bluuebunny/crossref_metadata_2025 huggingface dataset:
Novel closely spaced planar dual‐band frequency‐selective surface
Authors: Çimen Sibel
Published: August 2013

A novel, dual‐band frequency‐selective surface (FSS) is presented. The designed and fabricated FSS structure has closely spaced operation bands and high selective frequency response at each operation band. Two‐dimensional compact U‐shaped slot resonators are used in each FSS unit cell to obtain this ability. Also, each slot resonator is composed of straight slot section at the centre connected to two U‐shaped arms at each end. The dimensions of the slot resonators are optimised using via full‐wave electromagnetic simulations using CST Microwave Solver. The proposed FSS structure is independent of incident polarisation and angle because of 90° rotation of the unit cell. The validity of the proposed FSS is confirmed by both simulation and measurement results. In the experimental verification, a waveguide measurement setup is chosen. Both measurement and simulation results show that the FSS structure operates at the designed frequencies of 8.80 and 9.54 GHz, which are very close to each other.
Quantum conductance of armchair graphene nanopores with edge impurities
Authors: Qiu Wanzhi , Skafidas Efstratios
Published: August 2013

The quantum conductance of armchair graphene nanopores (aGNPs) with edge impurities is investigated using the tight-binding model and non-equilibrium Green's function method. We find that aGNPs are particularly interesting since their transmission spectra can be easily tuned by pore-edge shaping to produce a variety of electronic transport characteristics. We first examine the local density of states at individual impurity sites. We then study the quantum conductance of aGNPs with various transmission spectra in response to perturbations to on-site energies and hopping coefficients of edge atoms. Insights into transport properties of aGNPs are provided and implications of these findings for designing aGNP devices in interconnection and sensing applications are discussed.
Numerical study of sub-millimeter Gunn oscillations in InP and GaN vertical diodes: Dependence on bias, doping, and length
Authors: García S. , Íñiguez-de-la-Torre I. , Pérez S. , Mateos J. , González T.
Published: August 2013

In this work, we report on Monte Carlo simulations of InP and GaN vertical Gunn diodes to optimize their oscillation frequency and DC to AC conversion efficiency. We show that equivalent operating conditions are achieved by the direct application of a sinusoidal AC voltage superimposed to the DC bias and by the simulation of the intrinsic device coupled with the consistent solution of a parallel RLC resonant circuit connected in series. InP diodes with active region about 1 μm offer a conversion efficiency up to 5.5% for frequencies around 225 GHz. By virtue of the larger saturation velocity, for a given diode length, oscillation frequencies in GaN diodes are higher than for InP structures. Current oscillations at frequencies as high as 675 GHz, with 0.1% efficiency, are predicted at the sixth generation band in a 0.9 μm-long GaN diode, corroborating the suitability of GaN to operate near the THz band. At the first generation band, structures with notch, in general, provide lower oscillation frequencies and efficiencies in comparison with the same structures without notch. However, a higher number of generation bands are originated in notched diodes, thus, typically reaching larger frequencies. Self-heating effects reduce the performance, but in GaN diodes the efficiency is not significantly degraded.
Valence instability and photochemical reaction at surface of strongly correlated MgTi2O4
Authors: Fujiwara H. , Ishige Y. , Mizokawa T. , Sasaki T. , Isobe M. , Ueda Y.
Published: August 2013

We have studied surface electronic structure and light illumination effect of spinel-type MgTi2O4 with Ti3+(d1) electronic configuration. Ti4+ species are found at surfaces of MgTi2O4, which show photo-induced effect similar to the Ti4+-oxide semiconductors with surface depletion layer. MgTi2O4 with the surface Ti4+ states reacts with water for H2 emission, which is largely enhanced by light illumination.
A new perturbed-chain equation of state for square-well chains in fluid and solid phases
Authors: Alavi Farzad , Feyzi Farzaneh
Published: August 2013

Considering the hard-chain system as reference, a perturbed-chain equation of state (EOS) is developed. The second-order thermodynamic perturbation theory EOS is applied to the reference system. Monte Carlo simulation data for average intra-molecular and inter-molecular segment-segment radial distribution function of hard-chain systems with a chain length of 3–10 in the range of packing fraction between 0.1 and 0.72, covering both fluid and solid phases, are reported. A disordered solid phase structure is considered in this work. These customized data are used to develop the perturbation term of square-well (SW) attractions. The performance of perturbed-chain EOS is tested against computer simulation data from the literature for compressibility factor and phase equilibrium in the systems of SW chains. Results within good accuracy are obtained for all the test cases. Global vapor-liquid-solid equilibrium diagrams for SW chain systems predicted by the new EOS are reported.
Explosive synchronization transitions in complex neural networks
Authors: Chen Hanshuang , He Gang , Huang Feng , Shen Chuansheng , Hou Zhonghuai
Published: August 2013

It has been recently reported that explosive synchronization transitions can take place in networks of phase oscillators [Gómez-Gardeñes et al. Phys. Rev. Lett. 106, 128701 (2011)] and chaotic oscillators [Leyva et al. Phys. Rev. Lett. 108, 168702 (2012)]. Here, we investigate the effect of a microscopic correlation between the dynamics and the interacting topology of coupled FitzHugh-Nagumo oscillators on phase synchronization transition in Barabási-Albert (BA) scale-free networks and Erdös-Rényi (ER) random networks. We show that, if natural frequencies of the oscillations are positively correlated with node degrees and the width of the frequency distribution is larger than a threshold value, a strong hysteresis loop arises in the synchronization diagram of BA networks, indicating the evidence of an explosive transition towards synchronization of relaxation oscillators system. In contrast to the results in BA networks, in more homogeneous ER networks, the synchronization transition is always of continuous type regardless of the width of the frequency distribution. Moreover, we consider the effect of degree-mixing patterns on the nature of the synchronization transition, and find that the degree assortativity is unfavorable for the occurrence of such an explosive transition.
Deterministic and stochastic bifurcations in the Hindmarsh-Rose neuronal model
Authors: Dtchetgnia Djeundam S. R. , Yamapi R. , Kofane T. C. , Aziz-Alaoui M. A.
Published: August 2013

