bluuebunny/crossref_metadata_2025 huggingface dataset:
The zero-frequency ion ring instability
Authors: Gary S. Peter , Pongratz Morris B. , Madland Christian D. , Swift Daniel W.
Published: July 2002

The electrostatic zero-frequency ion ring instability with wave vector perpendicular to a uniform magnetic field B is examined through linear and second-order theory as well as by computer simulation. In the simulation ions are taken as magnetized particles; the electrons are described as a massless fluid subject to E×B motion. Saturation of the instability is primarily due to broadening of the ion ring distribution. A second-order theory provides an approximate criterion for the saturation amplitude, as does a simple trapping argument. Thus, for the simulation presented here, both quasilinear and trapping effects contribute to saturation.
The motion of a spherical particle suspended in a turbulent flow near a plane wall
Authors: Rizk M. A. , Elghobashi S. E.
Published: July 2002

Analytical solution of the equations of motion of a spherical particle suspended in a turbulent flow near a plane wall has been obtained. The equations include the lift force and wall effects on the drag force. The solution shows that the particle turbulent motion is affected by the wall presence in the following manner: (a) The wall augments the response of the particle to fluid turbulence. The ratio between the particle rms velocity fluctuation near the wall and that of an identical particle in an unbounded flow is always greater than unity. This ratio increases by increasing the particle density and diameter and decreasing the particle distance from the wall. (b) Wall effects in a direction normal to it are more pronounced than those in the parallel direction. This is attributed mainly to the lift force acting in the normal direction. (c) Effects of the drag force on particle intensity are confined close to the wall whereas the lift effects extend to larger distances.
A simulation study of the Alfvén ion-cyclotron instability in high-beta plasmas
Authors: Ambrosiano John , Brecht Stephen H.
Published: July 2002

A one-dimensional electromagnetic particle-in-cell (PIC) model is used to investigate the Alfvén ion-cyclotron (AIC) instability in high-β plasmas with large ion temperature anisotropies. The simulation results are compared with linear and quasilinear theory. Nonlinear effects, such as the rate at which the anisotropy relaxes and the level of perturbed magnetic wave energy at saturation, are compared as a function of β and of the initial ion anisotropy. It is found that for large β⊥ (β⊥>3) the anisotropy decays to a residual level in a time comparable to an ion-cyclotron period. Simulations designed to drive the instability by replenishing particles from an anisotropic temperature reservoir are also discussed.
Numerical calculation of nonlinear axisymmetric standing waves in a circular basin
Authors: Tsai Wu-Ting , Yue Dick K. P.
Published: July 2002

The solution of fully nonlinear first-mode axisymmetric standing waves in a circular basin is obtained numerically by direct collocation of truncated Fourier and Dini series in time and radial coordinates, respectively. The method produces accurate results which offer substantial improvements, especially in shallow depth, over existing solutions based on perturbation expansions. Numerical results for the frequency, partition of energy, surface profiles, and particle trajectories are presented for both deep and shallow water for a range of steepnesses. It is found that salient features such as the nonmonotonic dependence of frequency on steepness at certain depths, characteristic of two-dimensional standing waves, are also observed for axisymmetric standing waves.
A note on kinetic energy, dissipation and enstrophy
Authors: Wu Jie-Zhi , Zhou Ye , Fan Meng
Published: July 2002

We show that the dissipation rate of a Newtonian fluid with constant shear viscosity has three constituents from dilatation, vorticity, and surface strain. The last one, being associated with the most complicated manners of fluid deformation, only contributes to the change of internal energy but not that of kinetic energy. The distinction of these dissipation constituents is used to identify typical compact flow structures at high Reynolds numbers, such as shock waves, thin vortex layers, and filaments. This identification is of particular interest in studying turbulence structures. We then cast the incompressible version of the simplified kinetic-energy transport equation to a novel form, which is free from the work rate done by surface stresses but in which the full dissipation reenters. This result is of relevance to the physical routes via which turbulent energy is transferred to small scales.
First high power experiments with the Dutch free electron maser
Authors: Verhoeven A. G. A. , Bongers W. A. , Bratman V. L. , Caplan M. , Denisov G. G. , van Dijk G. , van der Geer C. A. J. , Manintveld P. , Poelman A. J. , Pluygers J. , Shmelyov M. Yu. , Smeets P. H. M. , Sterk A. B. , Urbanus W. H.
Published: July 2002

A free electron maser (FEM) has been built as a mm-wave source for applications on future fusion research devices such as ITER, the international tokamak experimental reactor [M. A. Makowski, F. Elio, and D. Loeser, April 97, Proc. 10th Workshop on ECE and ECRH, EC10, 549-559. World Scientific (1998)]. A unique feature of the Dutch fusion-FEM is the possibility to tune the frequency over the entire range from 130 to 260 GHz at an output power exceeding 1 MW. In the first phase of the project, a so-called inverse setup is used. The electron gun is mounted inside the high-voltage terminal. The entire beam line was tested successfully with extremely low loss current, lower than 0.05%. This included the accelerating structure up to 2 MV level and the transport through the undulator. First generation of mm-waves was achieved in October 1997. With an electron beam current around 8 A and an accelerator voltage of 1.76 MV the mm-wave pulse starts after 3 μs and lasts for 3 μs, reaching a maximum saturated peak power level of more than 500 kW at a frequency of 200 GHz. Output power, start-up time, and frequency correspond well with simulation results.
Production of large-area plasmas by electron beams
Authors: Fernsler R. F. , Manheimer W. M. , Meger R. A. , Mathew J. , Murphy D. P. , Pechacek R. E. , Gregor J. A.
Published: July 2002

