bluuebunny/crossref_metadata_2025 huggingface dataset:
Closed, form invariant solutions of convective–diffusive systems with applications to impurity transport
Authors: Mahajan S. M. , Valanju P. M. , Rowan William L.
Published: July 2002

Closed form Green’s functions are derived for convective–diffusive systems in cylindrical geometries. They are of the form e−αr 2I0(βr), where I0 is the modified Bessel function and α and β depend on time via e−t. These solutions can be used to model a variety of time-dependent transport experiments, such as pulsed impurity transport, oscillating gas puff, perturbative rf heating, and sawtooth heat pulse propagation. A successful comparison of the authors’ profiles with pulsed impurity injection experiments on the Texas Experimental Tokamak (TEXT) [K. W. Gentle, Nucl. Technol./Fusion 1, 479 (1981)] is presented.
Studies of self-consistent field structure in a quasioptical gyrotron
Authors: Antonsen T. M. , Bondeson A. , Roulin M. , Tran M. Q.
Published: July 2002

The presence of an electron beam in a quasioptical gyrotron cavity alters the structure of the fields from that of the empty cavity. A computer code has been written that calculates this alteration for either an electron beam or a thin dielectric tube placed in the cavity. Experiments measuring the quality factor of such a cavity were performed for the case of a dielectric tube and the results agree with the predictions of the code. Simulations of the case of an electron beam indicate that self-consistent effects can be made small, in that almost all the power leaves the cavity in a symmetric Gaussian-like mode, provided the resonator parameters are chosen carefully.
Superradiant free-electron lasers in the high-gain strong pump regime
Authors: Tsui K. H.
Published: July 2002

Time-dependent high-gain free-electron laser equations are generalized to the strong pump regime where fast, 2π/kw, spatial oscillations can no longer be eliminated by spatial averages, and evolution of the radiation phase needs to be taken into account. The equations are solved numerically along the characteristics of the beam and the laser field for superradiant solutions in the strong pump regime using the parameters of the Electron Laser Facility (ELF) of the Lawrence Livermore National Laboratory, [Phys. Rev. Lett. 54, 889 (1985); Nucl. Instrum. Methods A 250, 144 (1986); 285, 217 (1989); Phys. Rev. A 35, 2184 (1987)]. With beam pulses short compared to the wiggler length and long compared to the cooperation length, numerical solutions show that the initial laser fields get amplified to the steady-state level intensity in only a few bucket distances in the slippage region along the beam characteristics. The underlying mechanism for this superradiant gain gradient is due to the amplification of the incoming fields by the electrons prebunched by the earlier outgoing fields. Contrary to the earlier results, no spikes on the laser amplitude is observed in the slippage region. On the other hand, the solutions do show spikes on the relative laser phase. However, it is not clear on the credibility and the significance of these spikes, whether in amplitude or in phase.
High gain strong pump free-electron lasers with fast 2π/k
w spatial oscillations
Authors: Tsui K. H.
Published: July 2002

In the high gain strong pump regime, the radiation field amplitude and phase can vary substantially over a wiggler period. Also, due to the periodic transverse motion, the longitudinal electron energy suffers large variations over the same distance. Because of these considerations, the spatial average that leads to the classical one-dimensional free-electron laser equations is not applicable here. Thus the rederived equations suitable for the high gain strong pump regime contain 2π/kw periodic terms. Comparisons with the Electron Laser Facility (ELF) experiments of the Lawrence Livermore National Laboratory [Phys. Rev. Lett. 54, 889 (1985); 57, 2172 (1986); Phys. Rev. A 35, 2184 (1987)] on the operational characteristics including the relative phase of the radiation field are made.
Dominance of convective heat transport in the core of TFTR (Tokamak Fusion Test Reactor) supershot plasmas
Authors: Kissick M. W. , Efthimion P. C. , Mansfield D. K. , Callen J. D. , Bush C. E. , Park H. K. , Schivell J. , Synakowski E. J. , Taylor G.
Published: July 2002

Using perturbations in electron density and temperature induced by small helium gas puffs in TFTR (Tokamak Fusion Test Reactor) [Plasma Phys. Controlled Nucl. Fusion Res. 1, 51 (1986)], the dominance of convective heat transport in the core (r/a<0.4) of supershot plasmas has been demonstrated in a new way. The transp [J. Comput. Phys. 43, 61 (1981)] transport code was used to calculate the time-dependent particle and heat fluxes. Perturbations in the calculated convective and total electron heat fluxes were compared. They demonstrate that the conductive component decreases moving into the supershot core, and the convective component dominates in the supershot core. These results suggest a different transport drive in the supershot core compared to that in the rest of the supershot plasma.
Decay of turbulence in a power-law fluid
Authors: Sherwood J. D.
Published: July 2002

The final stages of decay of isotropic homogeneous turbulence are studied in a generalized Newtonian fluid, with a viscosity that depends on the mean rate of energy dissipation. The analysis is similar to that for Newtonian fluids. For a power-law fluid with power-law index n, the turbulent kinetic energy varies with time t as t−10/(7n−3), agreeing with the classical result t−5/2 when n=1. The decay is exponential when n=3/7, and turbulence vanishes in a finite time if n<3/7.
Stability of flow over axisymmetric bodies with porous suction strips
Authors: Nayfeh Ali H. , Reed H. L.
Published: July 2002