We analyze the bifurcations occurring in the 3D Hindmarsh-Rose neuronal model with and without random signal. When under a sufficient stimulus, the neuron activity takes place; we observe various types of bifurcations that lead to chaotic transitions. Beside the equilibrium solutions and their stability, we also investigate the deterministic bifurcation. It appears that the neuronal activity consists of chaotic transitions between two periodic phases called bursting and spiking solutions. The stochastic bifurcation, defined as a sudden change in character of a stochastic attractor when the bifurcation parameter of the system passes through a critical value, or under certain condition as the collision of a stochastic attractor with a stochastic saddle, occurs when a random Gaussian signal is added. Our study reveals two kinds of stochastic bifurcation: the phenomenological bifurcation (P-bifurcations) and the dynamical bifurcation (D-bifurcations). The asymptotical method is used to analyze phenomenological bifurcation. We find that the neuronal activity of spiking and bursting chaos remains for finite values of the noise intensity.
Potential barrier and band structure of closed edge graphene
Authors: He Chunshan , Wang Weiliang
Published: August 2013

The atomic structure, electron distribution, work function, and band structure of closed edge graphene are investigated with density functional theory. Field emission performance of closed edge graphene is compared with that of open edge graphene. We provide a possible explanation for the field emission microscopy image change after high emission current, which appeals to the experimentalists for further investigation.
Dielectric behavior of graphene/BaTiO3/polyvinylidene fluoride nanocomposite under high electric field
Authors: Shen Yang , Guan Yuhan , Hu Yuhan , Lei Yuechuan , Song Yu , Lin Yuanhua , Nan Ce-Wen
Published: August 2013

Three-phase graphene/BaTiO3/polyvinylidene fluoride composites films are prepared with a simple solution-cast method. The effects of graphene oxides (GOs) and reduced graphene oxides (RGOs) on the dielectric behavior of the three-phase composites are investigated at elevated electric field up to 150 kV/mm. BaTiO3 nanoparticles isolate the graphene nanosheets, leading to much higher percolation threshold. The introduction of both RGOs and GOs increases the leakage current in the three-phase composites at elevated electric field. The positive-up-negative-down measurements indicate that the switched polarization slightly increases with the addition of RGOs or GOs and is much smaller than the remnant polarization obtained from the P-E loops.
Biogas production from broiler manure, wastewater treatment plant sludge, and greenhouse waste by anaerobic co-digestion
Authors: Sarikaya E. , Demirer G. N.
Published: August 2013

Inappropriate management of organic wastes can cause serious damage to the environment by polluting water and air, which can lower the quality of life. Ammonia and greenhouse gases (CH4 and CO2) emitted from the waste storage units can pollute the air. Inappropriate application of nitrogen and phosphorus on fields as manure can lead to eutrophication of surface water resources and pollution of soil and ground water. Conversion of the organic wastes to biogas through anaerobic digestion will, however, reduce the adverse impact on the environment and will contribute to a reduction in the consumption of fossil fuels. This study investigated the anaerobic digestion of broiler manure and greenhouse waste as well as anaerobic co-digestion of broiler manure and dewatered and non-dewatered wastewater treatment plant sludge for the production of biogas. To this purpose, biochemical methane potential experiments were performed. Moreover, the effects of nutrient and trace metal supplementation on the biogas yield were also examined. The results of this study indicated that (1) The obtained biogas values for broiler manure and greenhouse waste were very well in agreement with relevant literature values and comparable to that of food waste reported in the literature, respectively. (2) The experimental biogas production observed for co-digestion of broiler manure and dewatered and non-dewatered wastewater treatment plant sludge is higher than literature values.
Molecular dynamics study of amorphous Ga-doped In2O3: A promising material for phase change memory devices
Authors: Ramzan M. , Kaewmaraya T. , Ahuja R.
Published: August 2013

In this paper, we employ the ab-initio molecular dynamics simulations, within the framework of density functional theory, to construct and characterize the amorphous structure of gallium-doped indium oxide. The electronic properties of amorphous and crystalline structures of Ga:In2O3 are investigated and compared by calculating the density of states, radial distribution function, bond angle distribution, and Bader charge analysis. We observe the band gap closure in amorphous structure, which corresponds to semiconductor to metallic transition in this material on amorphization. Our calculated results show the same characteristics of Ga:In2O3 with other phase change memory (PCM) materials available in literature and suggest it a promising candidate for PCM devices.
Continuous and unconstrained manipulation of micro-particles using phase-control of bulk acoustic waves
Authors: Greenhall J. , Guevara Vasquez F. , Raeymaekers B.
Published: August 2013

