bluuebunny/crossref_metadata_2025 huggingface dataset:
Peroxisome biogenesis disorders: The role of peroxisomes and metabolic dysfunction in developing brain
Authors: Faust P. L. , Banka D. , Siriratsivawong R. , Ng V. G. , Wikander T. M.
Published: April 2005

Peroxisome biogenesis disorders, of which Zellweger syndrome is the most severe, result in severe neurological dysfunction associated with abnormal CNS neuronal migrations due to the lack of functional peroxisomes. The PEX2‐/‐ mouse model for Zellweger syndrome has enabled us to evaluate the role of peroxisomes in the development and functioning of the nervous system. These studies have shown that, in addition to disturbances in neuronal migration in developing cerebral cortex and cerebellum, defects in neuronal differentiation, proliferation and survival may also contribute to the CNS malformations. However, owing to the multiorgan dysfunction in peroxisomal disorders, it has been difficult to clearly define an intrinsic role for the peroxisome in brain cells. The use of several in vitro cell culture assays to evaluate the migration and differentiation of cerebellar neurons demonstrates a persistence of defects in peroxisome‐deficient neurons. The absence of potential systemically derived, extrinsic factors in these in vitro systems indicates that CNS intrinsic defects contribute to the pathogenesis of disease in these disorders. However, bile acid treatment also increases the survival and growth of PEX2‐/‐ mice and improves some aspects of cerebellar development, indicating that extrinsic factors also affect the developing peroxisome‐deficient brain. Therefore, the final phenotype of nervous system dysfunction in peroxisomal disorders will reflect a combination of both CNS intrinsic and extrinsic factors.
Molecular mechanisms of dominant expression in porphyria
Authors: Badminton M. N. , Elder G. H.
Published: April 2005

Summary:#Partial deficiency of enzymes in the haem synthetic pathway gives rise to a group of seven inherited metabolic disorders, the porphyrias. Each deficiency is associated with a characteristic increase in haem precursors that correlates with the symptoms associated with individual porphyrias and allows accurate diagnosis. Two types of clinical presentation occur separately or in combination; acute life‐threatening neurovisceral attacks and/or cutaneous symptoms. Five of the porphyrias are low‐penetrance autosomal dominant conditions in which clinical expression results from additional factors that act by increasing demand for haem or by causing an additional decrease in enzyme activity or by a combination of these effects. These include both genetic and environmental factors. In familial porphyria cutanea tarda (PCTF), environmental factors that include alcohol, exogenous oestrogens and hepatotropic viruses result in inhibition of hepatic enzyme activity via a mechanism that involves excess iron accumulation. In erythropoietic protoporphyria (EPP), co‐inheritance of a functional polymorphism in trans to a null ferrochelatase allele accounts for most clinically overt cases. In the autosomal dominant acute hepatic porphyrias (acute intermittent porphyria, variegate porphyria, hereditary coproporphyria), acute neurovisceral attacks occur in a minority of those who inherit one of these disorders. Although various exogenous (e.g. drugs, alcohol) and endogenous factors (e.g. hormones) have been identified as provoking acute attacks, these do not provide a full explanation for the low penetrance of these disorders. It seems probable that genetic background influences susceptibility to acute attacks, but the genes that are involved have not yet been identified.
Exposure to threat of war and terror, political attitudes, stress, and life satisfaction among teenagers in Israel☆
Authors: Shamai Michal , Kimhi Shaul
Published: April 2005

The study examines the pattern of relations between level of threat, political attitude (willingness for political compromise), and well‐being (level of stress and life satisfaction) among Israeli teenagers in reaction to Israel's withdrawal from Lebanon. Participants were 419 teenagers from two residential areas: the Northern area (close to the Israeli–Lebanese border) and the Central area (distant from the border). The results show: (a) significant differences between the two groups on: political attitude (Center scored higher), on level of stress (North scored higher), and on life satisfaction (North scored lower); (b) a significant negative correlation between exposure to threat and political attitude; (c) significant negative correlations between political attitude and measures of stress, and a significant positive correlation with life satisfaction; (d) political attitude mediates between level of threat and well‐being. Political and educational implications, and mental health policy are discussed.
STORKS AND OSTRICHES: AN EARLY PARODY OF STRAUSS'S ‘ARIADNE AUF NAXOS’
Authors: Walton Chris
Published: April 2005

The name of the conductor Max Conrad is hardly one to conjure with. He admitted as much himself in the first paragraph of his memoirs, published in 1956:
To be sure, I was born in Berlin, as was [Bruno Walter]; he attended the Askanische High School in Berlin – as did I; he left it without finishing his school leaving certificate – as did I; he studied music with teachers from the Stern Conservatory – as did I; and he soon became famous – and I … oh dear, now it becomes irregular, like a Greek verb …

For more than 30 years, Conrad (1872–1963) played a major role in the operatic life of Zurich. In the greater scheme of things, this might perhaps be regarded as of little significance, for Zurich's was then but a theatre in the outer provinces of the German-speaking world. Nor did Conrad achieve recognition by conducting important premières there, as did other men (such as Robert F. Denzler, who conducted the world premières of Hindemith's Mathis der Maler and Berg's Lulu in the 1930s, or Hans Rosbaud, who conducted the staged première of Schoenberg's Moses und Aron in 1957). Conrad did train the flower maidens for the first (legal) performance of Wagner's Parsifal outside Bayreuth, given in Zurich in 1913 – but the première itself was conducted by the chief kapellmeister, Lothar Kempter. To be sure, Conrad did conduct the world première of the revised version of Othmar Schoeck's fine expressionist opera Penthesilea in 1928; but that is hardly enough to ensure anyone posthumous fame.
Silvestrov and Kancheli
Authors: Quinn Peter
Published: April 2005

SILVESTROV: Silent Songs; Four Songs after Osip Mandelstam. Sergey Yakovenko (bar), Ilya Scheps, Valentin Silvestrov (pno). ECM New Series 982 1424.