A numerical-perturbation scheme is used to determine the stability of incompressible flows past axisymmetric bodies with porous strips. Linear triple deck, closed-form solutions are developed for the basic-flow quantities, which account for upstream influence. These solutions are linear superpositions of the flow past the body without suction plus the perturbations caused by the suction strips. The flow past the suctionless body is calculated using the Transition Analysis Program System (TAPS). A perturbation scheme is used to determine the increment aij in the complex wavenumber at a given location sj because of the presence of a strip centered at si. The end result is a set of influence coefficients that can be used to determine the growth rates and amplification factors for any suction levels without repeating the calculations. The influence coefficients are used to develop a simple linear optimization scheme to determine the number, spacing, and mass flow rate through the strips on an axisymmetric body.
Electric breakdown waves: Exact solutions. Part II
Authors: Hemmati Mostafa , Fowler Richard G.
Published: July 2002

The methods of numerical solution of the electron fluid dynamical equations that govern wave breakdown, and were described in a previous paper, are applied here to the various situations under which these waves have been observed. Samples of the detailed structures of the waves are presented. Solutions have been investigated for both negatively and positively driven waves, into both ionized and nonionized media, and for waves followed by a large current as in a lightning return stroke.
Statistical analysis of the anisotropy of scalar diffusion in turbulent shear flows
Authors: Yoshizawa Akira
Published: July 2002

Anisotropy of the diffusion of a passive scalar is studied statistically by means of the direct-interaction approximation incorporated with the multiscale expansion method. The scalar flux generated by turbulent shear flows is calculated up to the second order of a scale parameter. As a result, a relation between the anisotropy of scalar diffusion and the inhomogeneity of turbulent flows is shown.
Linear stability analysis of the Vlasov–Poisson equations in high density plasmas in the presence of crossed fields and density gradients
Authors: Kaup D. J. , Hansen P. J. , Choudhury S. Roy , Thomas Gary E.
Published: July 2002

A method is presented for the linear stability analysis of the Vlasov–Poisson equations in high density, finite temperature plasmas in the presence of inhomogeneous crossed fields and density gradients. The method is more general than earlier studies of high-β inhomogeneous plasmas in that various approximations employed therein such as the local approximation (JWKB), low-frequency, or small wavelength restrictions are not employed here. Although only the nonrelativistic electrostatic planar case is treated, the method, with due modification, could be extended into the relativistic electromagnetic regime. The method uses a singular perturbation expansion to construct the unperturbed single particle orbits. Then with these orbits the ‘‘integration over the unperturbed orbits’’ necessary for determining the perturbed distribution function is performed. The initial distribution function may be quite general, but the expansions used do assume a distribution close to that of a sheared laminar flow. The perturbed distribution function is obtained as a singular perturbation expansion also. Lastly, the application of the method is demonstrated by reducing the linearized Vlasov–Poisson equations, with inhomogeneous electric fields and density gradients, to a second-order ordinary differential equation where the frequency is an eigenvalue. Similarities to and differences from the cold-fluid equations are pointed out.
Comparison between experiments and numerical simulations of three-dimensional plane wakes
Authors: Meiburg E. , Lasheras J. C.
Published: July 2002

The three-dimensional structure of a moderate Reynolds number (100) plane wake behind a flat plate subjected to periodic spanwise perturbations has been studied both experimentally and numerically. Comparisons between experimental interface visualizations and numerical calculations demonstrate that important features of the development of the three-dimensional instability can be reproduced by numerical, inviscid vortex dynamics. This suggests a promising way to analyze the physical processes involved in the initiation of the three-dimensional transition in wake flows.
Ignition density bounds and collective modes of fusing nuclei
Authors: Coppi B. , Tang W. M.
Published: July 2002

In the process of achieving ignition conditions collective modes (microinstabilities), which lead to enhanced transport of the fusing nuclei, are likely to be present. When such phenomena are taken into account an appropriate formulation of ignition criteria becomes necessary, and two distinct bounds on the plasma density are identified.
Large-eddy simulation of rotating channel flows using a localized dynamic model
Authors: Piomelli Ugo , Liu Junhui
Published: July 2002

Most applications of the dynamic subgrid-scale stress model use volume- or planar-averaging to avoid ill-conditioning of the model coefficient, which may result in numerical instabilities. Furthermore, a spatially-varying coefficient is mathematically inconsistent with the original derivation of the model. A localization procedure is proposed here that removes the mathematical inconsistency to any desired order of accuracy in time. This model is applied to the simulation of rotating channel flow, and results in improved prediction of the turbulence statistics. The model coefficient vanishes in regions of quiescent flow, reproducing accurately the intermittent character of the flow on the stable side of the channel. Large-scale longitudinal vortices can be identified, consistent with the observation from experiments and direct simulations. The effect of the unresolved scales on higher-order statistics is also discussed.
Distribution of particles suspended in convective flow in differentially heated cavity
Authors: Yarin A. L. , Kowalewski T. A. , Hiller W. J. , Koch St.
Published: July 2002