A method of unconstrained and continuous manipulation of micro-particles in a fluid using bulk acoustic waves is theoretically derived and experimentally demonstrated. The method is based on phase-control of standing pressure waves created by two opposing transducers. Reflections are taken into account, removing the need for complex experiments. The operating domain of this method is characterized and compared to existing techniques. In contrast to methods based on linearly adjusting the phase difference between opposing transducers, it is shown that by independently controlling the phase of each transducer, particles can be manipulated in an unconstrained manner over multiple wavelengths.
Engineered knottin peptide enables noninvasive optical imaging of intracranial medulloblastoma
Authors: Moore Sarah J. , Hayden Gephart Melanie G. , Bergen Jamie M. , Su YouRong S. , Rayburn Helen , Scott Matthew P. , Cochran Jennifer R.
Published: August 2013

Central nervous system tumors carry grave clinical prognoses due to limited effectiveness of surgical resection, radiation, and chemotherapy. Thus, improved strategies for brain tumor visualization and targeted treatment are critically needed. We demonstrate that mouse cerebellar medulloblastoma (MB) can be targeted and illuminated with a fluorescent, engineered cystine knot (knottin) peptide that binds with high affinity to α
v
β
3
, α
v
β
5
, and α
5
β
1
integrin receptors. This integrin-binding knottin peptide, denoted EETI 2.5F, was evaluated as a molecular imaging probe in both orthotopic and genetic models of MB. Following tail vein injection, fluorescence arising from dye-conjugated EETI 2.5F was localized to the tumor compared with the normal surrounding brain tissue, as measured by optical imaging. The imaging signal intensity correlated with tumor volume. Due to its unique ability to bind to α
5
β
1
integrin, EETI 2.5F showed superior in vivo and ex vivo brain tumor imaging contrast compared with other engineered integrin-binding knottin peptides and with c(RGDfK), a well-studied integrin-binding peptidomimetic. Next, EETI 2.5F was fused to an antibody fragment crystallizable (Fc) domain (EETI 2.5F–Fc) to determine if a larger integrin-binding protein could also target intracranial brain tumors. EETI 2.5F–Fc, conjugated to a fluorescent dye, illuminated MB following i.v. injection and was able to distribute throughout the tumor parenchyma. In contrast, brain tumor imaging signals were not detected in mice injected with EETI 2.5F proteins containing a scrambled integrin-binding sequence, demonstrating the importance of target specificity. These results highlight the potential of using EETI 2.5F and EETI 2.5–Fc as targeted molecular probes for brain tumor imaging.
The effects of topical Arnica on performance, pain and muscle damage after intense eccentric exercise
Authors: Pumpa Kate L. , Fallon Kieran E. , Bensoussan Alan , Papalia Shona
Published: August 2013

The aim of the study was to determine if topical Arnica is effective in reducing pain, indicators of inflammation and muscle damage, and in turn improve performance in well‐trained males experiencing delayed onset muscle soreness (DOMS). Twenty well‐trained males matched by maximal oxygen uptake (V̇O2 Max) completed a double‐blind, randomised placebo‐controlled trial. Topical Arnica was applied to the skin superficial to the quadriceps and gastrocnemius muscles immediately after a downhill running protocol designed to induce DOMS. Topical Arnica was reapplied every 4 waking hours for the duration of the study. Performance measures (peak torque, countermovement and squat jump), pain assessments (visual analogue scale (VAS) and muscle tenderness) and blood analysis (interleukin‐1 beta, interleukin‐6, tumour necrosis factor‐alpha, C‐reactive protein, myoglobin and creatine kinase) were assessed at seven time points over five days (pre‐, post‐, 4, 24, 48, 72 and 96 hours after the downhill run). Participants in the topical Arnica group reported less pain as assessed through muscle tenderness and VAS 72 hours post‐exercise. The application of topical Arnica did not affect any performance assessments or markers of muscle damage or inflammation. Topical Arnica used immediately after intense eccentric exercise and for the following 96 hours did not have an effect on performance or blood markers. It did however demonstrate the possibility of providing pain relief three days post‐eccentric exercise.
Regulation of HSP70 synthesis by messenger RNA degradation.
Authors: Petersen R B , Lindquist S
Published: August 2013

When Drosophila cells are heat shocked, hsp70 messenger RNA (mRNA) is stable and is translated at high efficiencies. During recovery from heat shock, hsp70 synthesis is repressed and its messenger RNA (mRNA) is degraded in a highly regulated fashion. Dramatic differences in the timing of repression and degradation are observed after heat treatments of different severities. The 3' untranslated region (UTR) of the hsp70 mRNA was sufficient to transfer this regulated degradation to heterologous mRNAs. Altering the translational efficiency of the message or changing its natural translation-termination site did not alter its pattern of regulation, although in some cases it changed the absolute rate of degradation. We have previously shown that hsp70 mRNA is very unstable when it is expressed at normal growth temperatures (from a metallothionein promoter). We report here that the 3' untranslated region of the hsp70 mRNA is responsible for this instability as well. We postulate that a mechanism for degrading hsp70 mRNA pre-exists in Drosophila cells, that it is inactivated by heat shock and that it is the reactivation of this mechanism that is responsible for hsp70 repression during recovery. This degradation system may be the same as that used by other unstable mRNAs.
Nuclear proteins TREF1 and TREF2 bind to the transcriptional control element of the transferrin receptor gene and appear to be associated as a heterodimer.
Authors: Roberts M R , Miskimins W K , Ruddle F H
Published: August 2013

