William R.
Published: June 2004

A convergent, highly stereoselective total synthesis of (–)-spinosyn A (
1
) is described. Key features of the synthesis include the transannular Diels–Alder reaction of macrocyclic pentaene 11 and the transannular Morita–Baylis–Hillman cyclization of 12 that generates tetracycle 26. The total synthesis of (–)-spinosyn A was completed by a sequence involving the highly β-selective glycosidation reaction of 13 and glycosyl imidate 30.
Bilayers merge even when exocytosis is transient
Authors: Taraska Justin W. , Almers Wolfhard
Published: June 2004

During exocytosis, the lumen of secretory vesicles connects with the extracellular space. In some vesicles, this connection closes again, causing the vesicle to be recaptured mostly intact. The degree to which the bilayers of such vesicles mix with the plasma membrane is unknown. Work supporting the kiss-and-run model of transient exocytosis implies that synaptic vesicles allow neither lipid nor protein to escape into the plasma membrane, suggesting that the two bilayers never merge. Here, we test whether neuroendocrine granules behave similarly. Using two-color evanescent field microscopy, we imaged the lipid probe FM4-64 and fluorescent proteins in single dense core granules. During exocytosis, granules lost FM4-64 into the plasma membrane in small fractions of a second. Although FM4-64 was lost, granules retained the membrane protein, GFP-phogrin. By using GFP-phogrin as a probe for resealing, it was found that even granules that reseal lose FM4-64. We conclude that the lipid bilayers of the granule and the plasma membrane become continuous even when exocytosis is transient.
Lethal impairment of cholinergic neurotransmission in hemicholinium-3-sensitive choline transporter knockout mice
Authors: Ferguson Shawn M. , Bazalakova Mihaela , Savchenko Valentina , Tapia Juan Carlos , Wright Jane , Blakely Randy D.
Published: June 2004

Presynaptic acetylcholine (ACh) synthesis and release is thought to be sustained by a hemicholinium-3-sensitive choline transporter (CHT). We disrupted the murine CHT gene and examined CHT–/– and +/– animals for evidence of impaired cholinergic neurotransmission. Although morphologically normal at birth, CHT–/– mice become immobile, breathe irregularly, appear cyanotic, and die within an hour. Hemicholinium-3-sensitive choline uptake and subsequent ACh synthesis are specifically lost in CHT–/– mouse brains. Moreover, we observe a time-dependent loss of spontaneous and evoked responses at CHT–/– neuromuscular junctions. Consistent with deficits in synaptic ACh availability, we also observe developmental alterations in neuromuscular junction morphology reminiscent of changes in mutants lacking ACh synthesis. Adult CHT+/– mice overcome reductions in CHT protein levels and sustain choline uptake activity at wild-type levels through posttranslational mechanisms. Our results demonstrate that CHT is an essential and regulated presynaptic component of cholinergic signaling and indicate that CHT warrants consideration as a candidate gene for disorders characterized by cholinergic hypofunction.
The phosphodiesterase inhibitor rolipram delivered after a spinal cord lesion promotes axonal regeneration and functional recovery
Authors: Nikulina Elena , Tidwell J. Lille , Dai Hai Ning , Bregman Barbara S. , Filbin Marie T.
Published: June 2004

Although there is no spontaneous regeneration of mammalian spinal axons after injury, they can be enticed to grow if cAMP is elevated in the neuronal cell bodies before the spinal axons are cut. Prophylactic injection of cAMP, however, is useless as therapy for spinal injuries. We now show that the phosphodiesterase 4 (PDE4) inhibitor rolipram (which readily crosses the blood–brain barrier) overcomes inhibitors of regeneration in myelin in culture and promotes regeneration
in vivo
. Two weeks after a hemisection lesion at C3/4, with embryonic spinal tissue implanted immediately at the lesion site, a 10-day delivery of rolipram results in considerable axon regrowth into the transplant and a significant improvement in motor function. Surprisingly, in rolipram-treated animals, there was also an attenuation of reactive gliosis. Hence, because rolipram promotes axon regeneration, attenuates the formation of the glial scar, and significantly enhances functional recovery, and because it is effective when delivered s.c., as well as post-injury, it is a strong candidate as a useful therapy subsequent to spinal cord injury.
Stability and dynamics of crystals and glasses of motorized particles
Authors: Shen Tongye , Wolynes Peter G.
Published: June 2004