An analysis is presented for the production of weakly ionized plasmas by electron beams, with an emphasis on the production of broad, planar plasmas capable of reflecting X-band microwaves. Considered first in the analysis is the ability of weakly ionized plasmas to absorb, emit and reflect electromagnetic radiation. Following that is a determination of the electron beam parameters needed to produce plasmas, based on considerations of beam ionization, range, and stability. The results of the analysis are then compared with a series of experiments performed using a sheet electron beam to produce plasmas up to 0.6 m square by 2 cm thick. The electron beam in the experiments was generated using a long hollow-cathode discharge operating in an enhanced-glow mode. That mode has only recently been recognized, and a brief analysis of it is given for completeness. The conclusion of the study is that electron beams can produce large-area, planar plasmas with high efficiency, minimal gas heating, low electron temperature, high uniformity, and high microwave reflectivity, as compared with plasmas produced by other sources.
Tests of local transport theory and reduced wall impurity influx with highly radiative plasmas in the Tokamak Fusion Test Reactor
Authors: Hill K. W. , Scott S. D. , Bell M. , Budny R. , Bush C. E. , Clark R. E. H. , Denne-Hinnov B. , Ernst D. R. , Hammett G. W. , Mikkelsen D. R. , Mueller D. , Ongena J. , Park H. K. , Ramsey A. T. , Synakowski E. J. , Taylor G. , Zarnstorff M. C. ,   the TFTR Group
Published: July 2002

The electron temperature (Te) profile in neutral beam-heated supershot plasmas (Te0∼6–7 keV ion temperature Ti0∼15–20 keV, beam power Pb∼16 MW) was remarkably invariant when radiative losses were increased significantly through gas puffing of krypton and xenon in the Tokamak Fusion Test Reactor [McGuire et al., Phys. Plasmas 2, 2176 (1995)]. Trace impurity concentrations (nz/ne∼10−3) generated almost flat and centrally peaked radiation profiles, respectively, and increased the radiative losses to 45%–90% of the input power (from the normal ∼25%). Energy confinement was not degraded at radiated power fractions up to 80%. A 20%–30% increase in Ti, in spite of an increase in ion–electron power loss, implies a factor of ∼3 drop in the local ion thermal diffusivity. These experiments form the basis for a nearly ideal test of transport theory, since the change in the beam heating power profile is modest, while the distribution of power flow between (1) radiation and (2) conduction plus convection changes radically and is locally measurable. The decrease in Te was significantly less than predicted by two transport models and may provide important tests of more complete transport models. At input power levels of 30 MW, the increased radiation eliminated the catastrophic carbon influx (carbon “bloom”) and performance (energy confinement and neutron production) was improved significantly relative to that of matched shots without impurity gas puffing.
Efficiency reduction caused intense rf-induced E×B drift during relativistic magnetron operation
Authors: Riyopoulos Spilios
Published: July 2002

At high power relativistic magnetron operation the rf-induced drift velocity becomes of the same order as the dc E×B velocity, causing a significant increase in the relativistic factor γ and decrease in electronic efficiency. The relativistic guiding center flow above the hub is analyzed. A second order expansion in eVrf/mc2 reveals that the first order (parapotential) rf-drift introduces a second order effective potential equal to the rf-drift kinetic energy, causing deviations from the parapotential flow. A formula for high power magnetron efficiency with full-relativistic effects is derived and discussed.
Response of internal transport barriers to central electron heating and current drive on ASDEX Upgrade
Authors: Wolf R. C. , Günter S. , Leuterer F. , Peeters A. , Pereverzev G. , Gruber O. , Kaufmann M. , Lackner K. , Maraschek M. , McCarthy P. J. , Meister H. , Salzmann H. , Schade S. , Schweinzer J. , Suttrop W. ,   ASDEX Upgrade Team
Published: July 2002

Internal transport barriers with the central electron temperature as large as the central ion temperature both in excess of 10 keV have been achieved in the Axi-symmetric divertor experiment (ASDEX Upgrade) [H. Vernickel et al., J. Nucl. Mater. 128, 71 (1984)]. By applying central electron cyclotron heating and current drive to negative central shear discharges, established by neutral beam heating in the current ramp, the core electron temperatures could be raised by more than a factor of 2. Despite the fivefold increase of the central electron heat flux, the ion and electron energy and also angular momentum transport did not deteriorate. For neutral beam injection alone and also with additional central electron cyclotron counter-current drive, a double tearing mode and the associated detrimental effect on the plasma confinement is destabilized only transiently, when the minimum of the safety factor (qmin) passes through 2. For co-current drive, however, the confinement does not recover after qmin has dropped below 2, as the (2,1) mode persists due to the influence of the current profile modification in the very plasma center.
High-power second harmonic emission and frequency locking in a 28-GHz gyrotron
Authors: Danly B. G. , Mulligan W. J. , Temkin R. J. , Sollner T. C. L. G.
Published: July 2002

Two important new results are presented from a study of second harmonic (2ωc) emission from a pulsed gyrotron designed for fundamental (ωc) emission of 200 kW at 28 GHz in the TE021 mode. Simultaneous output at ω1≊ωc and ω2≊2ωc, with the condition ω2=2ω1 applying as ω1 was varied, has been observed for the first time. Strong, single mode 2ωc emission (80 kW at 50 GHz) is also observed, and such emission is shown to be characteristic of gyrotrons designed for high-power emission at the fundamental.
Pyroelectric effects in LiNbO3 channel-waveguide devices
Authors: Bulmer C. H. , Burns W. K. , Hiser S. C.
Published: July 2002