Our aim is to explore, both experimentally and theoretically, the cumulative effects of small particle–liquid density difference, where the particles are used as tracers in recirculating flow. As an example we take a flow field generated in a differentially heated cavity. The main flow structure in such a cavity consists in one or two spiraling motions. Long-term observations of such structures with the help of tracers (small particles) indicated that accumulation of the particles may set in at some flow regions. For theoretical insight into the phenomenon, a simple analytical model of recirculating (rotating) flow was studied. It was assumed that particles are spherical and rigid, and their presence does not affect the flow field. The particle Reynolds number is negligibly small, hence only the effects of particle–liquid density difference are of importance. Besides buoyancy, the effects of Saffman’s force and the inertial forces are also taken into account when calculating particle trajectories. Both cases were analyzed, particles with density slightly higher and lower than the fluid. It was found that in our case the inertial forces are egligible. In the numerical experiment trajectories of particles were investigated. The particles were allocated at random in the flow field obtained by numerical solution of the natural convection in the differentially heated cavity. In the experimental part, behavior of a dilute particle suspension in the convective cell was explored. In the model-analytical study of a simple spiraling motion, it was found that due to the interaction of the recirculating convective flow field and the gravity-buoyancy force, the particles may be trapped in some flow regions, whereas the rest of the flow field becomes particle-free. This prediction agrees fairly well with the numerical and experimental findings.
Spatio-temporal dynamics of forced periodic flows in a confined domain
Authors: Nakamura Yukio
Published: July 2002

Nonlinear dynamics of a localized linear array of vortices is investigated by numerical simulations. The setup consists of a thin fluid layer (electrolyte) enclosed in a rectangular box and driven by the injection of homogeneous electric currents in an alternating magnetic field. The spectral model simulates the Navier–Stokes equations in two dimensions with steady forcing and linear bottom friction. The model provides an accurate representation of the evolution of flow pattern. Fourier decomposition of the streamfunction shows that a subharmonic instability occurs in the same symmetry subspace as that of a basic flow in the primary instability regime and a shear flow mode appears in a different symmetry subspace in the secondary instability regime. The exploration of temporal behavior shows that the system produces Hopf-type bifurcations. Chaos and frequency locking due to the mutual interaction between the unstable modes are observed. A general scenario of the dynamics of forced periodic flows is discussed by the amplitude equations, which are modeled using formal group theoretical techniques.
Derivation and investigation of a new explicit algebraic model for the passive scalar flux
Authors: Wikström P. M. , Wallin S. , Johansson A. V.
Published: July 2002

An algebraic relation for the scalar flux, in terms of mean flow quantities, is formed by applying an equilibrium condition in the transport equations for the normalized scalar flux. This modeling approach is analogous to explicit algebraic Reynolds stress modeling (EARSM) for the Reynolds stress anisotropies. The assumption of negligible advection and diffusion of the normalized passive scalar flux gives, in general, an implicit, nonlinear set of algebraic equations. A method to solve this implicit relation in a fully explicit form is proposed, where the nonlinearity in the scalar-production-to-dissipation ratio is considered and solved. The nonlinearity, in the algebraic equations for the normalized scalar fluxes, may be eliminated directly by using a nonlinear term in the model of the pressure scalar-gradient correlation and the destruction and thus results in a much simpler model for both two-and three-dimensional mean flows. The performance of the present model is investigated in three different flow situations. These are homogeneous shear flow with an imposed mean scalar gradient, turbulent channel flow, and the flow field downstream a heated cylinder. The direct numerical simulation (DNS) data are used to analyze the passive scalar flux in the homogeneous shear and channel flow cases and experimental data are used in the case of the heated cylinder wake. Sets of parameter values giving very good predictions in all three cases are found.
Magnetron theory
Authors: Riyopoulos Spilios
Published: July 2002

A guiding center fluid theory is applied to model steady-state, single mode, high-power magnetron operation. A hub of uniform, prescribed density, feeds the current spokes. The spoke charge follows from the continuity equation and the incompressibility of the guiding center flow. Included are the spoke self-fields (DC and AC), obtained by an expansion around the unperturbed (zero-spoke charge) flow in powers of ν/V1, ν, and V1 being the effective charge density and AC amplitude. The spoke current is obtained as a nonlinear function of the detuning from the synchronous (Buneman–Hartree, BH) voltage Vs; the spoke charge is included in the self-consistent definition of Vs. It is shown that there is a DC voltage region of width ‖V−Vs‖∼V1, where the spoke width is constant and the spoke current is simply proportional to the AC voltage. The magnetron characteristic curves are ‘‘flat’’ in that range, and are approximated by a linear expansion around Vs. The derived formulas differ from earlier results [J. F. Hull, in Cross Field Microwave Devices, edited by E. Okress (Academic, New York, 1961), pp. 496–527] in (a) there is no current cutoff at synchronism; the tube operates well below as well above the BH voltage; (b) the characteristics are single valued within the synchronous voltage range; (c) the hub top is not treated as virtual cathode; and (d) the hub density is not equal to the Brillouin density; comparisons with tube measurements show the best agreement for hub density near half the Brillouin density. It is also shown that at low space charge and low power the gain curve is symmetric relative to the voltage (frequency) detuning. While symmetry is broken at high-power/high space charge magnetron operation, the BH voltage remains between the current cutoff voltages.
Feedback equations for the wall modes of a rotating plasma
Authors: Boozer Allen H.
Published: July 2002

Important magnetohydrodynamic instabilities, called wall modes, have growth rates that are determined by the resistivity of the wall that surrounds a toroidal plasma. Wall modes can be feedback stabilized, but plasma rotation modifies the theory of feedback stabilization and can even stabilize the modes. This paper formulates the wall mode problem in a rotating plasma in a manner that can be incorporated in feedback stabilization codes. The formulation utilizes the concept of a permeability, which relates the normal magnetic field on the surface of the plasma to the normal field produced by external circuits. The permeability provides all of the information about the plasma that is required to calculate the stability and the feedback stabilization of wall modes.
X-ray photoelectron spectroscopy study of the effects of ultrapure water on GaAs
Authors: Massies J. , Contour J. P.
Published: July 2002