Many of the large structures of the cell, such as the cytoskeleton, are assembled and maintained far from equilibrium. We study the stabilities of various structures for a simple model of such a far-from-equilibrium organized assembly in which spherical particles move under the influence of attached motors. From the variational solutions of the many-body master equation for Brownian motion with motorized kicking we obtain a closed equation for the order parameter of localization. Thus, we obtain the transition criterion for localization and stability limits for the crystalline phase and frozen amorphous structures of motorized particles. The theory also allows an estimate of nonequilibrium effective temperatures characterizing the response and fluctuations of motorized asemblies.
Plasmodium falciparum
cysteine protease falcipain-1 is not essential in erythrocytic stage malaria parasites
Authors: Sijwali Puran S. , Kato Kentaro , Seydel Karl B. , Gut Jiri , Lehman Julie , Klemba Michael , Goldberg Daniel E. , Miller Louis H. , Rosenthal Philip J.
Published: May 2004

Among potential new targets for antimalarial chemotherapy are
Plasmodium falciparum
cysteine proteases, known as falcipains. Falcipain-2 and falcipain-3 are food vacuole hemoglobinases that may have additional functions. The function of falcipain-1 remains uncertain. To better characterize the role of falcipain-1 in erythrocytic parasites, we disrupted the falcipain-1 gene and characterized recombinant parasites. Disruption of the falcipain-1 gene was confirmed with Southern blots, and loss of expression of falcipain-1 was confirmed with immunoblots and by loss of labeling with a specific protease inhibitor. Compared with wild-type parasites, falcipain-1 knockout parasites developed normally, with the same morphology, multiplication rate, and invasion efficiency, and without significant differences in sensitivity to cysteine protease inhibitors. In wild-type and knockout parasites, cysteine protease inhibitors blocked hemoglobin hydrolysis in trophozoites, with a subsequent block in rupture of erythrocytes by mature schizonts, but they did not inhibit erythrocyte invasion by merozoites. Our results indicate that although falcipain-1 is expressed by erythrocytic parasites, it is not essential for normal development during this stage or for erythrocyte invasion.
In planta
functions of the
Arabidopsis
cytokinin receptor family
Authors: Higuchi Masayuki , Pischke Melissa S. , Mähönen Ari Pekka , Miyawaki Kaori , Hashimoto Yukari , Seki Motoaki , Kobayashi Masatomo , Shinozaki Kazuo , Kato Tomohiko , Tabata Satoshi , Helariutta Ykä , Sussman Michael R. , Kakimoto Tatsuo
Published: May 2004

Since their discovery as cell-division factors in plant tissue culture about five decades ago, cytokinins have been hypothesized to play a central role in the regulation of cell division and differentiation in plants. To test this hypothesis
in planta
, we isolated
Arabidopsis
plants lacking one, two, or three of the genes encoding a subfamily of histidine kinases (
CRE1
,
AHK2
, and
AHK3
) that function as cytokinin receptors. Seeds were obtained for homozygous plants containing mutations in all seven genotypes, namely single, double, and triple mutants, and the responses of germinated seedlings in various cytokinin assays were compared. Both redundant and specific functions for the three different cytokinin receptors were observed. Plants carrying mutations in all three genes did not show cytokinin responses, including inhibition of root elongation, inhibition of root formation, cell proliferation in and greening of calli, and induction of cytokinin primary-response genes. The triple mutants were small and infertile, with a reduction in meristem size and activity, yet they possessed basic organs: roots, stems, and leaves. These results confirm that cytokinins are a pivotal class of plant growth regulators but provide no evidence that cytokinins are required for the processes of gametogenesis and embryogenesis.
Activation of Aurora-A kinase by protein phosphatase inhibitor-2, a bifunctional signaling protein
Authors: Satinover David L. , Leach Craig A. , Stukenberg P. Todd , Brautigan David L.
Published: June 2004

Aurora-A kinase is necessary for centrosome maturation, for assembly and maintenance of a bipolar spindle, and for proper chromosome segregation during cell division. Aurora-A is an oncogene that is overexpressed in multiple human cancers. Regulation of kinase activity apparently depends on phosphorylation of Thr-288 in the T-loop. In addition, interactions with targeting protein for
Xenopus
kinesin-like protein 2 (TPX2) allosterically activate Aurora-A. The Thr-288 phosphorylation is reversed by type-1 protein phosphatase (PP1). Mutations in the yeast Aurora, Ipl1, are suppressed by overexpression of Glc8, the yeast homolog of phosphatase inhibitor-2 (I-2). In this study, we show that human I-2 directly and specifically stimulated recombinant human Aurora-A activity
in vitro
. The I-2 increase in kinase activity was not simply due to inhibition of PP1 because it was not mimicked by other phosphatase inhibitors. Furthermore, activation of Aurora-A was unaffected by deletion of the I-2 N-terminal PP1 binding motif but was eliminated by deletion of the I-2 C-terminal domain. Aurora-A and I-2 were recovered together from mitotic HeLa cells. Kinase activation by I-2 and TPX2 was not additive and occurred without a corresponding increase in T-loop phosphorylation. These results suggest that both I-2 and TPX2 function as allosteric activators of Aurora-A. This implies that I-2 is a bifunctional signaling protein with separate domains to inhibit PP1 and directly stimulate Aurora-A kinase.
ATM, a DNA damage-inducible kinase, contributes to activation by high NaCl of the transcription factor TonEBP/OREBP
Authors: Irarrazabal Carlos E. , Liu Jennifer C. , Burg Maurice B. , Ferraris Joan D.
Published: June 2004