The thermal stability of Ti-indiffused LiNbO3 channel-waveguide devices is described. In Z-cut interferometers the pyroelectric effect causes large output instabilities. However, both Z-cut directional couplers and X-cut interferometers show good thermal stability. We discuss the pyroelectric effect and explain device thermal behavior. We describe the decay of induced voltage or of change in interferometer phase bias after a temperature change and deduce that LiNbO3 resistivities can be electric field dependent.
Optical absorption of some polymers in the region 240–170 nm
Authors: Philipp H. R. , Cole H. S. , Liu Y. S. , Sitnik T. A.
Published: July 2002

Absorption coefficients for some technologically important polymer materials are given in the wavelength range ∼240–170 nm. Absorption coefficients at 193 nm for these polymers show a wide range of values from ∼2×102 cm−1 for polytetrafluoroethylene (Teflon) to ∼4×105 cm−1 for polyimide. The general nature of the optical properties of polymers in the vacuum ultraviolet is also discussed.
Costimulation of
N
-methyl-
d
-aspartate and muscarinic neuronal receptors modulates gap junctional communication in striatal astrocytes
Authors: Rouach N. , Tencé M. , Glowinski J. , Giaume C.
Published: July 2002

Cocultures of neurons and astrocytes from the rat striatum were used to determine whether the stimulation of neuronal receptors could affect the level of intercellular communication mediated by gap junctions in astrocytes. The costimulation of
N
-methyl-D-asparte (NMDA) and muscarinic receptors led to a prominent reduction of astrocyte gap junctional communication (GJC) in coculture. This treatment was not effective in astrocyte cultures, these cells being devoid of NMDA receptors. Both types of receptors contribute synergistically to this inhibitory response, as the reduction in astrocyte GJC was not observed after the blockade of either NMDA or muscarinic receptors. The involvement of a neuronal release of arachidonic acid (AA) in this inhibition was investigated because the costimulation of neuronal NMDA and muscarinic receptors markedly enhanced the release of AA in neuronal cultures and in cocultures. In addition, both the reduction of astrocyte GJC and the release of AA evoked by NMDA and muscarinic receptor costimulation were prevented by mepacrine, a phospholipase A
2
inhibitor, and this astrocyte GJC inhibition was mimicked by the exogenous application of AA. Metabolites of AA formed through the cyclooxygenase pathway seem to be responsible for the effects induced by either the costimulation of NMDA and muscarinic neuronal receptors or the application of exogenous AA because, in both cases, astrocyte GJC inhibition was prevented by indomethacin. Altogether, these data provide evidence for a neuronal control of astrocytic communication and open perspectives for the understanding of the modalities through which cholinergic interneurons and glutamatergic inputs affect local circuits in the striatum.
Amplification–mutagenesis: Evidence that “directed” adaptive mutation and general hypermutability result from growth with a selected gene amplification
Authors: Hendrickson Heather , Slechta E. Susan , Bergthorsson Ulfar , Andersson Dan I. , Roth John R.
Published: July 2002

When a particular
lac
mutant of
Escherichia coli
starves in the presence of lactose, nongrowing cells appear to direct mutations preferentially to sites that allow growth (adaptive mutation). This observation suggested that growth limitation stimulates mutability. Evidence is provided here that this behavior is actually caused by a standard Darwinian process in which natural selection acts in three sequential steps. First, growth limitation favors growth of a subpopulation with an amplification of the mutant
lac
gene; next, it favors cells with a
lac
+
revertant allele within the amplified array. Finally, it favors loss of mutant copies until a stable haploid
lac
+
revertant arises and overgrows the colony. By increasing the
lac
copy number, selection enhances the likelihood of reversion within each developing clone. This sequence of events appears to direct mutations to useful sites. General mutagenesis is a side-effect of growth with an amplification (SOS induction). The F′ plasmid, which carries
lac
, contributes by stimulating gene duplication and amplification. Selective stress has no direct effect on mutation rate or target specificity, but acts to favor a succession of cell types with progressively improved growth on lactose. The sequence of events—amplification, mutation, segregation—may help to explain both the origins of some cancers and the evolution of new genes under selection.
The lethal and edema factors of anthrax toxin bind only to oligomeric forms of the protective antigen
Authors: Mogridge Jeremy , Cunningham Kristina , Lacy D. Borden , Mourez Michael , Collier R. John
Published: July 2002

The three proteins that comprise anthrax toxin, edema factor (EF), lethal factor (LF), and protective antigen (PA), assemble at the mammalian cell surface into toxic complexes. After binding to its receptor, PA is proteolytically activated, yielding a carboxyl-terminal 63-kDa fragment (PA
63
) that coordinates assembly of the complexes, promotes their endocytosis, and translocates EF and LF to the cytosol. PA
63
spontaneously oligomerizes to form symmetric ring-shaped heptamers that are capable of binding three molecules of EF and/or LF as competing ligands. To determine whether binding of these ligands depends on oligomerization of PA
63
, we prepared two oligomerization-deficient forms of this protein, each mutated on a different PA
63
–PA
63
contact face. In solution or when bound to receptors on Chinese hamster ovary K1 cells, neither mutant alone bound ligand, but a mixture of them did. After the two mutants were proteolytically activated and mixed with ligand in solution, a ternary complex was isolated containing one molecule of each protein. Thus EF and LF bind stably only to PA
63
dimers or higher order oligomers. These findings are relevant to the kinetics and pathways of assembly of anthrax toxin complexes.
The optimal measure of allelic association
Authors: Morton N. E. , Zhang W. , Taillon-Miller P. , Ennis S. , Kwok P.-Y. , Collins A.
Published: July 2002