KANCHELI: Diplipito; Valse Boston. Thomas Demenga (vc), Derek Lee Ragin (counter-ten), Dennis Russell Davies (pno), Stuttgarter Kammerorchester c. Dennis Russell Davies. ECM New Series 472 0822.
An upper stratospheric layer of enhanced HNO3 following exceptional solar storms
Authors: Orsolini Y. J. , Manney G. L. , Santee M. L. , Randall C. E.
Published: April 2005

An analysis of stratospheric nitric acid (HNO3) observed by the Michelson Interferometer for Passive Atmospheric Sounding (MIPAS) reveals a distinct, high‐altitude maximum, that appeared in late November 2003 in the polar upper stratosphere. Confined to the polar vortex, the enhanced HNO3 layer intensified while descending to the middle stratosphere, and disappeared between mid‐January and mid‐February. The high‐altitude maximum is considerably enhanced compared to the weak, secondary maxima previously reported in the literature. Analysis of MIPAS stratospheric nitrogen dioxide (NO2) and correlations with the geomagnetic Ap index suggest that particle precipitation from the intense solar storms of October–November 2003 was responsible for this extraordinary high‐altitude HNO3 layer. The detailed morphology and time‐evolution of such a distinct, high‐altitude HNO3 layer have not been observed before, nor the link to geomagnetic activity through contemporaneous upper stratospheric NO2 measurements.
The Aksay segment of the northern Altyn Tagh fault: Tectonic geomorphology, landscape evolution, and Holocene slip rate
Authors: Mériaux A.‐S. , Tapponnier P. , Ryerson F. J. , Xiwei Xu , King G. , Van der Woerd J. , Finkel R. C. , Haibing Li , Caffee M. W. , Zhiqin Xu , Wenbin Chen
Published: April 2005

Millennial slip rates have been determined for the Altyn Tagh fault (ATF) at three sites near Aksay (∼94°E) in northeastern Tibet by dating fluvial channels and terrace riser offsets with radiocarbon and 10Be‐26Al surface exposure dating. Up to nine main surfaces are defined on the basis of morphology, elevation, and dating. The abandonment age of some surfaces is constrained by radiocarbon dating, which typically coincides with the youngest cosmogenic ages for a particular surface. Older surface exposure ages are taken to represent the duration of terrace emplacement. Cumulative offsets range from 20 to 260 m and fall in distinct groups, indicative of climatically modulated regional landscape formation. Most samples are younger than ∼14 ka and postdate the Last Glacial Maximum. The end of the early Holocene optimum marks the boundary between the ages of the two main terrace levels at 5–6 ka. At this longitude the ATF is divided into a northern and southern branch. The northern ATF should thus yield a minimum rate for the ATF system. Slip rate estimates using the abandonment age of the overlying level for fill terraces or channels and the emplacement of the underlying level for strath terraces give 30 consistent results, yielding an average Holocene rate of 17.8 ± 3.6 mm/yr. It is ∼9 mm/yr less than the long‐term rate obtained near Tura at ∼87°E (26.9 ± 6.9 mm/yr), in keeping with the inference of an eastward decreasing rate on the ATF, due to increased thrusting to the south. However, it remains twice the rate determined by GPS studies.
Numerical study on the Kuroshio path states in the northern Okinawa Trough of the East China Sea
Authors: Nakamura Hirohiko
Published: April 2005

Previous observations have indicated that the Kuroshio in the northern Okinawa Trough is quasibimodal in character, with distinct northern and southern path states. The Kuroshio path states are examined here through simulations using a primitive inflow/outflow model for 16 inflow structures parameterized by the location of maximum surface velocity and the rate of velocity decrease with depth. The simulations reveal three distinct path states in the northern Okinawa Trough: a weakly stable northern path state associated with blocking of an anticyclonic Kuroshio meander, a highly stable southern path state maintained by blocking of a cyclonic Kuroshio meander, and an intermediate unstable path state. The variations in the unstable state, occurring as either periodic transitions between the northern and southern path states or periodic fluctuations of an otherwise stable path, are driven by self‐sustained internal oscillation in the northern Okinawa Trough. Cyclogenesis in the continental slope area due to baroclinic instability appears to play an important role in Kuroshio path variations in the unstable state, and this instability is effectively controlled by the location of the Kuroshio mean path on the slope rather than its velocity as conventionally thought.
Seasonal heat budget in the mixed layer of the southeastern tropical Indian Ocean in a high‐resolution ocean general circulation model
Authors: Du Yan , Qu Tangdong , Meyers Gary , Masumoto Yukio , Sasaki Hideharu
Published: April 2005

Using results from a high‐resolution ocean general circulation model, this study identifies the impact of ocean dynamics on the mean seasonal cycle of sea surface temperature (SST) in the southeastern tropical Indian Ocean. An important question about the mechanisms of regional SST variation arises in a quick look at satellite‐observed SST and ocean color. Although ocean color indicates a distinctive upwelling season from July to September, the SST depression off Java and Sumatra is small, forming a sharp contrast with other eastern boundary regions. The model results suggest an explanation of the process responsible for the small SST depression. Our analysis indicates that there are three dynamically different regimes within the region. First, near the coast of northwest Australia, surface heat flux controls the seasonal variation of SST, while horizontal advection and vertical entrainment are relatively weak. This result is consistent with previous studies. Second, south of Java and farther to the east, warm horizontal advection of the Indonesian throughflow (ITF) neutralizes the cold upwelling. The transport of the ITF, especially the outflow from the Lombok Strait, reaches its seasonal maximum in July–September, at the same time that the maximum upwelling occurs. Third, west of Sumatra, a thick barrier layer exists, which impedes the cold thermocline water from entering the mixed layer (ML). Although upwelling occurs, it has no significant effect on the SST as a result of the small vertical temperature gradient at the bottom of the ML.
Underthrusting and exhumation: A comparison between the External Hellenides and the “hot” Cycladic and “cold” South Aegean core complexes (Greece)
Authors: van Hinsbergen D. J. J. , Zachariasse W. J. , Wortel M. J. R. , Meulenkamp J. E.
Published: April 2005