Two novel proteins that bind specifically to the transferrin receptor (TR) promoter, have been isolated from HeLa cell nuclear extract using a combination of ion exchange and oligonucleotide-affinity chromatography. TREF1 and TREF2, which have apparent molecular weights of 82 and 62 kDa, respectively, appear to be associated as a heterocomplex (TREF), and both proteins are able to contact target DNA directly. TREF interacts specifically with a region of the TR promoter which contains the TR transcriptional control element. This region is similar in sequence to the cAMP-responsive and phorbol ester-responsive elements found in several viral and cellular genes. Binding of TREF to the TR promoter results in modification of DNA topology over multiple helical turns, including a sequence revealed by a helical periodicity map as having an unusual structure.
Distribution and isolation of four laminin variants; tissue restricted distribution of heterotrimers assembled from five different subunits.
Authors: Engvall E , Earwicker D , Haaparanta T , Ruoslahti E , Sanes J R
Published: August 2013

The distribution of subunits of the basement membrane proteins laminin and merosin in human and rabbit tissue was studied by immunofluorescence using monoclonal antibodies. The laminin A chain is present in epithelial, endothelial, and smooth muscle basement membranes. Merosin, as defined by its heavy chain M, is present in striated muscle and peripheral nerve. The A subunit colocalizes with at least two B subunits: B2 plus either B1 or the recently discovered B1 homologue S. The M subunit most often colocalizes with B1 and B2. Exceptions include the myotendinous junction, where M colocalizes with S, and the trophoblast basement membrane, where the M subunit colocalizes with S as well as B1. The presence of all five known subunits of the laminin family in placenta allowed isolation of their parent molecules in native form by the use of monoclonal antibodies in affinity chromatography. Four different heterotrimeric proteins could be identified: B1 chain-containing laminin (A-B1-B2), S chain-containing laminin (A-S-B2), B1-containing merosin (M-B1-B2), and S-containing merosin (M-S-B2). The data show that the proteins in the laminin family are heterotrimers composed of one heavy and two light chains; that most basement membranes contain predominantly one protein of the laminin family; and that laminin, as defined by the A subunit, has a much more restricted distribution than previously thought.
Specific growth stimulation of cultured smooth muscle cells from spontaneously hypertensive rats by platelet-derived growth factor A-chain homodimer.
Authors: Resink T J , Scott-Burden T , Hahn A W , Rouge M , Hosang M , Powell J S , Bühler F R
Published: August 2013

Cultured vascular smooth muscle cells (VSMC)1 from spontaneously hypertensive rats (SHR) possess specific cell surface receptors for both homodimeric forms of platelet-derived growth factor (PDGF-AA and PDGF-BB), in contrast to cells from normotensive Wistar Kyoto (WKY) animals, which express receptors only for the B-chain form of PDGF. Stimulation of quiescent VSMC from SHR with PDGF-AA resulted in activation of S6-kinase and induction of phosphoinositide catabolism, as well as cellular proliferation when cultures were maintained for prolonged periods with daily supplementation of the growth factor. WKY-derived VSMC showed no response to PDGF-AA, which was consistent with their lack of specific receptors for this homodimer. The responsiveness of quiescent cells from SHR and WKY to the B-chain homodimer was similar. The enhanced growth responsiveness of SHR-derived cells to fetal calf serum, as compared with cells from their normotensive counterparts, may be accounted for in part by their expression of receptors for the AA homodimer of PDGF.
Heparanase activity expressed by platelets, neutrophils, and lymphoma cells releases active fibroblast growth factor from extracellular matrix.
Authors: Ishai-Michaeli R , Eldor A , Vlodavsky I
Published: August 2013

Incubation of platelets, neutrophils, and lymphoma cells with Descemet's membranes of bovine corneas and with the extracellular matrix (ECM) produced by cultured corneal endothelial cells resulted in release of basic fibroblast growth factor (bFGF), which stimulated the proliferation of 3T3 fibroblasts and vascular endothelial cells. Similar requirements were observed for release of endogenous bFGF stored in Descemet's membrane and of exogenous bFGF sequestered by the subendothelial ECM. Release of ECM-resident bFGF by platelets, neutrophils, and lymphoma cells was inhibited by carrageenan lambda, but not by protease inhibitors, in correlation with the inhibition of heparanase activity expressed by these cells. Degradation of the ECM-heparan sulfate side chains by this endo-beta-D-glucuronidase is thought to play an important role in cell invasion, particularly in the extravasation of blood-borne tumor cells and activated cells of the immune system. We propose that both heparanase and ECM-resident bFGF may modulate the cell response to contact with its local environment. Heparanase-mediated release of active bFGF from storage in basement membranes provides a novel mechanism for a localized induction of neovascularization in various normal and pathological processes, such as wound healing, inflammation, and tumor development.
Plasminogen activation by t-PA on the surface of human melanoma cells in the presence of alpha 2-macroglobulin secretion.
Authors: Bizik J , Lizonová A , Stephens R W , Grófová M , Vaheri A
Published: August 2013