Allelic association between pairs of loci is derived in terms
of the association probability ρ as a function of recombination θ,
effective population size
N
, linear systematic pressure
v
, and time
t
, predicting both
ρ

rt

, the decrease of association from
founders and ρ

ct

, the increase by genetic
drift, with ρ

t

=
ρ

rt

+ ρ

ct

.
These results conform to the Malecot equation, with time replaced by
distance on the genetic map, or on the physical map if recombination in
the region is uniform. Earlier evidence suggested that ρ is less
sensitive to variations in marker allele frequencies than alternative
metrics for which there is no probability theory. This robustness is
confirmed for six alternatives in eight samples. In none of these 48
tests was the residual variance as small as for ρ. Overall,
efficiency was less than 80% for all alternatives, and less than 30%
for two of them. Efficiency of alternatives did not increase when
information was estimated simultaneously. The swept radius within which
substantial values of ρ are conserved lies between 385 and 893 kb,
but deviation of parameters between measures is enormously significant.
The large effort now being devoted to allelic association has little
value unless the ρ metric with the strongest theoretical basis and
least sensitivity to marker allele frequencies is used for mapping of
marker association and localization of disease loci.
Steroid hormones induce HMG1 overexpression and sensitize breast cancer cells to cisplatin and carboplatin
Authors: He Qing , Liang Cynthia H. , Lippard Stephen J.
Published: July 2002

Cisplatin is an anticancer drug that has enjoyed remarkable success against testicular tumors, but dose limiting side-effects have limited its application against a broader range of cancers. Previous studies have shown that high-mobility group (HMG) domain proteins such as HMG1 sensitize cells to cisplatin by shielding its major DNA adducts from nucleotide excision repair. Estrogen treatment increases HMG1 mRNA levels in breast cancer MCF-7 cells. Herein, we describe that treatment of human cancer cells having steroid hormone receptors with the appropriate hormone, estrogen and/or progesterone, significantly increases the potency of cisplatin and its analogue carboplatin by causing the overexpression of HMG1. These findings suggest that the proper combination of these drugs, which are already approved by the Food and Drug Administration, could have potential benefit in treating tumors such as ovarian or breast that carry the hormone receptors.
Maintenance of caspase-3 proenzyme dormancy by an intrinsic “safety catch” regulatory tripeptide
Authors: Roy Sophie , Bayly Christopher I. , Gareau Yves , Houtzager Vicky M. , Kargman Stacia , Keen Sabina L. C. , Rowland Kathleen , Seiden Isolde M. , Thornberry Nancy A. , Nicholson Donald W.
Published: July 2002

Caspase-3 is synthesized as a dormant proenzyme and is maintained
in an inactive conformation by an Asp-Asp-Asp “safety-catch”
regulatory tripeptide contained within a flexible loop near the
large-subunit/small-subunit junction. Removal of this “safety
catch” results in substantially enhanced autocatalytic maturation as
well as increased vulnerability to proteolytic activation by upstream
proteases in the apoptotic pathway such as caspase-9 and
granzyme B. The safety catch functions through multiple ionic
interactions that are disrupted by acidification, which occurs in the
cytosol of cells during the early stages of apoptosis. We
propose that the caspase-3 safety catch is a key regulatory checkpoint
in the apoptotic cascade that regulates terminal events in the
caspase cascade by modulating the triggering of caspase-3 activation.
Sense- and antisense-mediated gene silencing in tobacco is inhibited by the same viral suppressors and is associated with accumulation of small RNAs
Authors: Di Serio Francesco , Schöb Hanspeter , Iglesias Alejandro , Tarina Corina , Bouldoires Estelle , Meins Frederick
Published: July 2002

Antisense-mediated gene silencing (ASGS) and posttranscriptional
gene silencing (PTGS) with sense transgenes markedly reduce the
steady-state mRNA levels of endogenous genes similar in
transcribed sequence. RNase protection assays established that
silencing in tobacco plants transformed with plant-defense-related
class I sense and antisense chitinase (
CHN
) transgenes
is at the posttranscriptional level. Infection of tobacco plants with
cucumber mosaic virus strain FN and a necrotizing strain of potato
virus Y, but not with potato virus X, effectively suppressed PTGS and
ASGS of both the transgenes and homologous endogenes. This suggests
that ASGS and PTGS share components associated with initiation and
maintenance of the silent state. Small,
ca.
25-nt RNAs
(smRNA) of both polarities were associated with PTGS and ASGS in
CHN
transformants as reported for PTGS in other
transgenic plants and for RNA interference in
Drosophila
. Similar results were obtained with an
antisense class I β-1,3-glucanase transformant showing that viral
suppression and smRNAs are a more general feature of ASGS. Several
current models hold that diverse signals lead to production of
double-stranded RNAs, which are processed to smRNAs that then trigger
PTGS. Our results provide direct evidence for mechanistic links between
ASGS and PTGS and suggest that ASGS could join a common PTGS pathway at
the double-stranded RNA step.
Stage-specific modulation of skeletal myogenesis by inhibitors of nuclear deacetylases
Authors: Iezzi Simona , Cossu Giulio , Nervi Clara , Sartorelli Vittorio , Puri Pier Lorenzo
Published: July 2002