After their emplacement in the course of the late Mesozoic and the Cenozoic, the Hellenic nappes became fragmented during late orogenic extension since the late Eocene. Here we focus on the transition of underthrusting during nappe emplacement to exhumation during late orogenic extension. To this end, we compared previously published data on the structural geological and metamorphic history of the underthrusted parts of the Tripolitza and Ionian nappes, which were exhumed in the Cycladic and South Aegean windows, with newly obtained data on the sedimentary, stratigraphic, and structural development of the part of these nappes in the foreland, in front of the subduction thrust. The results allow the identification of two major events: Event 1 took place around the Eocene‐Oligocene transition and marks the onset of underthrusting of the Tripolitza nappe below the Pindos nappe and the Ionian nappe below the Tripolitza nappe, respectively. This led to the uplift and erosion of the Pindos unit and the onset of deposition of the Tripolitza and Ionian flysch in front of the Pindos thrust, together with the formation of mylonites at the base of the metamorphosed portions of the Pindos unit related to the underthrusting of the Tripolitza unit. Event 2 occurred in the latest Oligocene to earliest Miocene and marks the decoupling of the Ionian unit from the underthrusting plate, the accretion of the Tripolitza and Ionian units to the overriding plate, and the onset of late orogenic extension and exhumation in the overriding plate. This led to the formation of the South Aegean and Cycladic core complexes and the subsidence of the Klematia‐Paramythia half‐graben throughout the early Miocene.
Sprouty-4 negatively regulates cell spreading by inhibiting the kinase activity of testicular protein kinase
Authors: TSUMURA Yoshikazu , TOSHIMA Jiro , LEEKSMA Onno C. , OHASHI Kazumasa , MIZUNO Kensaku
Published: April 2005

TESK1 (testicular protein kinase 1) is a serine/threonine kinase that phosphorylates cofilin and plays a critical role in integrin-mediated actin cytoskeletal reorganization and cell spreading. We previously showed that TESK1 interacts with Sprouty-4 (referred to as Spry4), an inhibitor of growth factor-induced Ras/MAP (mitogen-activated protein) kinase signalling, but the functional role of this interaction has remained unknown. In the present study, we show that Spry4 inhibits the kinase activity of TESK1 by binding to it through the C-terminal cysteine-rich region. Expression of Spry4 in cultured cells suppressed integrin-mediated cell spreading, and TESK1 reversed the inhibitory effect of Spry4 on cell spreading. Furthermore, Spry4 suppressed integrin- and TESK1-mediated cofilin phosphorylation during the spreading of cells on laminin. These findings suggest that Spry4 suppresses cell spreading by inhibiting the kinase activity of TESK1. Although tyrosine phosphorylation is required for the inhibitory activity of Spry4 on a Ras/MAP kinase pathway, mutation of the corresponding tyrosine residue (Tyr-75 in human Spry4) to an alanine had no apparent effect on its inhibitory actions on TESK1 activity and cell spreading, which suggests a novel cellular function of Spry to regulate the actin cytoskeleton, independent of its inhibitory activity on the Ras/MAP kinase signalling.
Ypr140wp, ‘the yeast tafazzin’, displays a mitochondrial lysophosphatidylcholine (lyso-PC) acyltransferase activity related to triacylglycerol and mitochondrial lipid synthesis
Authors: TESTET Eric , LAROCHE-TRAINEAU Jeanny , NOUBHANI Abdelmajid , COULON Denis , BUNOUST Odile , CAMOUGRAND Nadine , MANON Stephen , LESSIRE René , BESSOULE Jean-Jacques
Published: April 2005

When the yeast protein Ypr140w was expressed in Escherichia coli, a lyso-PC [lysophosphatidylcholine (1-acylglycerophosphorylcholine)] acyltransferase activity was found associated with the membranes of the bacteria. To our knowledge, this is the first identification of a protein capable of catalysing the acylation of lyso-PC molecules to form PC. Fluorescence microscopy analysis of living yeasts revealed that the fusion protein Ypr140w–green fluorescent protein is targeted to the mitochondria. Moreover, in contrast with wild-type cells, in the absence of acyl-CoA, the yeast mutant deleted for the YPR140w gene has no lyso-PC acyltransferase activity associated with the mitochondrial fraction. When yeast cells were grown in the presence of lactate, the mutant synthesized 2-fold more triacylglycerols when compared with the wild-type. Moreover, its mitochondrial membranes contained a lesser amount of PC and cardiolipin, and the fatty acid composition of these latter was greatly changed. These modifications were accompanied by a 2-fold increase in the respiration rates (states 3 and 4) of the mitochondria. The relationship between the deletion of the YPR140w gene and the lipid composition of the ypr140wΔ cells is discussed.
Orientation and mode of lipid-binding interaction of human apolipoprotein E C-terminal domain
Authors: RAUSSENS Vincent , DRURY Jessica , FORTE Trudy M. , CHOY Nicole , GOORMAGHTIGH Erik , RUYSSCHAERT Jean-Marie , NARAYANASWAMI Vasanthy
Published: April 2005