Several human melanoma cell lines produced tissue-type plasminogen activator (t-PA), as detected by zymography and immunocapture assay of culture media and cell lysates. Urokinase (u-PA) was found at only less than or equal to 1% the level of t-PA. Acid eluates of the cell surface indicated that the melanoma cells had t-PA bound on their surface, but no u-PA, and also had a very low capacity to bind exogenous u-PA. After incubation of the melanoma cells with 10% plasminogen-depleted fetal calf serum and human plasminogen, bound plasmin activity could be eluted from the cell surface with tranexamic acid, an analogue of lysine. This indicated that plasminogen was activated on the cell surface. The cell-surface plasmin formation was inhibited by an anti-catalytic monoclonal antibody to human t-PA, and not by an anti-catalytic antibody to u-PA. The melanoma cells also synthesized and secreted alpha 2-macroglobulin (alpha 2M), as shown by alpha 2M-specific mRNA in Northern blotting and detection of alpha 2M protein in conditioned cell culture media. The media were found to inhibit u-PA but not t-PA. This inhibition was related to their alpha 2M content, and immunoabsorption of alpha 2M removed the inhibitory activity. These studies suggest that t-PA can bind to the surface of melanoma cells and generate surface-bound plasmin. Because t-PA and cell-bound plasmin are unaffected by alpha 2M, t-PA may, in the case of melanoma cells, serve an analogous function to u-PA in supporting tumor cell invasion.
The transmembrane anchor of the T-cell antigen receptor beta chain contains a structural determinant of pre-Golgi proteolysis.
Authors: Wileman T , Carson G R , Shih F F , Concino M F , Terhorst C
Published: August 2013

Studies with the T-cell antigen receptor (TCR) have shown that the endoplasmic reticulum, or an organelle closely associated with it, can retain and degrade membrane proteins selectively. The observation that only three (alpha, beta, and delta) of the six (alpha beta gamma delta epsilon zeta) subunits of the TCR are susceptible to proteolysis implies that structural features within the labile proteins mark them for degradation. The TCR beta chain is degraded in the endoplasmic reticulum, and, in this study, we have started to define the domains of the protein that make it susceptible to proteolysis. The experiments show that the transmembrane anchor and short five-amino-acid cytoplasmic tail of the protein contain a dominant determinant of proteolysis. When these residues were removed from the beta chain, the protein became resistant to proteolysis. Even though the resulting ectodomain of the beta chain lacked a transmembrane anchor, it was not secreted by cells and was retained in the endoplasmic reticulum. We conclude that retention in the endoplasmic reticulum alone does not lead to degradation. The results suggest that structural features within the membrane anchor of the protein predispose the beta chain to proteolysis. This was confirmed by replacing the membrane anchor of the interleukin 2 (IL2) receptor, a protein that was stable within the secretory pathway, with that of the TCR beta chain. The unmodified IL2 receptor was transported efficiently to the surface of cells, and an "anchor minus" construct was secreted quantitatively into the culture media. When the membrane anchor of the IL2 receptor was replaced with that of the TCR beta chain, the chimera was unable to reach the Golgi apparatus and was degraded rapidly.
Expression of transfected transforming growth factor alpha induces a motile fibroblast-like phenotype with extracellular matrix-degrading potential in a rat bladder carcinoma cell line.
Authors: Gavrilović J , Moens G , Thiery J P , Jouanneau J
Published: August 2013

Acquisition of cell motility is often correlated with the malignant progression of a transformed cell. To investigate some of the mechanisms involved in the development of a migratory state, we transfected the NBTII rat carcinoma cell line, which forms stationary epithelial clusters in culture, with the gene encoding human transforming growth factor alpha (TGF alpha). Expression of TGF alpha in NBTII cells resulted in cells of motile and vimentin-positive phenotype with internalized desmosomal components, analogous to the treatment of cells with exogenous TGF alpha. The clones expressed a 5.2-kb TGF alpha message and synthesized an 18-kDa form of TGF alpha. Supernatants of TGF alpha-producing clones induced the internalization of desmosomal components, the production of vimentin, and increased motility in untransfected epithelial NBTII cells, indicating that the factor produced by the clones was in a biologically active form. TGF alpha-producing clones secreted significant levels of a 95-kDa gelatinolytic metal-loproteinase, virtually absent in untransfected cell supernatants. In contrast, levels of inhibitors of metalloproteinases and of a plasminogen activator were similar in untransfected and TGF alpha-transfected NBTII cells. These results suggest that expression of TGF alpha in an epithelial tumor cell results in the development of a motile, fibroblast-like phenotype with matrix-degrading potential, which could result in a more aggressive tumor in vivo.
Lysosomal degradation of receptor-bound urokinase-type plasminogen activator is enhanced by its inhibitors in human trophoblastic choriocarcinoma cells.
Authors: Jensen P H , Christensen E I , Ebbesen P , Gliemann J , Andreasen P A
Published: August 2013

We have studied the effect of plasminogen activator inhibitors PAI-1 and PAI-2 on the binding of urokinase-type plasminogen activator (u-PA) to its receptor in the human choriocarcinoma cell line JAR. With 125I-labeled ligands in whole-cell binding assays, both uncomplexed u-PA and u-PA-inhibitor complexes bound to the receptor with a Kd of approximately 100 pM at 4 degrees C. Transferring the cells to 37 degrees C led to degradation to amino acids of up to 50% of the cell-bound u-PA-inhibitor complexes, whereas the degradation of uncomplexed u-PA was 15%; the remaining ligand was recovered in an apparently intact form in the medium or was still cell associated. The degradation could be inhibited by inhibitors of vesicle transport and lysosomal hydrolases. By electron microscopic autoradiography, both 125I-u-PA and 125I-u-PA-inhibitor complexes were located over the cell membrane at 4 degrees C, with the highest density of grains over the membrane at cell-cell interphases, but, after incubation at 37 degrees C, 17 and 27% of the grains for u-PA and u-PA-PAI-1 complexes, respectively, appeared over lysosomal-like bodies. These findings suggest that the u-PA receptor possesses a clearance function for the removal of u-PA after its complex formation with a specific inhibitor. The data suggest a novel mechanism by which receptor-mediated endocytosis is initiated by the binding of a secondary ligand.
A highly conserved intronic sequence is involved in transcriptional regulation of the alpha 1(I) collagen gene.
Authors: Liska D J , Slack J L , Bornstein P
Published: August 2013