Nuclear acetyltransferases promote and deacetylases inhibit skeletal muscle-gene expression, suggesting the potential effectiveness of deacetylase inhibitors (DIs) in modulating skeletal myogenesis. Surprisingly, previous studies have indicated that DIs suppress myogenesis. The recent observations that histone deacetylases associate with the muscle-regulatory proteins MyoD and MEF2C only in undifferentiated myoblasts prompted us to evaluate the effect of DIs at distinct stages of the myogenic program. We found that exposure of established rodent and human muscle cells to distinct DIs has stage-specific effects. Exposure of undifferentiated skeletal myoblasts to DIs, followed by incubation in differentiation medium, enhanced the expression of muscle-specific reporters and increased the levels of endogenous muscle proteins, leading to a dramatic increase in the formation of multinucleated myotubes. By contrast, simultaneous exposure of muscle cells to differentiation medium and DIs inhibited the myogenic program. Likewise, embryos exposed
in utero
to nonteratogenic doses of DI at the early stages of somitic myogenesis (embryonic day 8.5) exhibited an increased number of somites and augmented expression of a muscle-specific transgene as well as endogenous muscle genes. The functional effects induced by DIs were mirrored by changes in the state of acetylation of histones present at a muscle-gene enhancer and of MyoD itself. These results represent the first evidence that DIs can enhance muscle differentiation and suggest the rationale for their use in manipulating adult and embryonic skeletal myogenesis.
Evolutionary genomics of
Staphylococcus aureus
: Insights into the origin of methicillin-resistant strains and the toxic shock syndrome epidemic
Authors: Fitzgerald J. Ross , Sturdevant Daniel E. , Mackie Stacy M. , Gill Steven R. , Musser James M.
Published: July 2002

An emerging theme in medical microbiology is that extensive variation exists in gene content among strains of many pathogenic bacterial species. However, this topic has not been investigated on a genome scale with strains recovered from patients with well-defined clinical conditions.
Staphylococcus aureus
is a major human pathogen and also causes economically important infections in cows and sheep. A DNA microarray representing >90% of the
S. aureus
genome was used to characterize genomic diversity, evolutionary relationships, and virulence gene distribution among 36 strains of divergent clonal lineages, including methicillin-resistant strains and organisms causing toxic shock syndrome. Genetic variation in
S. aureus
is very extensive, with ≈22% of the genome comprised of dispensable genetic material. Eighteen large regions of difference were identified, and 10 of these regions have genes that encode putative virulence factors or proteins mediating antibiotic resistance. We find that lateral gene transfer has played a fundamental role in the evolution of
S. aureus.
The
mec
gene has been horizontally transferred into distinct
S. aureus
chromosomal backgrounds at least five times, demonstrating that methicillin-resistant strains have evolved multiple independent times, rather than from a single ancestral strain. This finding resolves a long-standing controversy in
S. aureus
research. The epidemic of toxic shock syndrome that occurred in the 1970s was caused by a change in the host environment, rather than rapid geographic dissemination of a new hypervirulent strain. DNA microarray analysis of large samples of clinically characterized strains provides broad insights into evolution, pathogenesis, and disease emergence.
Frataxin activates mitochondrial energy conversion and oxidative phosphorylation
Authors: Ristow Michael , Pfister Markus F. , Yee Andrew J. , Schubert Markus , Michael Laura , Zhang Chen-Yu , Ueki Kojhiro , Michael M. Dodson , Lowell Bradford B. , Kahn C. Ronald
Published: July 2002

Friedreich's ataxia (FA) is an autosomal recessive disease caused
by decreased expression of the mitochondrial protein frataxin. The
biological function of frataxin is unclear. The homologue of frataxin
in yeast,
YFH1
, is required for cellular respiration and
was suggested to regulate mitochondrial iron homeostasis. Patients
suffering from FA exhibit decreased ATP production in skeletal muscle.
We now demonstrate that overexpression of frataxin in mammalian cells
causes a Ca
2+
-induced up-regulation of tricarboxylic acid
cycle flux and respiration, which, in turn, leads to an increased
mitochondrial membrane potential (Δψ
m
) and results in
an elevated cellular ATP content. Thus, frataxin appears to be a key
activator of mitochondrial energy conversion and oxidative
phosphorylation.
Detection of DNA point mutations and mRNA expression levels by rolling circle amplification in individual cells
Authors: Christian Allen T. , Pattee Melissa S. , Attix Christina M. , Reed Beth E. , Sorensen Karen J. , Tucker James D.
Published: July 2002

Rolling circle amplification has been useful for detecting point
mutations in isolated nucleic acids, but its application in cytological
preparations has been problematic. By pretreating cells with a
combination of restriction enzymes and exonucleases, we demonstrate
that rolling circle amplification
in situ
can detect
gene copy number and single base mutations in fixed cells with
efficiencies up to 90%. It can also detect and quantify transcribed
RNA in individual cells, making it a versatile tool for cell-based
assays.
Hemodynamic effects of epidermal growth factor in conscious rats and monkeys.
Authors: Keiser J A , Ryan M J
Published: July 2002

The therapeutic application of growth factors to human disease has become closer to reality with the advent of faster means of synthesizing these molecules and novel drug delivery strategies. Epidermal growth factor (EGF) belongs to a large family of molecules with the ability to modulate growth. Purified extracts of EGF have been used clinically to modulate gastrointestinal secretion of hormones and accelerate healing. EGF is also reported to have both vascular smooth muscle contractile and relaxing activity Cardiovascular studies were performed with the bioactive 48-amino acid fragment of human EGF in rodents and primates to determine the effects of EGF on blood pressure and heart rate in conscious animals. Intravenous infusion of EGF induced an initial pressor response in rats followed by a prolonged decrease in blood pressure. In contrast, in monkeys, EGF had dose-related blood pressure-lowering effects only; significant hypotension was observed at doses ranging from 3 to 300 microg/kg i.v. Hypotension was associated with modest tachycardia in both species. To our knowledge, this is the first report of hemodynamic effects of EGF in primates, and it clearly documents that the mitogenic role of growth factors such as EGF is but one aspect of their physiology.
Exploring the active site of chorismate mutase by combinatorial mutagenesis and selection: the importance of electrostatic catalysis.
Authors: Kast P , Asif-Ullah M , Jiang N , Hilvert D
Published: July 2002