ApoE (apolipoprotein E) is an anti-atherogenic lipid transport protein that plays an integral role in lipoprotein metabolism and cholesterol homoeostasis. Lipid association educes critical functional features of apoE, mediating reduction in plasma and cellular cholesterol levels. The 10-kDa CT (C-terminal) domain of apoE facilitates helix–helix interactions in lipid-free state to promote apoE self-association and helix–lipid interactions during binding with lipoproteins, although the mode of lipid-binding interaction is not well understood. We investigated the mode of lipid-binding interaction and orientation of apoE CT domain on reconstituted lipoproteins. Isolated recombinant human apoE CT domain (residues 201–299) possesses a strong ability to interact with phospholipid vesicles, yielding lipoprotein particles with an apparent molecular mass of ∼600 kDa, while retaining the overall α-helical content. Electron microscopy and non-denaturing PAGE analysis of DMPC (dimyristoylphosphatidylcholine)–apoE CT domain lipoprotein complexes revealed discoidal complexes with a diameter of approx. 17 nm. Cross-linking apoE CT domain on discoidal particles yielded dimeric species as the major product. Attenuated total reflectance Fourier transform IR spectroscopy of phospholipid–apoE CT domain complexes reveals that the helical axis is oriented perpendicular to fatty acyl chains of the phospholipid. Fluorescence quenching analysis of DMPC–apoE CT domain discoidal complexes by spin-labelled stearic acid indicated a relatively superficial location of the native tryptophan residues with respect to the plane of the phospholipid bilayer. Taken together, we propose that apoE CT domain interacts with phospholipid vesicles, forming a long extended helix that circumscribes the discoidal bilayer lipoprotein complex.
Characterization of an archaeal family 4 uracil DNA glycosylase and its interaction with PCNA and chromatin proteins
Authors: DIONNE Isabelle , BELL Stephen D.
Published: April 2005

We describe the characterization of a family 4 UDG1 (uracil DNA glycosylase) from the crenarchaeote Sulfolobus solfataricus. UDG1 is found to have a marked preference for substrates containing a G:U base pair over either A:U or single-stranded uracil-containing DNA substrates. UDG1 is found to interact with the sliding clamp PCNA (proliferating cell nuclear antigen), and does so by a conserved motif in the C-terminus of the protein. S. solfataricus has a heterotrimeric PCNA, and only one of the subunits, PCNA3, interacts with UDG1. We have been unable to detect any stimulation of UDG activity by PCNA, in contrast with the observed effects of PCNA on a number of DNA metabolic enzymes. However, analysis of the effects of Sulfolobus chromatin proteins on UDG1 leads us to propose a mechanistic basis for coupling UDG1 to the replication fork.
Inhibitory effect of palmitate on the mitochondrial NADH:ubiquinone oxidoreductase (complex I) as related to the active–de-active enzyme transition
Authors: LOSKOVICH Maria V. , GRIVENNIKOVA Vera G. , CECCHINI Gary , VINOGRADOV Andrei D.
Published: April 2005

Palmitate rapidly and reversibly inhibits the uncoupled NADH oxidase activity catalysed by activated complex I in inside-out bovine heart submitochondrial particles (IC50 extrapolated to zero enzyme concentration is equal to 9 μM at 25 °C, pH 8.0). The NADH:hexa-ammineruthenium reductase activity of complex I is insensitive to palmitate. Partial (∼50%) inhibition of the NADH:external quinone reductase activity is seen at saturating palmitate concentration and the residual activity is fully sensitive to piericidin. The uncoupled succinate oxidase activity is considerably less sensitive to palmitate. Only a slight stimulation of tightly coupled respiration with NADH as the substrate is seen at optimal palmitate concentrations, whereas complete relief of the respiratory control is observed with succinate as the substrate. Palmitate prevents the turnover-induced activation of the de-activated complex I (IC50 extrapolated to zero enzyme concentration is equal to 3 μM at 25 °C, pH 8.0). The mode of action of palmitate on the NADH oxidase is qualitatively temperature-dependent. Rapid and reversible inhibition of the complex I catalytic activity and its de-active to active state transition are seen at 25 °C, whereas the time-dependent irreversible inactivation of the NADH oxidase proceeds at 37 °C. Palmitate drastically increases the rate of spontaneous de-activation of complex I in the absence of NADH. Taken together, these results suggest that free fatty acids act as specific complex I-directed inhibitors; at a physiologically relevant temperature (37 °C), their inhibitory effects on mitochondrial NADH oxidation is due to perturbation of the pseudo-reversible active–de-active complex I transition.
Chaperone-like properties of the prodomain of TNFα-converting enzyme (TACE) and the functional role of its cysteine switch
Authors: LEONARD Jennifer D. , LIN Frank , MILLA Marcos E.
Published: April 2005