High NaCl activates the transcription factor tonicity-responsive enhancer/osmotic response element-binding protein (TonEBP/OREBP), resulting in increased transcription of several protective genes, including the glycine betaine/γ-aminobutyric acid transporter (
BGT1
). High NaCl damages DNA, and DNA damage activates ataxia telangiectasia mutated (ATM) kinase through autophosphorylation on Ser-1981. TonEBP/OREBP contains ATM consensus phosphorylation sites at Ser-1197, Ser-1247, and Ser-1367. The present studies test whether ATM is involved in activation of TonEBP/OREBP by high NaCl. We find that raising osmolality from 300 to 500 mosmol/kg by adding NaCl activates ATM, as indicated by phosphorylation at Ser-1981. High urea and radiation also activate ATM, but they do not increase TonEBP/OREBP transcriptional activity like high NaCl does. Wortmannin, which inhibits ATM, reduces NaCl-induced TonEBP/OREBP transcriptional activation and
BGT1
mRNA increase. Overexpression of wild-type TonEBP/OREBP increases ORE/TonE reporter activity much more than does overexpression of TonEBP/OREBP S1197A, S1247A, or S1367A. In AT cells (which express nonfunctional ATM), TonEBP/OREBP transcriptional and transactivating activity are further increased by expression of wild-type ATM but not of S1981A ATM. TonEBP/OREBP reciprocally coimmunoprecipitates with ATM kinase, demonstrating physical association. Additionally, antibody to ATM kinase supershifts TonEBP/OREBP bound to its cognate ORE/TonE DNA element. In AT cells, wortmannin further decreases high NaCl-induced increase in transcriptional activity, consistent with participation of signaling kinase(s) in addition to ATM. In conclusion, signaling via ATM is necessary for full activation of TonEBP/OREBP by high NaCl, but it is not sufficient.
Isolation of hnRNP complexes from Drosophila melanogaster.
Authors: Matunis M J , Matunis E L , Dreyfuss G
Published: May 2004

Nascent RNA polymerase II transcripts, heterogeneous nuclear RNAs (hnRNAs), become associated with nuclear proteins (hnRNP Proteins), and their processing into mRNAs takes place in these hnRNP complexes. hnRNP complexes have previously been purified from vertebrate cells. Here we report the isolation of hnRNP complexes from an invertebrate organism, the fruitfly Drosophila melanogaster. Candidate hnRNP proteins were purified from D. melanogaster embryos by ssDNA affinity chromatography, and mAbs were produced to many of the major proteins. Genuine hnRNP proteins were identified by several criteria, including nucleoplasmic localization, association with nascent transcripts, crosslinking to poly(A)-containing RNA in living cells, and amino acid sequence. In addition, mAbs that cross-react between the fruitfly and human hnRNP proteins were obtained. Most importantly, using hnRNP-specific mAbs we have purified the hnRNP complexes from D. melanogaster cells. These RNAase-sensitive complexes contain at least 10 major proteins designated hrps, the most abundant proteins having apparent molecular masses of 36, 38, 39, 40, 44, 48, 54, 62, 70, and 75 kD. cDNAs and complete sequences for several of these proteins have been obtained and are presented in the accompanying paper (Matunis, E. L., M. J. Matunis, and G. Dreyfuss. 1992. J. Cell Biol. 116:257-269). The purification of D. melanogaster hnRNP complexes will facilitate genetic and cytological studies on the function of hnRNA-binding proteins and on the posttranscriptional regulation of gene expression.
Characterization of the major hnRNP proteins from Drosophila melanogaster.
Authors: Matunis E L , Matunis M J , Dreyfuss G
Published: May 2004