Chorismate mutase (EC 5.4.99.5) catalyzes the intramolecular rearrangement of chorismate to prephenate. Arg-90 in the active site of the enzyme from Bacillus subtilis is in close proximity to the substrate's ether oxygen and may contribute to efficient catalysis by stabilizing the presumed dipolar transition state that would result upon scission of the C--O bond. To test this idea, we have developed a novel complementation system for chorismate mutase activity in Escherichia coli by reengineering parts of the aromatic amino acid biosynthetic pathway. The codon for Arg-90 was randomized, alone and in combination with that for Cys-88, and active clones were selected. The results show that a positively charged residue either at position 88 (Lys) or 90 (Arg or Lys) is essential. Our data provide strong support for the hypothesis that the positive charge is required for stabilization of the transition state of the enzymatic chorismate rearrangement. The new selection system, in conjunction with combinatorial mutagenesis, renders the mechanism of the natural enzyme(s) accessible to further exploration and opens avenues for the improvement of first generation catalytic antibodies with chorismate mutase activity.
Purification and cDNA cloning of a second apoptosis-related cysteine protease that cleaves and activates sterol regulatory element binding proteins.
Authors: Pai J T , Brown M S , Goldstein J L
Published: July 2002

We have purified from hamster liver a second cysteine protease that cleaves and activates sterol regulatory element binding proteins (SREBPs). cDNA cloning revealed that this enzyme is the hamster equivalent of Mch3, a human enzyme that is related to the interleukin 1beta converting enzyme. We call this enzyme Mch3/SCA-2. It is 54% identical to hamster CPP32/SCA-1, a cysteine protease that was earlier shown to cleave SREBPs at a conserved Asp between the basic helix-loop-helix leucine zipper domain and the membrane attachment domain. This cleavage liberates an NH2-terminal fragment of approximately 460 amino acids that activates transcription of genes encoding the low density lipoprotein receptor and enzymes of cholesterol synthesis. Mch3/SCA-2 and CPP32/SCA-I are synthesized as inactive 30-35 kDa precursors that are thought to be cleaved during apoptosis to generate active fragments of approximately 20 and approximately 10 kDa. The current data lend further support to the notion that SREBPs are cleaved and activated as part of the program in programmed cell death.
Mutation of Pro-258 in transmembrane domain 6 constitutively activates the G protein-coupled alpha-factor receptor.
Authors: Konopka J B , Margarit S M , Dube P
Published: July 2002

The alpha-factor pheromone receptor stimulates MATa yeast cells to undergo conjugation. The receptor contains seven transmembrane domains that function in ligand binding and in transducing a signal to the cytoplasmic receptor sequences to mediate G protein activation. A genetic screen was used to isolate receptor mutations that constitutively signal in the absence of alpha-factor. The Pro-258-->Leu (P258L) mutation caused constitutive receptor signaling that was equivalent to about 45% of the maximum level observed in wild-type cells stimulated with alpha-factor. Mutations of both Pro-258 and the adjacent Ser-259 to Leu increased constitutive signaling to > or = 90% of the maximum level. Since Pro-258 occurs in the central portion of transmembrane domain 6, and since proline residues are expected to cause a kink in alpha-helical domains, the P258L mutation is predicted to alter the structure of transmembrane domain 6. The P258L mutation did not result in a global distortion of receptor structure because alpha-factor bound to the mutant receptors with high affinity and induced even higher levels of signaling. These results suggest that sequences surrounding Pro-258 may be involved in ligand activation of the receptor. Conformational changes in transmembrane domain 6 may effect a change in the adjacent sequences in the third intracellular loop that are thought to function in G protein activation. Greater than 90% of all G protein-coupled receptors contain a proline residue at a similar position in transmembrane domain 6, suggesting that this aspect of receptor activation may be conserved in other receptors.
Suppression of diet-induced atherosclerosis in low density lipoprotein receptor knockout mice overexpressing lipoprotein lipase.
Authors: Shimada M , Ishibashi S , Inaba T , Yagyu H , Harada K , Osuga J I , Ohashi K , Yazaki Y , Yamada N
Published: July 2002

Lipoprotein lipase (LPL) is a key enzyme in the hydrolysis of triglyceride-rich lipoproteins. Conflicting results have been reported concerning its role in atherogenesis. To determine the effects of the overexpressed LPL on diet-induced atherosclerosis, we have generated low density lipoprotein receptor (LDLR) knockout mice that overexpressed human LPL transgene (LPL/LDLRKO) and compared their plasma lipoproteins and atherosclerosis with those in nonexpressing LDLR-knockout mice (LDLRKO). On a normal chow diet, LPL/LDLRKO mice showed marked suppression of mean plasma triglyceride levels (32 versus 236 mg/dl) and modest decrease in mean cholesterol levels (300 versus 386 mg/dl) as compared with LDLRKO mice. Larger lipoprotein particles of intermediate density lipoprotein (IDL)/LDL were selectively reduced in LPL/LDLRKO mice. On an atherogenic diet, both mice exhibited severe hypercholesterolemia. But, mean plasma cholesterol levels in LPL/ LDLRKO mice were still suppressed as compared with that in LDLRKO mice (1357 versus 2187 mg/dl). Marked reduction in a larger subfraction of IDL/LDL, which conceivably corresponds to remnant lipoproteins, was observed in the LPL/LDLRKO mice. LDLRKO mice developed severe fatty streak lesions in the aortic sinus after feeding with the atherogenic diet for 8 weeks. In contrast, mean lesion area in the LPL/LDLRKO mice was 18-fold smaller than that in LDLRKO mice. We suggest that the altered lipoprotein profile, in particular the reduced level of remnant lipoproteins, is mainly responsible for the protection by LPL against atherosclerosis.
COOH-terminal processing of nascent polypeptides by the glycosylphosphatidylinositol transamidase in the presence of hydrazine is governed by the same parameters as glycosylphosphatidylinositol addition.
Authors: Ramalingam S , Maxwell S E , Medof M E , Chen R , Gerber L D , Udenfriend S
Published: July 2002