The first intron of the human alpha 1(I) collagen gene contains a positive, orientation-dependent cis-acting sequence located between bases +292 and +670. Transient transfection experiments indicate that this sequence is functional in both primary chicken tendon fibroblasts and in a human fibroblast-like cell line derived from SV40-transformed marrow stromal cells. DNase I footprint, methylation interference, and mobility shift analyses provide evidence for a sequence-specific binding activity and show that the region of binding corresponds to a 29-base-pair sequence that is also present in the rat alpha 1(I) collagen intron. This conserved sequence contains an AP1 consensus motif. Sequence-specific binding activity is present in nuclear extracts from HeLa and fibroblast cell lines but not in extracts from two lymphoid cell lines. Mutation of the AP1 consensus sequence indicates that this motif is required for function of the cis-acting element. These data indicate that transcriptional modulation of the alpha 1(I) collagen gene involves an interaction between an intronic AP1-containing sequence and its cognate transcription factors.
Lateral diffusion of nerve growth factor receptor: modulation by ligand-binding and cell-associated factors.
Authors: Venkatakrishnan G , McKinnon C A , Ross A H , Wolf D E
Published: August 2013

We compared the properties in human melanoma cell line A875 and rat pheochromocytoma cell line PC12 of nerve growth factor receptor (NGFr). We also analyzed NGFr and a truncated NGFR lacking the cytoplasmic domain, which were transiently expressed in COS cells. The full-length NGFR expressed in COS cells bound nerve growth factor (NGF) with positive cooperativity, but A875 NGFr and truncated NGFr in COS cells did not display positive cooperativity. The anti-human NGFr monoclonal antibody NGFR5 was characterized and found not to compete with NGF for binding to NGFr. Fabs were prepared from NGFR5 and 192, an anti-rat NGFR monoclonal antibody that was previously shown not to compete with NGF for binding. Fluorescein-labeled Fabs were used to measure the distribution and lateral diffusion of the NGFr. NGFr expressed on COS and A875 cells are diffusely distributed, but NGFr on the surface of PC12 cells appeared, for some cells, to be patched. In A875 cells, 51% of the NGFr was free to diffuse with diffusion coefficient (D) approximately 7 X 10(-10) cm2/s. In COS cells, 43% diffused with D approximately 5 X 10(-10) cm2/s. There was no significant difference in diffusibility between the full-length NGFr and the truncated NGFr. We compared NGFr diffusion on PC12 cells in suspension or adherent to collagen-coated coverslips. For suspension cells, we obtained 32% recovery with D approximately 2.5 X 10(-9) cm2/s. On adherent cells, we obtained 17% recovery with 6 X 10(-9) cm2/s. Binding of NGF enhanced lateral diffusion of NGFr in A875 cells and in PC12 cells in suspension but did not alter lateral diffusion of NGFr in COS cells or in adherent PC12 cells. NGF had no effect on the diffusing fraction or the distribution of NGFR for any cell line.
Tenascin variants: differential binding to fibronectin and distinct distribution in cell cultures and tissues.
Authors: Chiquet-Ehrismann R , Matsuoka Y , Hofer U , Spring J , Bernasconi C , Chiquet M
Published: August 2013

In the chicken, three tenascin variants have been characterized that are generated by alternative splicing of 3 of its 11 fibronectin type III repeats. Using monoclonal antibodies that react with common regions versus extra repeats of tenascin, we could distinguish and separate tenascin variants and investigate their interaction with fibronectin using multiple experimental procedures. Interestingly, in all assays used the smallest tenascin variant bound more strongly to fibronectin than the larger ones. These biochemical data were paralleled by the observation that in chick embryo fibroblast cultures only the smallest form of tenascin could be detected in the fibronectin-rich extracellular matrix network laid down by the cells. Furthermore, each tissue present in adult chicken gizzard contained a distinct set of tenascin variants. Those tissues particularly rich in extracellular matrix, such as the tendon, contained the smallest tenascin only. Intermediate-sized tenascin was present in smooth muscle, whereas the largest form was exclusively detectable underneath the epithelial lining of the villi. Thus it appears that cell type-specific forms of tenascin exist that are appropriate for the functional requirements of the respective extracellular matrices.
Fibronectin/integrin interaction induces tyrosine phosphorylation of a 120-kDa protein.
Authors: Guan J L , Trevithick J E , Hynes R O
Published: August 2013

We describe a 120-kDa protein (pp120) that is phosphorylated on tyrosine in cells attached to fibronectin-coated surfaces. The protein appears to be located in focal contacts where it codistributes with beta 1 integrins. pp120 is distinct from the beta 1 subunit of integrins and from vinculin and alpha-actinin. pp120 is rapidly dephosphorylated in cells suspended by trypsinization but becomes rapidly phosphorylated in cells attaching and spreading on fibronectin. Attachment of cells to RGD-containing peptides, polylysine, or concanavalin A is not sufficient to induce phosphorylation of pp120. The 120-kDa cell-binding domain of fibronectin can induce some phosphorylation of pp120, but further phosphorylation occurs in the presence also of the heparin-binding domain of fibronectin. Phosphorylation of pp120 precedes, but is correlated with, subsequent cell spreading. Phosphorylation of pp120 can also be triggered by attachment of cells to anti-integrin antibodies, and this requires the cytoplasmic domain of the integrin beta 1 subunit. Thus interaction of beta 1 integrins with extracellular ligands (fibronectin or antibodies) triggers phosphorylation of an intracellular 120-kDa protein, pp120, that may be involved in the responses of cells to attachment.
Alpha 2-macroglobulin restricts plasminogen activation to the surface of RC2A leukemia cells.
Authors: Stephens R W , Tapiovaara H , Reisberg T , Bizik J , Vaheri A
Published: August 2013