X-ray photoelectron spectroscopy has been used to investigate the effects of de-ionized water on chemical etched GaAs surfaces. When the treatment with water is performed in static conditions (stagnant water) a Ga-rich oxide layer is formed on GaAs at the rate of 10–20 Å h−1. In contrast, when the GaAs surface is treated in dynamic conditions (running water), no oxide buildup is observed. Moreover, running water can remove the oxide film formed in static conditions, as well as oxidized layers due to air exposure. These results are discussed in the framework of cleaning prior to molecular beam epitaxy.
Cycling of defects between trapped negative charge and interface states at the Si-SiO2 interface
Authors: Sung J. M. , Lyon S. A.
Published: July 2002

We have investigated the states produced at the Si-SiO2 interface by Fowler–Nordheim tunneling and by UV photoinjection at low temperature (90 K). After the interface states are generated, subsequent application of a negative gate bias (5–6 MV/cm) converts them entirely into trapped negative charge. The reverse conversion into interface states occurs if a positive gate bias (5–6 MV/cm) is applied. Cycling of the defects between interface states and trapped negative charge is observed in both experiments. The data strongly suggest that both low-temperature Fowler–Nordheim tunneling and UV photoinjection generate essentially the same structural defect at the interface between Si and SiO2.
Subpicosecond dynamics of electron injection into GaAs/AlGaAs quantum wells
Authors: Goodnick S. M. , Lugli P.
Published: July 2002

We model the subpicosecond evolution of a nonthermal electron distribution injected into a GaAs/AlGaAs quantum well using an ensemble Monte Carlo simulation which includes electron-electron scattering. The calculated results are in good agreement with the experimental time dependence of the carrier distribution function from recent bleaching experiments with carrier-carrier scattering the dominant mechanism contributing to band filling.
Fiber-optic evanescent field absorption sensor
Authors: Paul Phillip H. , Kychakoff George
Published: July 2002

Waveguide evanescent field absorption has been measured as a function of cladding refractive index and bulk absorption coefficient. A simple theory for interpreting these results has been developed. An all-fiber species concentration sensor has been constructed and its performance characterized.
Determination of the critical layer thickness of Si1−xGex/Si heterostructures by direct observation of misfit dislocations
Authors: Kohama Y. , Fukuda Y. , Seki M.
Published: July 2002

The dependence of the critical layer thickness hc on mole fraction x of Si1−xGex/Si heterostructures is determined by direct observations of misfit dislocations by using the electron beam induced current (EBIC) technique and transmission electron microscopy (TEM). The EBIC images of the Si1−xGex/Si interfaces show square-grid patterns running in the 〈110〉 directions. These patterns are identified by TEM observation to be misfit dislocations generated at the Si1−xGex/Si interface. The dependence of hc on x is compared with the results reported by R. People and J. C. Bean [Appl. Phys. Lett. 47, 322 (1985) and 49, 229 (1986)]. The comparison reveals that inconsistency exists between them, especially in the range of x<0.3. This inconsistency is considered to arise from the difference in the techniques adopted to determine the hc’s.
Directed evolution of the site specificity of Cre recombinase
Authors: Santoro Stephen W. , Schultz Peter G.
Published: July 2002

Cre recombinase from bacteriophage P1 recognizes a 34-bp recombination site,
loxP
, with exquisite sequence specificity and catalyzes the site-specific insertion, excision, or rearrangement of DNA. To better understand the molecular basis of protein–DNA recognition and generate recombinases with altered specificities, we have developed a directed evolution strategy that can be used to identify recombinases that recognize variant
loxP
sites. To be selected, members of a library of Cre variants produced by targeted random mutagenesis must rapidly catalyze recombination,
in vivo
, between two variant
loxP
sites that are located on a reporter plasmid. Recombination results in an altered pattern of fluorescent protein expression that can be identified by flow cytometry. Fluorescence-activated cell sorting can be used either to screen positively for recombinase variants that recognize a novel
loxP
site, or negatively for variants that cannot recognize the wild-type
loxP
site. The use of positive screening alone resulted in a relaxation of recombination site specificity, whereas a combination of positive and negative screening resulted in a switching of specificity. One of the identified recombinases selectively recombines a novel recombination site and operates at a rate identical to that of wild-type Cre. Analysis of the sequences of the resulting Cre variants provides insight into the evolution of these altered specificities. This and other systems should contribute to our understanding of protein–DNA recognition and may eventually be used to evolve custom-tailored recombinases that can be used for gene study and inactivation.
Femtosecond resolution of ligand-heme interactions in the high-affinity quinol oxidase
bd
: A di-heme active site?
Authors: Vos Marten H. , Borisov Vitaliy B. , Liebl Ursula , Martin Jean-Louis , Konstantinov Alexander A.
Published: July 2002