Proteins anchored to the cell membrane via a glycosylphosphatidylinositol (GPI) moiety are found in all eukaryotes. After NH2-terminal peptide cleavage of the nascent protein by the signal peptidase, a second COOH-terminal signal peptide is cleaved with the concomitant addition of the GPI unit. The proposed mechanism of the GPI transfer is a transamidation reaction that involves the formation of an activated carbonyl intermediate (enzyme-substrate complex) with the ethanolamine moiety of the preassembled GPI unit serving as a nucleophile. Other nucleophilic acceptors like hydrazine (HDZ) and hydroxylamine have been shown to be possible alternate substrates for GPI. Since GPI has yet to be purified, the use of readily available nucleophilic substitutes such as HDZ and hydroxylamine is a viable alternative to study COOH-terminal processing by the putative transamidase. As a first step in developing a soluble system to study this process, we have examined the amino acid requirements at the COOH terminus for the transamidation reaction using HDZ as the nucleophilic acceptor instead of GPI. The hydrazide-forming reaction shows identical amino acid requirement profiles to that of GPI anchor addition. Additionally, we have studied other parameters relating to the kinetics of the transamidation reaction in the context of rough microsomal membranes. The findings with HDZ provide further evidence for the transamidase nature of the enzyme and also provide a starting point for development of a soluble assay.
Interaction of the histone (H3-H4)2 tetramer of the nucleosome with positively supercoiled DNA minicircles: Potential flipping of the protein from a left- to a right-handed superhelical form.
Authors: Hamiche A , Carot V , Alilat M , De Lucia F , O'Donohue M F , Revet B , Prunell A
Published: July 2002

We have studied the ability of the histone (H3-H4)2 tetramer, the central part of the nucleosome of eukaryotic chromatin, to form particles on DNA minicircles of negative and positive superhelicities, and the effect of relaxing these particles with topoisomerase I. The results show that even modest positive torsional stress from the DNA, and in particular that generated by DNA thermal fluctuations, can trigger a major, reversible change in the conformation of the particle. Neither a large excess of naked DNA, nor a crosslink between the two H3s prevented the transition from one form to the other. This suggested that during the transition, the histones neither dissociated from the DNA nor were even significantly reshuffled. Moreover, the particles reconstituted on negatively and positively supercoiled minicircles look similar under electron microscopy. These data agree best with a transition involving a switch of the wrapped DNA from a left- to a right-handed superhelix. It is further proposed, based on the left-handed overall superhelical conformation of the tetramer within the octamer [Arents, G., Burlingame, R. W., Wang, B. C., Love, W. E. & Moudrianakis, E. N. (1991) Proc. Natl.Acad. Sci. USA 88, 10148-10152] that this change in DNA topology is mediated by a similar change in the topology of the tetramer itself, which may occur through a rotation (or a localized deformation) of the two H3-H4 dimers about their H3-H3 interface. Potential implications of this model for nucleosome dynamics in vivo are discussed.
Effect of interleukin 12 on tumor induction by 3-methylcholanthrene.
Authors: Noguchi Y , Jungbluth A , Richards E C , Old L J
Published: July 2002

Interleukin (IL)-12 has strong antitumor activity in transplantable tumor systems in the mouse. The present study was designed to determine whether tumor induction by 3-methylcholanthrene (3-MC), a carcinogenic hydrocarbon, can be inhibited by IL-12. BALB/cBy mice were injected subcutaneously with 25 micrograms or 100 micrograms of 3-MC and treated with 100 ng, 10 ng, or 1 ng of IL-12 for 5 days a week for 18 weeks, with a schedule of 3 weeks on and 1 week off. In mice injected with 25 micrograms of 3-MC, treatment with 100 ng of IL-12 delayed tumor appearance and reduced tumor incidence. Tumor appearance was also delayed in mice injected with 100 micrograms of 3-MC and treated with 100 ng of IL-12, but the final tumor incidence was the same as in non-IL-12-treated mice. In contrast to the characteristically round, hard, well-circumscribed, and protruding tumor induced by 3-MC, a percentage of tumors induced in IL-12-treated mice had atypical characteristics: flat, soft, and invasive. Atypical tumors had a longer latent period and were more frequently seen in mice injected with 100 micrograms of 3-MC and treated with 100 ng of IL-12. Interferon gamma, IL-10, and tumor necrosis factor could be induced throughout the treatment period by IL-12, indicating that repeated injections of IL-12 do not induce a state of tachyphylaxis. High production of interferon gamma by CD8 T cells and a TH2-->TH1 or TH0 shift in the cytokine secretion profile of CD4 T cells were also seen in the IL-12-treated mice. IL-12 provides a powerful new way to explore the defensive role of the immune system in tumorigenesis.
Inhibition of gastric acid secretion by stress: A protective reflex mediated by cerebral nitric oxide
Authors: Esplugues J. V. , Barrachina M. D. , Beltrán B. , Calatayud S. , Whittle B. J. R. , Moncada S.
Published: July 2002