To better understand the role(s) of hnRNP proteins in the process of mRNA formation, we have identified and characterized the major nuclear proteins that interact with hnRNAs in Drosophila melanogaster. cDNA clones of several D. melanogaster hnRNP proteins have been isolated and sequenced, and the genes encoding these proteins have been mapped cytologically on polytene chromosomes. These include the hnRNP proteins hrp36, hrp40, and hrp48, which together account for the major proteins of hnRNP complexes in D. melanogaster (Matunis et al., 1992, accompanying paper). All of the proteins described here contain two amino-terminal RNP consensus sequence RNA-binding domains and a carboxyl-terminal glycine-rich domain. We refer to this configuration, which is also found in the hnRNP A/B proteins of vertebrates, as 2 x RBD-Gly. The sequences of the D. melanogaster hnRNP proteins help define both highly conserved and variable amino acids within each RBD and support the suggestion that each RBD in multiple RBD-containing proteins has been conserved independently and has a different function. Although 2 x RBD-Gly proteins from evolutionarily distant organisms are conserved in their general structure, we find a surprising diversity among the members of this family of proteins. A mAb to the hrp40 proteins crossreacts with the human A/B and G hnRNP proteins and detects immunologically related proteins in divergent organisms from yeast to man. These data establish 2 x RBD-Gly as a prevalent hnRNP protein structure across eukaryotes. This information about the composition of hnRNP complexes and about the structure of hnRNA-binding proteins will facilitate studies of the functions of these proteins.
Translocation of oxysterol binding protein to Golgi apparatus triggered by ligand binding.
Authors: Ridgway N D , Dawson P A , Ho Y K , Brown M S , Goldstein J L
Published: May 2004

A cDNA encoding a cytoplasmic oxysterol binding protein was expressed at high levels by transfection in animal cells. This protein binds oxysterols such as 25-hydroxycholesterol that regulate sterol metabolism by transcriptional and posttranscriptional effects. In the transfected cells, some of the oxysterol binding protein (OSBP) was distributed diffusely in the cytoplasm, and some was bound to small vesicles near the nucleus, as revealed by indirect immunofluorescence. Upon addition of 25-hydroxycholesterol, most of the OSBP became concentrated in large perinuclear structures that stained with lentil lectin, a protein that stains the Golgi apparatus. The structures that contained OSBP were disrupted by brefeldin A, confirming their identification as Golgi. A mutant OSBP lacking the COOH-terminal oxysterol binding domain localized to the Golgi spontaneously, suggesting that this domain normally occludes the domain that binds to the Golgi and that sterols relieve this occlusion. The previously noted potential leucine zipper sequence in OSBP was not required for Golgi localization, nor was it essential for homodimer formation. We conclude that OSBP is triggered to bind extrinsically to Golgi membranes when it binds oxysterols and speculate that this translocation may play a role in the transport, metabolism, or regulatory actions of oxysterols.
Bovine chromaffin granule membranes undergo Ca(2+)-regulated exocytosis in frog oocytes.
Authors: Scheuner D , Logsdon C D , Holz R W
Published: May 2004

We have devised a new method that permits the investigation of exogenous secretory vesicle function using frog oocytes and bovine chromaffin granules, the secretory vesicles from adrenal chromaffin cells. Highly purified chromaffin granule membranes were injected into Xenopus laevis oocytes. Exocytosis was detected by the appearance of dopamine-beta-hydroxylase of the chromaffin granule membrane in the oocyte plasma membrane. The appearance of dopamine-beta-hydroxylase on the oocyte surface was strongly Ca(2+)-dependent and was stimulated by coinjection of the chromaffin granule membranes with InsP3 or Ca2+/EGTA buffer (18 microM free Ca2+) or by incubation of the injected oocytes in medium containing the Ca2+ ionophore ionomycin. Similar experiments were performed with a subcellular fraction from cultured chromaffin cells enriched with [3H]norepinephrine-containing chromaffin granules. Because the release of [3H]norepinephrine was strongly correlated with the appearance of dopamine-beta-hydroxylase on the oocyte surface, it is likely that intact chromaffin granules and chromaffin granule membranes undergo exocytosis in the oocyte. Thus, the secretory vesicle membrane without normal vesicle contents is competent to undergo the sequence of events leading to exocytosis. Furthermore, the interchangeability of mammalian and amphibian components suggests substantial biochemical conservation of the regulated exocytotic pathway during the evolutionary progression from amphibians to mammals.
Myosin-I moves actin filaments on a phospholipid substrate: implications for membrane targeting.
Authors: Zot H G , Doberstein S K , Pollard T D
Published: May 2004