Interaction of the two high-spin hemes in the oxygen reduction site
of the
bd
-type quinol oxidase from
Escherichia
coli
has been studied by femtosecond multicolor transient
absorption spectroscopy. The previously unidentified Soret band of
ferrous heme
b
595
was determined to be
centered around 440 nm by selective excitation of the fully reduced
unliganded or CO-bound cytochrome
bd
in the α-band of
heme
b
595
. The redox state of the
b
-type hemes strongly affects both the line shape and
the kinetics of the absorption changes induced by photodissociation of
CO from heme
d
. In the reduced enzyme, CO
photodissociation from heme
d
perturbs the spectrum of
ferrous cytochrome
b
595
within a few ps,
pointing to a direct interaction between hemes
b
595
and
d
. Whereas in the
reduced enzyme no heme
d
-CO geminate recombination is
observed, in the mixed-valence CO-liganded complex with heme
b
595
initially oxidized, a significant part
of photodissociated CO does not leave the protein and recombines with
heme
d
within a few hundred ps. This caging effect may
indicate that ferrous heme
b
595
provides a
transient binding site for carbon monoxide within one of the routes by
which the dissociated ligand leaves the protein. Taken together, the
data indicate physical proximity of the hemes
d
and
b
595
and corroborate the possibility of a
functional cooperation between the two hemes in the dioxygen-reducing
center of cytochrome
bd
.
Concurrent MRI and diffuse optical tomography of breast after indocyanine green enhancement
Authors: Ntziachristos Vasilis , Yodh A. G. , Schnall Mitchell , Chance Britton
Published: July 2002

We present quantitative optical images of human breastin vivo. The images were obtained by using near-infrared diffuse optical tomography (DOT) after the administration of indocyanine green (ICG) for contrast enhancement. The optical examination was performed concurrently with a magnetic resonance imaging (MRI) exam on patients scheduled for excisional biopsy or surgery so that accurate image coregistration and histopathological information of the suspicious lesions was available. The ICG-enhanced optical images coregistered accurately with Gadolinium-enhanced magnetic resonance images validating the ability of DOT to image breast tissue. In contrast to simple transillumination, we found that DOT provides for localization and quantification of exogenous tissue chromophore concentrations. Additionally our use of ICG, an albumin bound absorbing dye in plasma, demonstrates the potential to differentiate disease based on the quantified enhancement of suspicious lesions.
Interactions between double positive thymocytes and high affinity ligands presented by cortical epithelial cells generate double negative thymocytes with T cell regulatory activity
Authors: Wang Rongfang , Wang-Zhu Yiran , Grey Howard
Published: July 2002

Previous studies on thymocyte differentiation by using reaggregate cultures (RC) of double positive T cell receptor (TCR) transgenic thymocytes and the thymic epithelial cell line ANV indicated that low concentrations of high affinity ligands for the TCR were efficient inducers of thymocyte maturation to CD4 single positive (SP) functional cells. In this study, it is demonstrated that, when high concentrations of high affinity ligands are used in this RC system, double positive (DP) cells down-modulate expression of both coreceptors and that, as a result, large numbers of double negative (DN) cells are generated. These DN cells proliferated modestly in response to stimulation by antigen, and this response was considerably augmented by the addition of IL-2 to the cultures. Notably, these antigen-stimulated DN cells produced large amounts of IL-10. When the DN cells generated in RC were cocultured with naive TCR transgenic T cells in the presence of antigen, they suppressed the proliferative response of the naive T cells. Thus, high affinity ligands, when presented to DP thymocytes by cortical thymic epithelial cells in reaggregate cultures, rather than causing deletion of the immature thymocytes, induce their differentiation into immunoregulatory DN cells, suggesting a distinct mechanism by which self tolerance may be maintained.
Activation of latent Kaposi's sarcoma-associated herpesvirus by demethylation of the promoter of the lytic transactivator
Authors: Chen Jiguo , Ueda Keiji , Sakakibara Shuhei , Okuno Toshiomi , Parravicini Carlo , Corbellino Mario , Yamanishi Koichi
Published: July 2002

Kaposi's sarcoma-associated herpesvirus (KSHV) is strongly linked
to Kaposi's sarcoma, primary effusion lymphomas, and a subset of
multicentric Castleman's disease. The mechanism by which this virus
establishes latency and reactivation is unknown. KSHV Lyta (lytic
transactivator, also named KSHV/Rta), mainly encoded by the ORF 50
gene, is a lytic switch gene for viral reactivation from latency,
inasmuch as it is both essential and sufficient to drive the entire
viral lytic cycle. Here we show that the Lyta promoter region was
heavily methylated in latently infected cells. Treatment of primary
effusion lymphoma-delivered cell lines with tetradecanoylphorbol
acetate caused demethylation of the Lyta promoter and induced KSHV
lytic phase
in vitro
. Methylation cassette assay shows
demethylation of the Lyta promoter region was essential for the
expression of Lyta.
In vivo
, biopsy samples obtained
from patients with KSHV-related diseases show the most demethylation in
the Lyta promoter region, whereas samples from a latently infected KSHV
carrier remained in a methylated status. These results suggest a
relationship among a demethylation status in the Lyta promoter, the
reactivation of KSHV, and the development of KSHV-associated diseases.
Neuroprotection in cerebral ischemia by neutralization of 3-aminopropanal
Authors: Ivanova Svetlana , Batliwalla Franak , Mocco J. , Kiss Szilard , Huang Judy , Mack William , Coon Alexander , Eaton John W. , Al-Abed Yousef , Gregersen Peter K. , Shohami Esther , Connolly E. Sander , Tracey Kevin J.
Published: July 2002