Moderate somatic stress inhibits gastric acid secretion. We have
investigated the role of endogenously released NO in this
phenomenon. Elevation of body temperature by 3°C or a reduction of 35
mmHg (1 mmHg = 133 Pa) in blood pressure for 10 min produced a
rapid and long-lasting reduction of distension-stimulated acid
secretion in the rat perfused stomach
in vivo
. A similar
inhibitory effect on acid secretion was produced by the intracisternal
(i.c.) administration of oxytocin, a peptide known to be released
during stress. Intracisternal administration of the NO-synthase
inhibitor,
N
G
-nitro-
l
-arginine
methyl ester (
l
-NAME) reversed the antisecretory effect
induced by all these stimuli, an action prevented by intracisternal
coadministration of the NO precursor,
l
-arginine.
Furthermore, microinjection of
l
-NAME into the dorsal motor
nucleus of the vagus nerve reversed the acid inhibitory effects of mild
hyperthermia, i.v. endotoxin, or i.c. oxytocin, an action
prevented by prior microinjection of
l
-arginine. By
contrast, microinjection of
l
-NAME into the nucleus tractus
solitarius failed to affect the inhibitory effects of hyperthermia,
i.v. endotoxin, or i.c. oxytocin. Immunohistochemical techniques
demonstrated that following hyperthermia there was a significant
increase in immunoreactivity to neuronal NO synthase in different areas
of the brain, including the dorsal motor nucleus of the vagus. Thus,
our results suggest that the inhibition of gastric acid secretion, a
defense mechanism during stress, is mediated by a nervous reflex
involving a neuronal pathway that includes NO synthesis in the brain,
specifically in the dorsal motor nucleus of the vagus.
SpoIIE governs the phosphorylation state of a protein regulating transcription factor sigma F during sporulation in Bacillus subtilis.
Authors: Arigoni F , Duncan L , Alper S , Losick R , Stragier P
Published: July 2002

Cell-specific activation of the transcription factor sigma F during sporulation in Bacillus subtilis is controlled by a regulatory pathway involving the proteins SpoIIE, SpoIIAA, and SpoIIAB. SpoIIAB is an antagonist of sigma F, and SpoIIAA, which is capable of overcoming SpoIIAB-mediated inhibition of sigma F, is an antagonist of SpoIIAB. SpoIIAA is, in turn, negatively regulated by SpoIIAB, which phosphorylates SpoIIAA on serine 58. SpoIIAA is also positively regulated by SpoIIE, which dephosphorylates SpoIIAA-P, the phosphorylated form of SpoIIAA. Here, isoelectric focusing and Western blot analysis were used to examine the phosphorylation state of SpoIIAA in vivo. SpoIIAA was found to be largely in the phosphorylated state during sporulation in wild-type cells but a significant portion of the protein that was unphosphorylated could also be detected. Consistent with the idea that SpoIIE governs dephosphorylation of SpoIIAA-P, SpoIIAA was entirely in the phosphorylated state in spoIIE mutant cells. Conversely, overexpression of spoIIE led to an increase in the ratio of unphosphorylated SpoIIAA to SpoIIAA-P and caused inappropriate activation of sigma F in the predivisional sporangium. We also show that a mutant form of SpoIIAA (SpoIIAA-S58T) in which serine 58 was replaced with threonine was present exclusively as SpoIIAA-P, a finding that confirms previous biochemical evidence that the mutant protein is an effective substrate for the SpoIIAB kinase but that SpoIIAA-S58T-P cannot be dephosphorylated by SpoIIE. We conclude that SpoIIE plays a crucial role in controlling the phosphorylation state of SpoIIAA during sporulation and thus in governing the cell-specific activation of sigma F.
A molecular mechanism for energy coupling in a membrane transport protein, the lactose permease of
Escherichia coli
Authors: Kaback H. Ronald
Published: July 2002

A mechanism for the coupled translocation of substrate and H
+
by the lactose permease of
Escherichia coli
is proposed, based on a variety of experimental observations. The permease is composed of 12 α-helical rods that traverse the membrane with the N and C termini on the cytoplasmic face. Four residues are irreplaceable with respect to coupling, and the residues are paired—Arg-302 (helix IX) with Glu-325 (helix X) and His-322 (helix X) with Glu-269 (helix VIII). In an adjacent region of the molecule at the interface between helices VIII and V is the substrate translocation pathway. Because of this arrangement, interfacial changes between helices VIII and V are transmitted to the interface between helices IX and X and vice versa. Upon ligand binding, a structural change at the interface between helices V and VIII disrupts the interaction between Glu-269 and His-322, Glu-269 displaces Glu-325 from Arg-302, and Glu-325 is protonated. Simultaneously, protonated Glu-325 becomes inaccessible to water, which drastically increases its pK
a
. In this configuration, the permease undergoes a freely reversible conformational change that corresponds to translocation of the ternary complex. To return to ground state after release of substrate, the Arg-302–Glu-325 interaction must be reestablished, which necessitates loss of H
+
from Glu-325. The H
+
is released into a water-filled crevice between helices IX and X which becomes transiently accessible to both sides of the membrane due to a change in helix tilt, where it is acted upon equally by either the membrane potential or the pH gradient across the membrane.