Acanthamoeba myosin-I bound to substrates of nitrocellulose or planar lipid membranes on glass moved actin filaments at an average velocity of 0.2 micron/s. This movement required ATP and phosphorylation of the myosin-I heavy chain. We prepared planar lipid membranes on a glass support by passive fusion of lipid vesicles (Brian, A. A., and H. M. McConnell. 1984. Proc. Natl. Acad. Sci. USA. 81:6159-6163) composed of phosphatidylcholine and containing 0-40% phosphatidylserine. The mass of lipid that bound to the glass was the same for membranes of 2 and 20% phosphatidylserine in phosphatidylcholine and was sufficient to form a single bilayer. Myosin-I moved actin filaments on planar membranes of 5-40% but not 0-2% phosphatidylserine. At the low concentrations of phosphatidylserine, actin filaments tended to detach suggesting that less myosin-I was bound. We used the cooperative activation of Acanthamoeba myosin-I ATPase by low concentrations of actin to assess the association of phospholipids with myosin-I. Under conditions where activity depends on the binding of actin to the tail of myosin-I (Albanesi, J. P., H. Fujisaki, and E. D. Korn. 1985. J. Biol. Chem. 260:11174-11179), phospholipid vesicles with 5-40% phosphatidylserine inhibited ATPase activity. The motility and ATPase results demonstrate a specific interaction of the tail of myosin-I with physiological concentrations of phosphatidylserine. This interaction is sufficient to support motility and may provide a mechanism to target myosin-I to biological membranes.
The HB-6, CDw75, and CD76 differentiation antigens are unique cell-surface carbohydrate determinants generated by the beta-galactoside alpha 2,6-sialyltransferase.
Authors: Bast B J , Zhou L J , Freeman G J , Colley K J , Ernst T J , Munro J M , Tedder T F
Published: May 2004

Expression of the beta-galactoside alpha 2,6-sialyltransferase (alpha 2,6-ST) was shown to regulate the generation of multiple cell-surface differentiation antigens (Ags) that may be necessary for lymphocyte function. A new mAb was produced, termed HB-6, that was shown to identify a novel neuraminidase-sensitive cell-surface Ag expressed by subpopulations of human lymphocytes and erythrocytes. In attempting to isolate a cDNA encoding the HB-6 antigen by expression cloning, a cDNA encoding the alpha 2,6-ST (EC 2.4.99.1) was obtained. Since expression of the alpha 2,6-ST protein was shown to be limited to the Golgi apparatus, the cell-surface HB-6 Ag was demonstrated to be the product of alpha 2,6-ST activity. Interestingly, alpha 2,6-ST expression also generated two other neuraminidase-sensitive lymphocyte cell-surface differentiation Ags, CDw75, and CD76. The HB-6, CDw75, and CD76 mAb identified distinct Ags that were differentially expressed by different B cell lines and exhibited different patterns of expression in tissue sections. These results indicate that alpha 2,6-ST expression is a critical regulatory step in the formation of the Ags that are recognized by these mAb, and that an alpha 2,6-linked sialic acid residue is an essential component of each Ag. Thus, expression of a single ST can result in the generation of multiple distinct antigenic determinants on the cell surface which can be distinguished by mAb and may have regulatory roles in lymphocyte function.
Compartmentalization of PDGF on extracellular binding sites dependent on exon-6-encoded sequences.
Authors: Raines E W , Ross R
Published: May 2004

The PDGFs are a family of molecules assembled as disulfide-bonded homo- and heterodimers from two distinct but highly homologous polypeptide chains (PDGF-A and PDGF-B). Two PDGF A-chain transcripts, which arise from alternative usage of the 69-bp exon 6 and exon 7, give rise to two forms of PDGF-A. In spite of the conservation of two PDGF A-chain forms over at least 350 million years, no differences in their biological activities have been identified. We have investigated the activity of the sequence encoded by the alternatively spliced exon 6 of the PDGF A-chain (peptide AL). Addition of peptide AL at 10(-5)-10(-9) M to cultured endothelium and smooth muscle induced a dose-dependent, 3-20-fold increase in PDGF in conditioned media within 30 min. Peptide AL had no detectable effect on A- or B-chain transcript levels, and decrease in culture temperature did not prevent rapid release of PDGF. In human umbilical vein endothelial cells treated with peptide AL, the PDGF release was principally PDGF-BB, while in smooth muscle cells it was primarily PDGF-AA. The capacity to induce release of PDGF is shared by the homologous peptide encoded by exon 6 of the B-chain of PDGF. Binding studies and cross-linking analysis are consistent with a charge-based association of exon 6 sequences with membrane- and matrix-associated heparan-sulfate proteoglycans. We hypothesize that translation of exon 6 of the A- or B-chain of PDGF results in compartmentalization of these forms of PDGF with HS-PG, whereas forms lacking this sequence would be soluble and diffuse.
Human basement membrane heparan sulfate proteoglycan core protein: a 467-kD protein containing multiple domains resembling elements of the low density lipoprotein receptor, laminin, neural cell adhesion molecules, and epidermal growth factor.
Authors: Kallunki P , Tryggvason K
Published: May 2004