Cerebral ischemia stimulates increased activity of polyamine oxidase, a ubiquitous enzyme that catabolizes polyamines to produce 3-aminopropanal. 3-Aminopropanal is a reactive aldehyde that mediates progressive neuronal necrosis and glial apoptosis. Here we report that increased levels of 3-aminopropanal-modified protein levels in humans after aneurysmal subarachnoid hemorrhage correlate with the degree of cerebral injury as measured by admission Hunt/Hess grade.
In vitro
screening of clinically approved drugs reveals that
N
-2-mercaptopropionyl glycine (
N
-2-MPG), an agent clinically approved for prevention of renal stones in patients with cysteinuria, significantly inhibits the cytotoxicity of 3-aminopropanal.
N
-2-MPG reacts with 3-aminopropanal to yield a nontoxic thioacetal adduct, as confirmed by electrospray ionization mass spectroscopy. Administration of
N
-2-MPG in clinically relevant doses to rats significantly reduces cerebral 3-aminopropanal-modified protein immunoreactivity and infarct volume in a standardized model of middle cerebral artery occlusion, even when the agent is administered after the onset of ischemia. These results implicate 3-aminopropanal as a therapeutic target for cerebral ischemia.
Probing alkali metal–π interactions with the side chain residue of tryptophan
Authors: Hu Jiaxin , Barbour Leonard J. , Gokel George W.
Published: July 2002

Feeble forces play a significant role in the organization of proteins. These include hydrogen bonding, hydrophobic interactions, salt bridge formation, and steric interactions. The alkali metal cation-π interaction is a force of potentially profound importance but its consideration in biology has been limited by the lack of experimental evidence. Our previous studies of cation–π interactions with Na
+
and K
+
involved the side arms of tryptophan (indole), tyrosine (phenol), and phenylalanine (benzene) as the arene donors. The receptor system possesses limiting steric constraints. In this report, we show that direct interactions between alkali metals and arenes occur at or within the van der Waals contact distance.
Cyclin A activates the DNA polymerase δ-dependent elongation machinery
in vitro
: A parvovirus DNA replication model
Authors: Bashir Tarig , Hörlein Rita , Rommelaere Jean , Willwand Kurt
Published: July 2002

Replication of the single-stranded linear DNA genome of parvovirus minute virus of mice (MVM) starts with complementary strand synthesis from the 3′-terminal snap-back telomere, which serves as a primer for the formation of double-stranded replicative form (RF) DNA. This DNA elongation reaction, designated conversion, is exclusively dependent on cellular factors. In cell extracts, we found that complementary strand synthesis was inhibited by the cyclin-dependent kinase inhibitor p21
WAF1/CIP1
and rescued by the addition of proliferating cell nuclear antigen, arguing for the involvement of DNA polymerase (Pol) δ in the conversion reaction.
In vivo
time course analyses using synchronized MVM-infected A9 cells allowed initial detection of MVM RF DNA at the G
1
/S phase transition, coinciding with the onset of cyclin A expression and cyclin A-associated kinase activity. Under
in vitro
conditions, formation of RF DNA was efficiently supported by A9 S cell extracts, but only marginally by G
1
cell extracts. Addition of recombinant cyclin A stimulated DNA conversion in G
1
cell extracts, and correlated with a concomitant increase in cyclin A-associated kinase activity. Conversely, a specific antibody neutralizing cyclin A-dependent kinase activity, abolished the capacity of S cell extracts for DNA conversion. We found no evidence for the involvement of cyclin E in the regulation of the conversion reaction. We conclude that cyclin A is necessary for activation of complementary strand synthesis, which we propose as a model reaction to study the cell cycle regulation of the Pol δ-dependent elongation machinery.
Desmuslin, an intermediate filament protein that interacts with α-dystrobrevin and desmin
Authors: Mizuno Yuji , Thompson Terri G. , Guyon Jeffrey R. , Lidov Hart G. W. , Brosius Melissa , Imamura Michihiro , Ozawa Eijiro , Watkins Simon C. , Kunkel Louis M.
Published: July 2002

Dystrobrevin is a component of the dystrophin-associated protein
complex and has been shown to interact directly with dystrophin,
α1-syntrophin, and the sarcoglycan complex. The precise role of
α-dystrobrevin in skeletal muscle has not yet been determined. To
study α-dystrobrevin's function in skeletal muscle, we used the
yeast two-hybrid approach to look for interacting proteins. Three
overlapping clones were identified that encoded an intermediate
filament protein we subsequently named desmuslin (DMN). Sequence
analysis revealed that DMN has a short N-terminal domain, a conserved
rod domain, and a long C-terminal domain, all common features of type 6
intermediate filament proteins. A positive interaction between DMN and
α-dystrobrevin was confirmed with an
in vitro
coimmunoprecipitation assay. By Northern blot analysis, we find that
DMN is expressed mainly in heart and skeletal muscle, although there is
some expression in brain. Western blotting detected a 160-kDa protein
in heart and skeletal muscle. Immunofluorescent microscopy localizes
DMN in a stripe-like pattern in longitudinal sections and in a mosaic
pattern in cross sections of skeletal muscle. Electron microscopic
analysis shows DMN colocalized with desmin at the Z-lines. Subsequent
coimmunoprecipitation experiments confirmed an interaction with desmin.
Our findings suggest that DMN may serve as a direct linkage between the
extracellular matrix and the Z-discs (through plectin) and may play an
important role in maintaining muscle cell integrity.
Dual HLA class I and class II restricted recognition of alloreactive T lymphocytes mediated by a single T cell receptor complex
Authors: Heemskerk Mirjam H. M. , de Paus Roelof A. , Lurvink Ellie G. A. , Koning Frits , Mulder Arend , Willemze Roel , van Rood Jon J. , Falkenburg J. H. Frederik
Published: July 2002