Human RC2A myelomonocytic leukemia cells are able to activate the prourokinase (pro-u-PA) they secrete so that active u-PA is present both in serum-free conditioned medium from these cells, as well as on the cell surface. When the cells are grown in serum-containing medium, no u-PA activity can be found in the medium but active u-PA is found bound to the cell surface where it can generate bound plasmin. This distribution of u-PA activity was shown to be, first, the net result of slow inactivation of free active u-PA by serum inhibitor(s) and simultaneous rapid uptake of u-PA onto the cell surface. Binding to cells was at least six times faster than inactivation by 10% serum. The principal serum inhibitor of u-PA was identified as alpha 2-macroglobulin (alpha 2M), and prior inactivation of u-PA by purified human alpha 2M was also shown to prevent uptake of u-PA activity onto cells. Second, although endogenous u-PA could form covalent complexes with purified alpha 2M in the culture medium of RC2A cells, covalent alpha 2M complexes were not formed by u-PA on the cell surface; the u-PA taken up in this compartment was protected against alpha 2M inhibition. u-PA anchored to plastic surfaces via monoclonal antibodies to the amino-terminal region of u-PA was also protected against alpha 2M, suggesting that the protection of cell surface u-PA results from a steric effect. These results provide evidence as to how the active u-PA produced by leukemia cells can contribute to proteolytic activity on their cell surface in the presence of serum inhibitors.
Cellular partitioning of beta-1 integrins and their phosphorylated forms is altered after transformation by Rous sarcoma virus or treatment with cytochalasin D.
Authors: Haimovich B , Aneskievich B J , Boettiger D
Published: August 2013

A sequential extraction procedure of 3-[(3-cholamidopropyl)-dimethylammonio]-1-propane sulfonate (CHAPS) buffer followed by RIPA or Laemmli sample buffer was developed to define two distinct subpopulations of beta-1 integrins in primary chicken embryo fibroblasts. Extraction of cells in culture revealed an association of adhesion plaque-localized integrin with the CHAPS-insoluble fraction. Phosphorylated integrins were found in both fractions, but the specific phosphorylation was 12-fold higher in the CHAPS insoluble fraction. The phosphorylation was evenly distributed between phosphoserine and phosphotyrosine. Transformation by Rous sarcoma virus caused a redistribution of integrin to rosettes and an increase in total integrin phosphorylation. Treatment with cytochalasin D caused a redistribution of the adhesion plaque-associated integrin into lacelike structures and reduced the level of integrin phosphorylation. These treatments also caused an altered distribution of phosphorylated integrin between the CHAPS soluble and insoluble fractions. These results suggest a role for integrin phosphorylation in the assembly and disassembly of cellular adhesion structures.
Tumor cell surface alpha 4 beta 1 integrin mediates adhesion to vascular endothelium: demonstration of an interaction with the N-terminal domains of INCAM-110/VCAM-1.
Authors: Taichman D B , Cybulsky M I , Djaffar I , Longenecker B M , Teixidó J , Rice G E , Aruffo A , Bevilacqua M P
Published: August 2013

Hematogenous metastasis involves adhesive interactions between blood-borne tumor cells and the vessel wall. By the use of in vitro assays, the adhesion of human melanoma, osteosarcoma, and kidney carcinoma (but not colon carcinoma) cell lines was shown to involve the cytokine-inducible endothelial cell surface protein inducible cell adhesion molecule 110 (INCAM-110) and the alpha 4 beta 1 integrin, molecules normally involved in endothelial-leukocyte interactions. Tumor adhesion to human endothelial cell monolayers was increased 1.9- to 8.2-fold by endothelial activation with the cytokine tumor necrosis factor (TNF) and inhibited by the anti-INCAM-110 monoclonal antibody (mAb) E1/6. Each of these tumor cells expressed members of the beta 1 integrin family of adhesion molecules, and antibodies to the alpha 4 and beta 1 integrin subunits inhibited tumor-endothelial adhesion (48-87% inhibition). A cDNA encompassing the three N-terminal Ig-like domains of vascular cell adhesion molecule 1 (VCAM-1) encoded a protein recognized by the anti-INCAM-110 mAb E1/6 and, when captured onto plastic, supported melanoma cell adhesion by an alpha 4 integrin-dependent mechanism. In contrast to mAb E1/6, a second anti-INCAM-110 mAb Hu8/4 neither inhibited adhesion to activated endothelium nor bound the first three Ig-like domains of INCAM-110/VCAM-1. These data indicate that the adherence of several human tumors to activated endothelium is mediated by an interaction of alpha 4 beta 1 integrin and the N-terminal Ig-like domains of endothelial INCAM-110/VCAM-1. Tumor acquisition of the alpha 4 integrin subunit and endothelial expression of INCAM-110 may affect the frequency and distribution of metastasis.
Identification of multiple extracellular signal-regulated kinases (ERKs) with antipeptide antibodies.
Authors: Boulton T G , Cobb M H
Published: August 2013