The primary structure of the large human basement membrane heparan sulfate proteoglycan (HSPG) core protein was determined from cDNA clones. The cDNA sequence codes for a 467-kD protein with a 21-residue signal peptide. Analysis of the amino acid sequence showed that the protein consists of five domains. The amino-terminal domain I contains three putative heparan sulfate attachment sites; domain II has four LDL receptor-like repeats; domain III contains repeats similar to those in the short arms of laminin; domain IV has lg-like repeats resembling those in neural cell adhesion molecules; and domain V contains sequences resembling repeats in the G domain of the laminin A chain and repeats in the EGF. The domain structure of the human basement membrane HSPG core protein suggests that this mosaic protein has evolved through shuffling of at least four different functional elements previously identified in other proteins and through duplication of these elements to form the functional domains. Comparison of the human amino acid sequence with a partial amino acid sequence from the corresponding mouse protein (Noonan, D. M., E. A. Horigan, S. R. Ledbetter, G. Vogeli, M. Sasaki, Y. Yamada, and J. R. Hassell. 1988. J. Biol. Chem. 263:16379-16387) shows a major difference between the species in domain IV, which contains the Ig repeats: seven additional repeats are found in the human protein inserted in the middle of the second repeat in the mouse sequence. This suggests either alternative splicing or a very recent duplication event in evolution. The multidomain structure of the basement membrane HSPG implies a versatile role for this protein. The heparan sulfate chains presumably participate in the selective permeability of basement membranes and, additionally, the core protein may be involved in a number of biological functions such as cell binding, LDL-metabolism, basement membrane assembly, calcium binding, and growth- and neurite-promoting activities.
Activated oncogenes promote and cooperate with chromosomal instability for neoplastic transformation
Authors: Woo Richard A. , Poon Randy Y.C.
Published: June 2004

Most cancer cells are aneuploid. The chromosomal instability hypothesis contends that aneuploidy is the catalyst for transformation, whereas the gene mutation hypothesis asserts that cancer is driven by mutations to proto-oncogenes and tumor-suppressor genes, with the aneuploidy a side effect of tumorigenesis. Because genotoxic stress induced by “culture shock” can obscure the transforming potential of exogenous genes, we cultured wild-type and p53-/- mouse embryo fibroblasts in a more physiological (serum-free) environment. Under these conditions, the cells were immortal and, more importantly, chromosomally stable. Expression of oncogenic H-RasV12 did not induce senescence, but sensitized these cells to p53-dependent apoptosis. In addition, H-RasV12 induced chromosomal instability, as well as accumulation and phosphorylation of p53. Significantly, whereas cells grown under standard conditions could be transformed by coexpression of H-RasV12 and E1A, the chromosomally stable cells were refractory to transformation, as measured by anchorage-independent growth and tumor formation in nude mice. These oncogenes required a third genetic alteration that abolished the p53 pathway to create a permissive environment that promotes rapid chromosomal instability and transformation. Oncogene-induced chromosomal instability and transformation was attenuated by antioxidants. These data indicate that chromosomal instability could be a catalyst for oncogenic transformation, and bring together aspects of the chromosomal instability hypothesis and the gene mutation hypothesis for tumorigenesis.
Signaling hierarchy downstream of retinoic acid that independently regulates vascular remodeling and endothelial cell proliferation
Authors: Bohnsack Brenda L. , Lai Lihua , Dolle Pascal , Hirschi Karen K.
Published: June 2004

We previously demonstrated that during vascular morphogenesis, retinoic acid (RA) is required for the control of endothelial cell proliferation and capillary plexus remodeling. Herein, we investigate the mechanisms by which RA regulates these processes in the yolk sac. We found that although the enzyme required for RA production during early embryogenesis, retinaldehyde dehydrogenase-2 (Raldh2), was expressed in the visceral endoderm, RA receptors α1 and α2 were expressed in endothelial cells in the mesoderm, indicating that they are direct targets of RA. In Raldh2-/- embryos, there was down-regulation of TGF-β1, fibronectin (Fn) and integrin α5, which was associated with decreased visceral endoderm survival and production of VEGF-A, Indian hedgehog (IHH), and bFGF. Exogenous provision of RA or Fn to Raldh2-/- explants in whole mouse embryo culture restored vascular remodeling, visceral endoderm survival, as well as integrin α5 expression and its downstream signaling that controls endothelial growth. Exogenous provision of visceral endoderm-derived factors (VEGF-A, IHH, and bFGF) failed to rescue endothelial cell proliferative control but collectively promoted vascular remodeling, suggesting that these processes are independently regulated via a signaling hierarchy downstream of RA.
Rad54 and DNA Ligase IV cooperate to maintain mammalian chromatid stability
Authors: Mills Kevin D. , Ferguson David O. , Essers Jeroen , Eckersdorff Mark , Kanaar Roland , Alt Frederick W.
Published: June 2004