The alloreactive human T cell clone MBM15 was found to exhibit dual specificity recognizing both an antigen in the context of the HLA class I A2 molecule and an antigen in the context of the HLA class II DR1. We demonstrated that the dual reactivity that was mediated via a single clonal T cell population depended on specific peptide binding. For complete recognition of the HLA-A2-restricted specificity the interaction of CD8 with HLA class I is essential. Interestingly, interaction of the CD8 molecule with HLA class I contributed to the HLA-DR1-restricted specificity. T cell clone MBM15 expressed two in-frame T cell receptor (TCR) Vα transcripts (Vα1 and Vα2) and one TCR Vβ transcript (Vβ13). To elucidate whether two TCR complexes were responsible for the dual recognition or one complex, cytotoxic T cells were transduced with retroviral vectors encoding the different TCR chains. Only T cells transduced with the TCR Vα1Vβ13 combination specifically recognized both the HLA-A2+and HLA-DR1+target cells, whereas the Vα2Vβ13 combination did not result in a TCR on the cell surface. Thus a single TCRαβ complex can have dual specificity, recognizing both a peptide in the context of HLA class I as well as a peptide in the context of HLA class II. Transactivation of T cells by an unrelated antigen in the context of HLA class II may evoke an HLA class I-specific T cell response. We propose that this finding may have major implications for immunotherapeutic interventions and insight into the development of autoimmune diseases.
Constitutive telomerase expression promotes mammary carcinomas in aging mice
Authors: Artandi Steven E. , Alson Scott , Tietze Maja K. , Sharpless Norman E. , Ye Siqin , Greenberg Roger A. , Castrillon Diego H. , Horner James W. , Weiler Sarah R. , Carrasco Ruben D. , DePinho Ronald A.
Published: July 2002

Telomerase is up-regulated in the vast majority of human cancers and serves to halt the progressive telomere shortening that ultimately blocks would-be cancer cells from achieving a full malignant phenotype. In contrast to humans, the laboratory mouse possesses long telomeres and, even in early generation telomerase-deficient mice, the level of telomere reserve is sufficient to avert telomere-based checkpoint responses and to permit full malignant progression. These features in the mouse provide an opportunity to determine whether enforced high-level telomerase activity can serve functions that extend beyond its ability to sustain telomere length and function. Here, we report the generation and characterization of transgenic mice that express the catalytic subunit of telomerase (mTERT) at high levels in a broad variety of tissues. Expression of mTERT conferred increased telomerase enzymatic activity in several tissues, including mammary gland, splenocytes, and cultured mouse embryonic fibroblasts. In mouse embryonic fibroblasts, mTERT overexpression extended telomere lengths but did not prevent culture-induced replicative arrest, thus reinforcing the view that this phenomenon is not related to occult telomere shortening. Robust telomerase activity, however, was associated with the spontaneous development of mammary intraepithelial neoplasia and invasive mammary carcinomas in a significant proportion of aged females. These data indicate that enforced mTERT expression can promote the development of spontaneous cancers even in the setting of ample telomere reserve.
Global modulation of cellular transcription by human cytomegalovirus is initiated by viral glycoprotein B
Authors: Simmen Kenneth A. , Singh Jasbir , Luukkonen B. G. Mattias , Lopper Matt , Bittner Anton , Miller Nancy E. , Jackson Michael R. , Compton Teresa , Früh Klaus
Published: July 2002

Human cytomegalovirus (HCMV) infection alters the expression of many cellular genes, including IFN-stimulated genes (ISGs) [Zhu, H., Cong, J.-P., Mamtora, G., Gingeras, T. & Shenk, T. (1998)Proc. Natl. Acad. Sci. USA95, 14470–14475]. By using high-density cDNA microarrays, we show that the HCMV-regulated gene expression profile in fibroblasts does not differ substantially from the response generated by IFN. Furthermore, we identified the specific viral component triggering this response as the envelope glycoprotein B (gB). Cells treated with gB, but not other herpesviral glycoproteins, exhibited the same transcriptional profile as HCMV-infected cells. Thus, the interaction of gB with its as yet unidentified cellular receptor is the principal mechanism by which HCMV alters cellular gene expression early during infection. These findings highlight a pioneering paradigm for the consequences of virus–receptor interactions.
Screening insertion libraries for mutations in many genes simultaneously using DNA microarrays
Authors: Mahalingam Ramamurthy , Fedoroff Nina
Published: July 2002

We describe a method to screen pools of DNA from multiple transposon lines for insertions in many genes simultaneously. We use thermal asymmetric interlaced–PCR, a hemispecific PCR amplification protocol that combines nested, insertion-specific primers with degenerate primers, to amplify DNA flanking the transposons. In reconstruction experiments with previously characterized
Arabidopsis
lines carrying insertions of the maize
Dissociation
(
Ds
) transposon, we show that fluorescently labeled, transposon-flanking fragments overlapping ORFs hybridize to cognate expressed sequence tags (ESTs) on a DNA microarray. We further show that insertions can be detected in DNA pools from as many as 100 plants representing different transposon lines and that all of the tested, transposon-disrupted genes whose flanking fragments can be amplified individually also can be detected when amplified from the pool. The ability of a transposon-flanking fragment to hybridize declines rapidly with decreasing homology to the spotted DNA fragment, so that only ESTs with >90% homology to the transposon-disrupted gene exhibit significant cross-hybridization. Because thermal asymmetric interlaced–PCR fragments tend to be short, use of the present method favors recovery of insertions in and near genes. We apply the technique to screening pools of new
Ds
lines using cDNA microarrays containing ESTs for ≈1,000 stress-induced and -repressed
Arabidopsis
genes.
Lack of hepcidin gene expression and severe tissue iron overload in upstream stimulatory factor 2 (
USF2
) knockout mice
Authors: Nicolas Gaël , Bennoun Myriam , Devaux Isabelle , Beaumont Carole , Grandchamp Bernard , Kahn Axel , Vaulont Sophie
Published: July 2002