The prodomain of TACE [TNFα (tumour necrosis factor α)-converting enzyme] is essential for the secretion of the functional enzyme. Previously, we showed that a TACE truncate was not secreted in the absence of the prodomain and that it was subjected to intracellular degradation. In the present study, we show that full-length TACE was also degraded when expressed without the prodomain. We demonstrate that the prodomain can rescue TACE's secretion in trans, suggesting an intramolecular chaperone function. We addressed the question whether a cysteine switch consensus motif is needed for the secretion of active TACE. The cysteine switch mutants [C184A (Cys184→Ala)] of TACE resembled the wild-type functionally and in their sensitivity to inhibitors. Interestingly, TACE zymogen forms expressed in the context of the C184A mutation were susceptible to intracellular degradation, suggesting that the prodomain-bound TACE zymogen may be more accessible to intracellular proteinases when compared with mature TACE. Two independent findings confirmed that the catalytic domain of TACE is in a more open state when bound to its prodomain: (i) core tryptophan residues were exposed to the solvent in the procatalytic domain complex and (ii) LysC rapidly proteolysed the procatalytic domain complex but not mature TACE. Therefore the prodomain of TACE is a specific intramolecular chaperone that aids in the secretion of this enzyme, while keeping the catalytic domain in a relatively open conformation. The cysteine switch of TACE is not essential for the secretion of the functional enzyme, but may prevent intracellular degradation of the TACE zymogen.
Mutational analysis of the active-site residues crucial for catalytic activity of adenosine kinase from Leishmania donovani
Authors: DATTA Rupak , DAS Ishita , SEN Banibrata , CHAKRABORTY Anutosh , ADAK Subrata , MANDAL Chhabinath , DATTA Alok K.
Published: April 2005

Leishmania donovani adenosine kinase (LdAdK) plays a pivotal role in scavenging of purines from the host. Exploiting interspecies homology and structural co-ordinates of the enzyme from other sources, we generated a model of LdAdK that led us to target several amino acid residues (namely Gly-62, Arg-69, Arg-131 and Asp-299). Replacement of Gly-62 with aspartate caused a drastic reduction in catalytic activity, with decreased affinity for either substrate. Asp-299 was found to be catalytically indispensable. Mutation of either Arg-131 or Arg-69 caused a significant reduction in kcat. R69A (Arg-69→Ala) and R131A mutants exhibited unaltered Km for either substrate, whereas ATP Km for R69K increased 6-fold. Importance of both of the arginine residues was reaffirmed by the R69K/R131A double mutant, which exhibited approx. 0.5% residual activity with a large increase in ATP Km. Phenylglyoxal, which inhibits the wild-type enzyme, also inactivated the arginine mutants to different extents. Adenosine protected both of the Arg-69 mutants, but not the R131A variant, from inactivation. Binding experiments revealed that the AMP-binding property of R69K or R69A and D299A mutants remained largely unaltered, but R131A and R69K/R131A mutants lost their AMP binding ability significantly. The G62D mutant did not bind AMP at all. Free energy calculations indicated that Arg-69 and Arg-131 are functionally independent. Thus, apart from the mandatory requirement of flexibility around the diglycyl (Gly-61–Gly-62) motif, our results identified Asp-299 and Arg-131 as key catalytic residues, with the former functioning as the proton abstractor from the 5′-OH of adenosine, while the latter acts as a bidentate electrophile to stabilize the negative charge on the leaving group during the phosphate transfer. Moreover, the positive charge distribution of Arg-69 probably helps in maintaining the flexibility of the α-3 helix needed for proper domain movement. These findings provide the first comprehensive biochemical evidence implicating the mechanistic roles of the functionally important residues of this chemotherapeutically exploitable enzyme.
Regulation of the prolyl hydroxylase domain protein 2 (phd2/egln-1) gene: identification of a functional hypoxia-responsive element
Authors: METZEN Eric , STIEHL Daniel P. , DOEGE Kathrin , MARXSEN Jan H. , HELLWIG-BÜRGEL Thomas , JELKMANN Wolfgang
Published: April 2005

The HIFs (hypoxia-inducible factors) are a family of heterodimeric transcription factors essential for the adaptation of cells to reduced oxygen supply. Three human PHDs (prolyl hydroxylase domain proteins, PHD1–PHD3) initiate oxygen-dependent degradation of HIF-α-subunits in normoxia. RNA interference directed against PHD2, but not PHD1 or PHD3, is sufficient to stabilize HIF-1α in normoxia. Therefore PHD2 is regarded as the main cellular oxygen sensor. PHD2 itself is up-regulated by hypoxia and may thus limit hypoxic signalling. By sequence analysis, we predicted a promoter approx. 3.5 kb 5′ of the translation start codon and a second promoter located in a CpG island immediately upstream of the coding sequence. A consensus HIF-1-binding site that is conserved in the murine phd2 gene was detected in the CpG island. By electrophoretic mobility-shift assay, we demonstrated binding of HIF-1 to the putative HIF-1-binding site. In luciferase reporter vectors, the isolated upstream promoter was inactive in all cell lines tested unless 200 bp were deleted at the 3′-end. The downstream promoter was active and induced by hypoxia. In reporter vectors containing both promoter sequences, luciferase activity was equal to vectors containing only the downstream promoter. In cells transfected with a vector containing both promoters, a single luciferase transcript was detectable. This transcript had the same length as transcripts from a vector containing the downstream promoter only. We conclude that the phd2 gene is transcribed exclusively from the downstream promoter that contains a functional hypoxia-responsive, cis-regulatory element. Our results establish that PHD2 is a direct HIF target gene.
The sustainability of interactions between the orexin-1 receptor and β-arrestin-2 is defined by a single C-terminal cluster of hydroxy amino acids and modulates the kinetics of ERK MAPK regulation
Authors: MILASTA Sandra , EVANS Nicholas A. , ORMISTON Laura , WILSON Shelagh , LEFKOWITZ Robert J. , MILLIGAN Graeme
Published: April 2005