Nonhomologous end joining (NHEJ) and homologous recombination (HR) represent the two major pathways of DNA double-strand break (DSB) repair in eukaryotic cells. NHEJ repairs DSBs by ligation of cognate broken ends irrespective of homologous flanking sequences, whereas HR repairs DSBs using an undamaged homologous template. Although both NHEJ and HR have been clearly implicated in the maintenance of genome stability, how these apparently independent and mechanistically distinct pathways are coordinated remains largely unexplored. To investigate the relationship between HR and NHEJ modes of DSB repair, we generated cells doubly deficient for the NHEJ factor DNA Ligase IV (Lig4) and the HR factor Rad54. We show that Lig4 and Rad54 cooperate to support cellular proliferation, repair spontaneous DSBs, and prevent chromosome and single chromatid aberrations. These findings demonstrate a role for NHEJ in the repair of DSBs that occur spontaneously during or after DNA replication, and reveal overlapping functions for NHEJ and Rad54-dependent HR in the repair of such DSBs.
Collaboration of homologous recombination and nonhomologous end-joining factors for the survival and integrity of mice and cells
Authors: Couëdel Chrystelle , Mills Kevin D. , Barchi Marco , Shen Lingbo , Olshen Adam , Johnson Roger D. , Nussenzweig André , Essers Jeroen , Kanaar Roland , Li Gloria C. , Alt Frederick W. , Jasin Maria
Published: June 2004

Homologous recombination (HR) and nonhomologous end-joining (NHEJ) are mechanistically distinct DNA repair pathways that contribute substantially to double-strand break (DSB) repair in mammalian cells. We have combined mutations in factors from both repair pathways, the HR protein Rad54 and the DNA-end-binding factor Ku80, which has a role in NHEJ. Rad54-/-Ku80-/- mice were severely compromised in their survival, such that fewer double mutants were born than expected, and only a small proportion of those born reached adulthood. However, double-mutant mice died at lower frequency from tumors than Ku80 single mutant mice, likely as a result of rapid demise at a young age from other causes. When challenged with an exogenous DNA damaging agent, ionizing radiation, double-mutant mice were exquisitely sensitive to low doses. Tissues and cells from double-mutant mice also showed indications of spontaneous DNA damage. Testes from some Rad54-/-Ku80-/- mice displayed enhanced apoptosis and reduced sperm production, and embryonic fibroblasts from Rad54-/-Ku80-/- animals accumulated foci of γ-H2AX, a marker for DSBs. The substantially increased DNA damage response in the double mutants implies a cooperation of the two DSB repair pathways for survival and genomic integrity in the animal.
Competency requirements for managerial development in manufacturing, assembly, and/or material processing functions
Authors: Barber Clifford S. , Tietje Brian C.
Published: June 2004

Organizations that include a manufacturing, assembly and/or material processing (MAMP) functions have become increasingly complex, competitive, and dynamic. These organizations demand increased emphasis on recruiting, developing, and retaining management talent to gain a competitive edge. This paper uses a modified Delphi triangulation approach to identify and categorize first‐level MAMP management competencies perceived to be important by three critical stakeholder groups – an expert MAMP upper management panel, first‐level MAMP managers who have been employed between one to three years, and business management faculty members. The results identify 14 knowledge, skill, and value‐based competencies and three higher‐order factors as essential components within the MAMP management function. There was considerable consistency in the perceptions of the three groups in evaluating the importance of these competencies, with the exception that first‐level managers placed greater importance on technical skills for daily MAMP activities than upper management.
Supercooling and cold‐hardiness in eggs of western and northern corn rootworms
Authors: Ellsbury M. M. , Lee R. E.
Published: May 2004

Oviposition by northern corn rootworms, Diabrotica barberi Smith and Lawrence, and western corn rootworms, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), key pests of corn in the Great Plains of the USA, occurs in the soil during late summer. Overwintering eggs are exposed to variable soil moisture and temperatures below −5 °C. The winter mortality of eggs in the soil is a primary factor that determines the potential for larval injury to corn the following spring. Our studies aimed to determine the comparative supercooling capacities of northern and western corn rootworm eggs and to assess egg mortality following brief exposure to extreme low temperature, ranging from −12.0 to −21.5 °C, under three moisture regimes. Eggs of northern corn rootworm were supercooled to a temperature as low as −27 °C, and survived supercooling to a greater extent than did western corn rootworm eggs. Moisture treatment prior to supercooling had little effect on northern corn rootworm eggs. Western corn rootworm eggs were more resistant than northern corn rootworm eggs to the effects of desiccation followed by supercooling. The survival of northern corn rootworm eggs was better than western corn rootworms under dry conditions, followed by exposure to temperatures of −12.0 and −17.5 °C, but was very low at −21.5 °C, regardless of the moisture regime. The results suggest that moisture and temperature may interact in the soil environment to determine the overwintering survival of corn rootworms. It is evident from these studies that both rootworm species experience mortality at temperatures well above the supercooling points of the eggs, but that differences exist in the effects of substrate moisture treatments on the cold‐hardiness of eggs from the two species.
Comparing floral nectar and aphid honeydew diets on the longevity and nutrient levels of a parasitoid wasp
Authors: Lee Jana C. , Heimpel George E. , Leibee Gary L.
Published: May 2004