We previously reported the disruption of the murine gene encoding the transcription factor USF2 and its consequences on glucose-dependent gene regulation in the liver. We report here a peculiar phenotype of
Usf2
−/−
mice that progressively develop multivisceral iron overload; plasma iron overcomes transferrin binding capacity, and nontransferrin-bound iron accumulates in various tissues including pancreas and heart. In contrast, the splenic iron content is strikingly lower in knockout animals than in controls. To identify genes that may account for the abnormalities of iron homeostasis in
Usf2
−/−
mice, we used suppressive subtractive hybridization between livers from
Usf2
−/−
and wild-type mice. We isolated a cDNA encoding a peptide, hepcidin (also referred to as LEAP-1, for liver-expressed antimicrobial peptide), that was very recently purified from human blood ultrafiltrate and from urine as a disulfide-bonded peptide exhibiting antimicrobial activity. Accumulation of iron in the liver has been recently reported to up-regulate hepcidin expression, whereas our data clearly show that a complete defect in hepcidin expression is responsible for progressive tissue iron overload. The striking similarity of the alterations in iron metabolism between
HFE
knockout mice, a murine model of hereditary hemochromatosis, and the
Usf2
−/−
hepcidin-deficient mice suggests that hepcidin may function in the same regulatory pathway as HFE. We propose that hepcidin acts as a signaling molecule that is required in conjunction with HFE to regulate both intestinal iron absorption and iron storage in macrophages.
Peroxynitrite does not decompose to singlet oxygen (
1
Δ
g
O
2
) and nitroxyl (NO
−
)
Authors: Martinez Glaucia R. , Di Mascio Paolo , Bonini Marcelo G. , Augusto Ohara , Briviba Karlis , Sies Helmut , Maurer Patrick , Röthlisberger Ursula , Herold Susanna , Koppenol Willem H.
Published: July 2002

According to Khan
et al.
[Khan, A. U.,
Kovacic, D., Kolbanovskiy, A., Desai, M., Frenkel, K. & Geacintov,
N. E. (2000)
Proc. Natl. Acad. Sci. USA
97,
2984–2989], peroxynitrite (ONOO
−
) decomposes after
protonation to singlet oxygen
(
1
Δ
g
O
2
) and singlet oxonitrate
(nitroxyl,
1
NO
−
) in high yield. They claimed
to have observed nitrosyl hemoglobin from the reaction of
NO
−
with methemoglobin; however, contamination with
hydrogen peroxide gave rise to ferryl hemoglobin, the spectrum of which
was mistakenly assigned to nitrosyl hemoglobin. We have carried out
UV–visible and EPR experiments with methemoglobin and hydrogen
peroxide-free peroxynitrite and find that no NO
−
is
formed. With this peroxynitrite preparation, no light emission from
singlet oxygen at 1270 nm is observed, nor is singlet oxygen chemically
trapped; however, singlet oxygen was trapped when hydrogen peroxide was
also present, as previously described [Di Mascio, P., Bechara, E.
J. H., Medeiros, M. H. G., Briviba, K. & Sies, H. (1994)
FEBS Lett.
355, 287–289]. Quantum mechanical and
thermodynamic calculations show that formation of the postulated
intermediate, a cyclic form of peroxynitrous acid (trioxazetidine), and
the products
1
NO
−
and
1
Δ
g
O
2
requires Gibbs energies of
ca.
+415 kJ⋅mol
−1
and
ca.
+180 kJ⋅mol
−1
, respectively. Our
results show that the results of Khan
et al.
are best
explained by interference from contaminating hydrogen peroxide left
from the synthesis of peroxynitrite.
SUMO-1 modification required for transformation by adenovirus type 5 early region 1B 55-kDa oncoprotein
Authors: Endter Christian , Kzhyshkowska Julia , Stauber Roland , Dobner Thomas
Published: July 2002

SUMO-1 is a small ubiquitin-related modifier protein that is
covalently linked to many cellular and viral protein targets.
Modification by SUMO-1 is proposed to play a role in protein targeting
and/or stability. We show here that adenovirus type 5 early region 1B
55-kDa (E1B-55kDa) oncoprotein can be covalently modified by SUMO-1
in vivo
through a major attachment site comprising a
single lysine residue at amino acid position 104. The sequence
surrounding this lysine matches the proposed ΨKxE consensus motif
required for SUMO-1 conjugation. A single mutation (K104R) that
abolishes SUMOylation of E1B-55kDa dramatically reduces the ability of
the adenovirus type 5 protein to transform primary baby rat kidney
cells in cooperation with E1A and to inhibit p53-mediated
transactivation. Overexpression of SUMO-1 in adenovirus type 5
E1A/E1B-55kDa-transformed baby rat kidney cells causes the
relocalization of E1B-55kDa from the cytoplasm to the nucleus, where it
accumulates with SUMO-1 in dot- or track-like structures.
Significantly, when SUMO-1 is ectopically expressed in transformed rat
cells no effect on the cytoplasmic localization of the E1B-K104R mutant
protein is observed. Our results demonstrate that SUMO-1 modification
is required for transformation by adenovirus type 5 E1B-55kDa and
provide further evidence for the idea that this posttranslational
modification plays a role in protein targeting to specific subcellular
sites.