The orexin-1 receptor interacts with β-arrestin-2 in an agonist-dependent manner. In HEK-293T cells, these two proteins became co-internalized into acidic endosomes. Truncations from the C-terminal tail did not prevent agonist-induced internalization of the orexin-1 receptor or alter the pathway of internalization, although such mutants failed to interact with β-arrestin-2 in a sustained manner or produce its co-internalization. Mutation of a cluster of three threonine and one serine residue at the extreme C-terminus of the receptor greatly reduced interaction and abolished co-internalization of β-arrestin-2–GFP (green fluorescent protein). Despite the weak interactions of this C-terminally mutated form of the receptor with β-arrestin-2, studies in wild-type and β-arrestin-deficient mouse embryo fibroblasts confirmed that agonist-induced internalization of this mutant required expression of a β-arrestin. Although without effect on agonist-mediated elevation of intracellular Ca2+ levels, the C-terminally mutated form of the orexin-1 receptor was unable to sustain phosphorylation of the MAPKs (mitogen-activated protein kinases) ERK1 and ERK2 (extracellular-signal-regulated kinases 1 and 2) to the same extent as the wild-type receptor. These studies indicate that a single cluster of hydroxy amino acids within the C-terminal seven amino acids of the orexin-1 receptor determine the sustainability of interaction with β-arrestin-2, and indicate an important role of β-arrestin scaffolding in defining the kinetics of orexin-1 receptor-mediated ERK MAPK activation.
Purification of a 75 kDa protein from the organelle matrix of human neutrophils and identification asN-acetylglucosamine-6-sulphatase
Authors: XU Shengyuan , ZHAO Linshu , LARSSON Anders , SMEDS Emanuel , KUSCHE-GULLBERG Marion , VENGE Per
Published: April 2005

A 75 kDa protein was purified to homogeneity from granule extracts of normal human granulocytes using Sephadex G-75 chromatography, Mono-S cation exchange chromatography and chromatofocusing. The protein consisted of one chain with a molecular mass of 75 kDa, as determined by SDS/PAGE. Tryptic peptide analysis by MALDI-TOF (matrix-assisted laser-desorption ionization–time-of-flight) MS and sequence analysis by MS/MS identified the protein to be N-acetylglucosamine-6-sulphatase (EC 3.1.6.14). The identity of the protein was confirmed by demostrating enzymatic activity towards the substrate N-acetylglucosamine 6-sulphate. The enzyme was active over a broad pH range with an optimum of pH 7.0, and showed a Km value of 13.0 mM and a Vmax value of ∼1.8 μM/min per mg. The enzyme also showed O-desulphation activity towards heparan sulphate-derived saccharides. Subcellular fractionation of neutrophil organelles showed the presence of enzymatic activity mainly in the same fractions as primary granules. Furthermore, PMA treatment of the neutrophils induced release of the enzyme, indicating its matrix protein nature. The presence of N-acetylglucosamine-6-sulphatase in human neutrophils implies that neutrophils may play a role in the modulation of cell surface molecules and extracellular matrix by O-desulphation.
Interaction of three-finger toxins with phospholipid membranes: comparison of S- and P-type cytotoxins
Authors: DUBOVSKII Peter V. , LESOVOY Dmitry M. , DUBINNYI Maxim A. , KONSHINA Anastasiya G. , UTKIN Yuri N. , EFREMOV Roman G. , ARSENIEV Alexander S.
Published: April 2005

The CTs (cytotoxins) I and II are positively charged three-finger folded proteins from venom of Naja oxiana (the Central Asian cobra). They belong to S- and P-type respectively based on Ser-28 and Pro-30 residues within a putative phospholipid bilayer binding site. Previously, we investigated the interaction of CTII with multilamellar liposomes of dipalmitoylphosphatidylglycerol by wide-line 31P-NMR spectroscopy. To compare interactions of these proteins with phospholipids, we investigated the interaction of CTI with the multilamellar liposomes of dipalmitoylphosphatidylglycerol analogously. The effect of CTI on the chemical shielding anisotropy and deformation of the liposomes in the magnetic field was determined at different temperatures and lipid/protein ratios. It was found that both the proteins do not affect lipid organization in the gel state. In the liquid crystalline state of the bilayer they disturb lipid packing. To get insight into the interactions of the toxins with membranes, Monte Carlo simulations of CTI and CTII in the presence of the bilayer membrane were performed. It was found that both the toxins penetrate into the bilayer with the tips of all the three loops. However, the free-energy gain on membrane insertion of CTI is smaller (by ≈7 kcal/mol; 1 kcal≡4.184 kJ) when compared with CTII, because of the lower hydrophobicity of the membrane-binding site of CTI. These results clearly demonstrate that the P-type cytotoxins interact with membranes stronger than those of the S-type, although the mode of the membrane insertion is similar for both the types.
Identification of a unique filamin A binding region within the cytoplasmic domain of glycoprotein Ibα
Authors: CRANMER Susan L. , PIKOVSKI Inna , MANGIN Pierre , THOMPSON Philip E. , DOMAGALA Teresa , FRAZZETTO Mark , SALEM Hatem H. , JACKSON Shaun P.
Published: April 2005

Binding of the platelet GPIb/V/IX (glycoprotein Ib/V/IX) receptor to von Willebrand factor is critical for platelet adhesion and aggregation under conditions of rapid blood flow. The adhesive function of GPIbα is regulated by its anchorage to the membrane skeleton through a specific interaction with filamin A. In the present study, we examined the amino acid residues within the cytoplasmic tail of GPIbα, which are critical for association with filamin A, using a series of 25-mer synthetic peptides that mimic the cytoplasmic tail sequences of wild-type and mutant forms of GPIbα. Peptide binding studies of purified human filamin A have demonstrated a major role for the conserved hydrophobic stretch L567FLWV571 in mediating this interaction. Progressive alanine substitutions of triple, double and single amino acid residues within the Pro561–Arg572 region suggested an important role for Trp570 and Phe568 in promoting GPIbα binding to filamin A. The importance of these two residues in promoting filamin A binding to GPIbα in vivo was confirmed from the study of Chinese-hamster ovary cells expressing GPIbα Trp570→Ala and Phe568→Ala substitutions. Phenotypic analysis of these cell lines in flow-based adhesion studies revealed a critical role for these residues in maintaining receptor anchorage to the membrane skeleton and in maintaining cell adhesion to a von Willebrand factor matrix under high-shear conditions. These studies demonstrate a novel filamin A binding motif in the cytoplasmic tail of GPIbα, which is critically dependent on both Trp570 and Phe568.
Identification and assessment of the role of a nominal phospholipid binding region of ORP1S (oxysterol-binding-protein-related protein 1 short) in the regulation of vesicular transport
Authors: FAIRN Gregory D. , McMASTER Christopher R.
Published: April 2005