A protein kinase characterized by its ability to phosphorylate microtubule-associated protein-2 (MAP2) and myelin basic protein (MBP) is thought to play a pivotal role in the transduction of signals from many receptors in response to their ligands. A kinase with such activity, named extracellular signal-regulated kinase 1 (ERK1), is activated rapidly by numerous extracellular signals, requires phosphorylation on tyrosine to be fully active, and in vitro can activate a kinase (a ribosomal S6 protein kinase) that is downstream in phosphorylation cascades. From the protein sequence predicted by the rat ERK1 cDNA, peptides were synthesized and used to elicit antibodies. The antibodies recognize both ERK1; a closely related kinase, ERK2; and a third novel ERK-related protein. Using these antibodies we have determined that ERK1 and ERK2 are ubiquitously distributed in rat tissues. Both enzymes are expressed most highly in brain and spinal cord as are their mRNAs. The third ERK protein was found in spinal cord and in testes. The antibodies detect ERKs in cell lines from multiple species, including human, mouse, dog, chicken, and frog, in addition to rat, indicating that the kinases are conserved across species. ERK1 and ERK2 have been separated by chromatography on Mono Q. Stimulation by insulin increases the phosphorylation of both kinases on tyrosine residues, as assessed by immunoblotting with phosphotyrosine antibodies, and retards their elution from Mono Q. Each of these ERKs appears to account for a distinct peak of MBP kinase activity. The activity in each peak is diminished by incubation with either phosphatase 2a or CD45. Therefore, both enzymes have similar modes of regulation and appear to contribute to the growth factor-stimulated MAP2/MBP kinase activity measured in cell extracts.
sar1, a gene from Schizosaccharomyces pombe encoding a protein that regulates ras1.
Authors: Wang Y , Boguski M , Riggs M , Rodgers L , Wigler M
Published: August 2013

Proper ras1 function is required for normal sexual function in the yeast Schizosaccharomyces pombe. We have found a gene in S. pombe, sar1, that encodes a product capable of regulating ras1 function. sar1 is a member of an expanding family of RAS GTPase-activating proteins (GAPs) that includes mammalian GAP, the yeast Saccharomyces cerevisiae IRA proteins, and the product of the human neurofibromatosis locus, NF1 sar1, like these other proteins, can complement the loss of IRA function in S. cerevisiae. Computer analysis shows that the highest degree of sequence conservation is restricted to a very small number of diagnostic residues represented by the motif Phe-Leu-Arg-X-X-X-Pro-Ala-X-X-X-Pro. We find no evidence that sar1 is required for the effector function of ras1.
Epidermal growth factor and transforming growth factor-alpha: differential intracellular routing and processing of ligand-receptor complexes.
Authors: Ebner R , Derynck R
Published: August 2013

Two structurally related but different polypeptide growth factors, epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha), exert their activities after interaction with a common cell-surface EGF/TGF-alpha-receptor. Comparative studies of the effects of both ligands have established that TGF-alpha is more potent than EGF in a variety of biological systems. This observation is not explained by differences in affinities of the ligands for the receptor, because the affinity-constants of both factors are very similar. We have compared the intracellular processing of ligand-receptor complexes using either EGF or TGF-alpha in two different cell systems. We found that TGF-alpha dissociates from the EGF/TGF-alpha-receptor at much higher pH than EGF, which may reflect the substantial difference in the calculated isoelectric points. After internalization, the intracellular TGF-alpha is more rapidly cleared than EGF, and a substantial portion of the released TGF-alpha represents undegraded TGF-alpha in contrast to the mostly degraded EGF. In addition, TGF-alpha did not induce a complete down-regulation of cell surface receptors, as observed with EGF, which is at least in part responsible for a much sooner recovery of the ligand-binding ability after down-regulation, in the case of TGF-alpha. These differences in processing of the ligand-receptor complexes may explain why TGF-alpha exerts quantitatively higher activities than EGF.
Degradation of T-cell receptor chains in the endoplasmic reticulum is inhibited by inhibitors of cysteine proteases.
Authors: Wileman T , Kane L P , Terhorst C
Published: August 2013

The endoplasmic reticulum, or an organelle closely associated with it, contains proteases that can be used to remove partially assembled or improperly folded proteins. Very little is known at present about the types of protease that degrade these proteins. The beta chain and cluster of differentiation (CD)3 delta subunit of the human T-cell antigen receptor (TCR) are degraded shortly after synthesis. In this study Chinese hamster ovary (CHO) cells transfected with either beta or delta were incubated with a panel of protease inhibitors, and the rates of degradation of the transfected proteins were followed using chain-specific enzyme-linked immunosorbent assays (ELISAs). Of the protease inhibitors tested, degradation of both chains was highly sensitive to sulfhydryl reagents and peptidyl inhibitors of cysteine proteases. Concentrations of inhibitors that produced near complete inhibition of degradation in the endoplasmic reticulum did not cause gross changes in cellular ATP levels nor did they significantly slow constitutive secretion from CHO cells. The inhibitors did not affect the ability of CHO cells to synthesize and assemble disulphide-linked TCR zeta dimers. We conclude that the protease inhibitors were not toxic to cells and did not affect the biosynthetic activity of the endoplasmic reticulum. Furthermore, they did not alter the ability of the endoplasmic reticulum to deliver its content to the Golgi apparatus. Taken together, these results suggest that the cysteine protease inhibitors slow degradation in the endoplasmic reticulum through an action on cysteine proteases. The results imply that the endoplasmic reticulum contains cysteine proteases that can be used to remove retained proteins.