We compared the effects of floral nectar from buckwheat, Fagopyrum esculentum Moench, and honeydew produced by the soybean aphid, Aphis glycines Matsumura (Homoptera: Aphididae), on longevity, nutrient levels, and egg loads of the parasitoid Diadegma insulare Cresson (Hymenoptera: Ichneumonidae). Diadegma insulare lived for 2 days in control treatments of water or clean soybean leaves, for 6–7 days with honeydew, and in excess of 2 weeks with buckwheat nectar. Potential reasons for the superiority of buckwheat nectar over soybean aphid honeydew for extending the longevity of parasitoids include: (i) parasitoids ingest more sugars from floral sources, (ii) oligosaccharides in honeydew have a lower nutritional value than nectar sugars, and (iii) honeydew has antagonistic compounds. Overall sugar levels were lower in honeydew‐ vs. nectar‐fed female wasps, suggesting a lower feeding rate, but other explanations cannot be excluded. Diadegma insulare eclosed with high levels of lipids and glycogen, and low levels of gut and storage sugars. All carbohydrates increased over the life of both nectar‐ and honeydew‐fed wasps, but remained low or decreased in starved wasps. Lipid levels declined over the lifespan of female wasps, but females fed floral nectar showed the slowest rate of lipid decline. Diet did not affect egg load, probably because the females were not given hosts in the experiment.
Feeding responses of specialist herbivores to plant extracts and pure allelochemicals: effects of prolonged exposure
Authors: Akhtar Yasmin , Isman Murray B.
Published: May 2004

We investigated the possibility of decreased feeding deterrent response following prolonged exposure in three specialist herbivores, Plutella xylostella (larvae), Pseudaletia unipuncta (larvae), and Epilachna varivestis (adults) through leaf disc choice and no‐choice tests. Neonate larvae (<24 h old) of the three species were reared on their host plants sprayed with the antifeedants until tested. Our results demonstrated that Ps. unipuncta did not show a decrease in feeding deterrent response to extracts of Melia volkensii (choice and no‐choice tests) or oregano (Origanum vulgare) (choice test) following prolonged exposure. Plutella xylostella exhibited a decrease in feeding deterrent response to M. volkensii in a choice test only, but not to oregano. Although not significant, we did find a trend toward decreasing feeding deterrent response to M. volkensii by both species in no‐choice tests. However, both species exhibited a significant decrease in feeding deterrent response to pure allelochemicals (Ps. unipuncta to thymol, and P. xylostella to thymol and toosendanin) compared with the naïve groups, following prolonged exposure in leaf disc choice tests. Epilachna varivestis showed a significant decrease in feeding deterrent responses to both oregano and thymol in leaf disc choice tests. We conclude from our present and previous studies that not only are there interspecific differences between generalist and specialist species but also among specialist species.
Influence of plant sulphur nutrition on oviposition and larval performance of the diamondback moth
Authors: Marazzi Cristina , Städler Erich
Published: May 2004

We measured adult oviposition preference, larval growth, and feeding behaviour of the crucifer specialist Plutella xylostella (L.) (Lepidoptera: Plutellidae) on plants of Brassica napus (L.) cv. Express (Brassicaceae), grown under three different sulphur regimes. The nutrient solutions used were the following: one sulphur‐free (S0), one normal sulphur (Sn, normal field concentration), and one sulphur‐rich (S+, double concentration of Sn). Females laid more eggs on Sn than on S0 plants, while only a slight, non‐significant difference was observed between Sn and S+ plants. Moreover, the development time from hatching to emergence was significantly shorter, and adults were heavier on Sn than on S0 plants. Comparing these same two parameters from Sn and S+ plants, we found a shorter development time on plants rich in sulphur, although this trend was not statistically significant. Larval feeding preferences were tested in a dual choice assay using leaf discs. A significantly higher number of larvae preferred leaf discs of Sn plants than those of S0 plants. Furthermore, the larvae preferred S+ to Sn discs. An optimal supply of sulphur to oilseed rape is necessary for a good seed harvest, and it also plays an important role in acceptance by P. xylostella of the host plant. Maintaining higher levels of sulphur in the plant nutrient solution benefits insect performance, both at the adult and larval stage.