The ORPs (oxysterol-binding-protein-related proteins) constitute an enigmatic family of intracellular lipid receptors that are related through a shared lipid binding domain. Emerging evidence suggests that ORPs relate lipid metabolism to membrane transport. Current data imply that the yeast ORP Kes1p is a negative regulator of Golgi-derived vesicular transport mediated by the essential phosphatidylinositol/phosphatidylcholine transfer protein Sec14p. Inactivation of Kes1p function allows restoration of growth and vesicular transport in cells lacking Sec14p function, and Kes1p function in this regard can be complemented by human ORP1S (ORP1 short). Recent studies have determined that Kes1p and ORP1S both bind phospholipids as ligands. To explore the function of distinct linear segments of ORP1S in phospholipid binding and vesicular transport regulation, we generated a series of 15 open reading frames coding for diagnostic regions within ORP1S. Purified versions of these ORP1S deletion proteins were characterized in vitro, and allowed the identification of a nominal phospholipid binding region. The in vitro analysis was interpreted in the context of in vivo growth and vesicle transport assays for members of the ORP1S deletion set. The results determined that the phospholipid binding domain per se was insufficient for inhibition of vesicular transport by ORP1S, and that transport of carboxypeptidase Y and invertase from the Golgi may be regulated differentially by specific regions of ORP1S/Kes1p.
Orientational dynamics of water confined on a nanometer length scale in reverse micelles
Authors: Tan Howe-Siang , Piletic Ivan R. , Fayer M. D.
Published: April 2005

The time-resolved orientational anisotropies of the OD hydroxyl stretch of dilute HOD in H2O confined on a nanometer length scale in sodium bis(2-ethylhexyl) sulfosuccinate (AOT) reverse micelles are studied using ultrafast infrared polarization and spectrally resolved pump-probe spectroscopy, and the results are compared to the same experiments on bulk water. The orientational anisotropy data for three water nanopool sizes (4.0, 2.4, and 1.7nm) can be fitted well with biexponential decays. The biexponential decays are analyzed using a wobbling-in-a-cone model that involves fast orientational diffusion within a cone followed by slower, full orientational relaxation. The data provide the cone angles, the diffusion constants for motion within the cones, and the final diffusion constants as a function of the nanopool size. The two processes can be interpreted as a local angular fluctuation of the OD and a global hydrogen bond network rearrangement process. The trend in the relative amplitudes of the long and short exponential decays suggest an increasing rigidity as the nanopool size decreases. The trend in the long decay constants indicates a longer hydrogen bond network rearrangement time with decreasing reverse micelle size. The anisotropy measurements for the reverse micelles studied extrapolate to ∼0.33 rather than the ideal value of 0.4, suggesting the presence of an initial inertial component in the anisotropy decay that is too fast to resolve. The very fast decay component is consistent with initial inertial orientational motion that is seen in published molecular-dynamics simulations of water in AOT reverse micelles. The angle over which the inertial orientational motion occurs is determined. The results are in semiquantitative agreement with the molecular-dynamics simulations.
“Crystal” magnetic structure in axisymmetric plasma accretion disks
Authors: Coppi B.
Published: April 2005

A general class of stationary magnetic configurations which can exist in (thin) plasma accretion disks is identified by solving analytically the coupled nonlinear equations that describe the radial and the vertical equilibrium conditions of the disk. These configurations are characterized by a “crystal” structure consisting of a sequence of toroidal current filaments that can involve null points of the poloidal magnetic field. The obtained solutions are valid in the limit where the magnetic energy density is smaller than the thermal energy density (β>1). In view of studying magnetic disk configurations from which jets can emerge, and for which the limit where β∼1 is important, the relevant equilibrium equations are derived and their symmetries are pointed out.
A random walk through the dynamics of homogeneous vapor-liquid nucleation
Authors: Huang David M. , Attard Phil
Published: April 2005

A method of calculating rates of homogeneous vapor-liquid nucleation based on Langevin dynamics of a few relevant degrees of freedom on a free-energy surface is proposed. The surface is obtained here from simulation and from a semi empirical expression. The mass and friction coefficients are derived from atomistic umbrella-sampling molecular-dynamics simulations. The calculated nucleation rate agrees with atomistic simulations for one particular state point of the Lennard-Jones fluid. The present method is about four orders of magnitude more computationally efficient than the direct atomistic simulation of the transmission coefficient.
Marshall Rosenbluth and the Metropolis algorithm
Authors: Gubernatis J. E.
Published: April 2005

The 1953 publication, “Equation of State Calculations by Very Fast Computing Machines” by N. Metropolis, A. W. Rosenbluth and M. N. Rosenbluth, and M. Teller and E. Teller [J. Chem. Phys. 21, 1087 (1953)] marked the beginning of the use of the Monte Carlo method for solving problems in the physical sciences. The method described in this publication subsequently became known as the Metropolis algorithm, undoubtedly the most famous and most widely used Monte Carlo algorithm ever published. As none of the authors made subsequent use of the algorithm, they became unknown to the large simulation physics community that grew from this publication and their roles in its development became the subject of mystery and legend. At a conference marking the 50th anniversary of the 1953 publication, Marshall Rosenbluth gave his recollections of the algorithm’s development. The present paper describes the algorithm, reconstructs the historical context in which it was developed, and summarizes Marshall’s recollections.
Edge localized mode control with an edge resonant magnetic perturbation
Authors: Moyer R. A. , Evans T. E. , Osborne T. H. , Thomas P. R. , Becoulet M. , Harris J. , Finken K.-H. , Boedo J. A. , Doyle E. J. , Fenstermacher M. E. , Gohil P. , Groebner R. J. , Groth M. , Jackson G. L. , La Haye R. J. , Lasnier C. J. , Leonard A. W. , McKee G. R. , Reimerdes H. , Rhodes T. L. , Rudakov D. L. , Schaffer M. J. , Snyder P. B. , Wade M. R. , Wang G. , Watkins J. G. , West W. P. , Zeng L.
Published: April 2005

A low amplitude (δbr∕BT=1 part in 5000) edge resonant magnetic field perturbation with toroidal mode number n=3 and poloidal mode numbers between 8 and 15 has been used to suppress most large type I edge localized modes (ELMs) without degrading core plasma confinement. ELMs have been suppressed for periods of up to 8.6 energy confinement times when the edge safety factor q95 is between 3.5 and 4. The large ELMs are replaced by packets of events (possibly type II ELMs) with small amplitude, narrow radial extent, and a higher level of magnetic field and density fluctuations, creating a duty cycle with long “active” intervals of high transport and short “quiet” intervals of low transport. The increased transport associated with these events is less impulsive and slows the recovery of the pedestal profiles to the values reached just before the large ELMs without the n=3 perturbation. Changing the toroidal phase of the perturbation by 60° with respect to the best ELM suppression case reduces the ELM amplitude and frequency by factors of 2–3 in the divertor, produces a more stochastic response in the H-mode pedestal profiles, and displays similar increases in small scale events, although significant numbers of large ELMs survive. In contrast to the best ELM suppression case where the type I ELMs are also suppressed on the outboard midplane, the midplane recycling increases until individual ELMs are no longer discernable. The ELM response depends on the toroidal phase of the applied perturbation because intrinsic error fields make the target plasma nonaxisymmetric, and suggests that at least some of the variation in ELM behavior in a single device or among different devices is due to differences in the intrinsic error fields in these devices. These results indicate that ELMs can be suppressed by small edge resonant magnetic field perturbations. Extrapolation to next-step burning plasma devices will require extending the regime of operation to lower collisionality and understanding the physical mechanism responsible for the ELM suppression.
Structure and electrical properties of sol-gel-derived (001)-oriented Pb[Yb1∕2Nb1∕2]O3–PbTiO3 thin films grown on LaNbO3∕Si(001) substrates
Authors: Zhou Q. F. , Shung K. K. , Zhang Q. Q. , Djuth F. T.
Published: April 2005

( 1 − x ) Pb [ Yb 1 ∕ 2 Nb 1 ∕ 2 ] O 3 – x Pb Ti O 3 (PYbN–PT, x=0.5) oriented thin films were deposited onto LaNiO3 (LNO)∕Si substrates by sol-gel processing. Highly (001)-oriented LNO thin films were prepared by a simple metal-organic decomposition technique when sample annealed at 650°C using a rapid thermal annealing. The room-temperature resistivity of LNO thin films was 0.65mΩcm. X-ray diffraction analysis revealed that the films of PYbN–PT were highly (001) oriented along LNO∕Si substrates. No pyrochlore phase was observed by X-ray diffraction and films had a uniform grain size of about 80–120nm. At 1kHz, the dielectric permittivity was 920 and dielectric loss is about 0.035. The ferroelectric films displayed good P-E hysteresis characteristic and better temperature stabilization compared with the films that have low Curie temperature.
Electrically tunable polymer stabilized liquid-crystal lens
Authors: Presnyakov Vladimir V. , Galstian Tigran V.
Published: April 2005

A tunable focal lens using flat electro-optical liquid-crystal cell with uniform pixel-free electrodes is developed. The lenslike gradient refractive index profile is created in the cell via the spatially distributed polymer network obtained by photopolymerization using a spatially nonuniform laser beam. The conditions of the polymer network generation are optimized to improve the optical quality of the lens and its focusing properties. Low optical loss (scattering) is achieved for a focal length smoothly tunable from infinity to 0.8m. Obtained results can be applied to develop lenses that have no moving parts and allow the electro-optical zooming.
Characterization of extreme ultraviolet emission using the fourth harmonic of a Nd:YAG laser
Authors: Yamaura Michiteru , Uchida Shigeaki , Sunahara Atsushi , Shimada Yoshinori , Nishimura Hiroaki , Fujioka Shinsuke , Okuno Tomoharu , Hashimoto Kazuhisa , Nagai Keiji , Norimatsu Takayoshi , Nishihara Katsunobu , Miyanga Noriaki , Izawa Yasukazu , Yamanaka Chiyoe
Published: April 2005

Characterization of an extreme ultraviolet (EUV) emission from laser-produced tin plasma was investigated for 266 and 1064nm laser wavelengths. The EUV emission exhibits a laser-wavelength dependence in terms of angular distribution and structures of emission spectra. Angular distributions expressed in a form of I(θ)∝cosαθ became α=1.3 and 0.5, respectively, for 266 and 1064nm laser wavelength. It is found that spectra from 266nm laser plasma show dips at around 13.5nm that had been well replicated in computer simulations. Both angular distribution and spectral structure at 13.5nm suggest the existence of an opaque plasma region in front of the EUV source plasma generated by 266nm radiation.