bluuebunny/crossref_metadata_2025 huggingface dataset:
Study of the consequence of excess indium in the active channel of InGaAs/InAlAs high electron mobility transistors on device properties
Authors: Ng G. I. , Pavlidis D. , Quillec M. , Chan Y. J. , Jaffe M. D. , Singh J.
Published: July 2002

A study of the properties of In0.52Al0.48As/In0.53+xGa0.47−xAs high electron mobility transistors is carried out for 0%, 7%, and 12% excess In values in the channel. Theoretical analysis shows that the enhanced In causes a biaxial compressive strain of 0.49% to 0.84% in the channel, increases the band-edge discontinuity from 0.437 to 0.500 eV, and reduces the carrier mass by 6%. Experimental characterizations support the theoretical predictions by demonstrating an increase of mobility from 9900 to 11 200 cm2/V s at 300 K, and a transconductance enhancement from 160 to at least 230 mS/mm.
Abietadiene synthase catalysis: Mutational analysis of a prenyl diphosphate ionization-initiated cyclization and rearrangement
Authors: Peters Reuben J. , Croteau Rodney B.
Published: July 2002

Abietadiene synthase catalyzes the committed step in resin acid biosynthesis, forming a mixture of abietadiene double-bond isomers by two sequential, mechanistically distinct cyclizations at separate active sites. The first reaction, protonation-initiated cyclization, converts the universal diterpene precursor geranylgeranyl diphosphate to the stable bicyclic intermediate copalyl diphosphate. In the second, magnesium ion-dependent reaction, diphosphate ester ionization-initiated cyclization generates the tricyclic perhydrophenanthrene-type backbone and is coupled, by intramolecular proton transfer within a transient pimarenyl intermediate, to a 1,2-methyl migration that generates the C13 isopropyl group characteristic of the abietane structure. Alternative deprotonations of the terminal abietenyl carbocation provide a mixture of abietadiene, levopimaradiene, and neoabietadiene, and this product profile varies as a function of pH. Mutational analysis of amino acids at the active site of a modeled structure has identified residues critical for catalysis, as well as several that play roles in specifying product formation, apparently by ligation of a magnesium ion cofactor. These results strongly suggest that choice between alternatives for deprotonation of the abietenyl intermediate depends more on the positioning effects of the carbocation–diphosphate anion reaction partners than on the pKa of multiple participating bases. In one extreme case, mutant N765A is unable to mediate the intramolecular proton transfer and aborts the reaction, without catalyzing 1,2-methyl migration, to produce only sandaracopimaradiene, thereby providing supporting evidence for the corresponding stereochemistry of the cryptic pimarenyl intermediate of the reaction pathway.
A comparison of experimental, theoretical, and numerical simulation Rayleigh–Taylor mixing rates
Authors: George E. , Glimm J. , Li X.-L. , Marchese A. , Xu Z.-L.
Published: July 2002

We present a Rayleigh–Taylor mixing rate simulation with an acceleration rate falling within the range of experiments. The simulation uses front tracking to prevent interfacial mass diffusion. We present evidence to support the assertion that the lower acceleration rate found in untracked simulations is caused, at least to a large extent, by a reduced buoyancy force due to numerical interfacial mass diffusion. Quantitative evidence includes results from a time-dependent Atwood number analysis of the diffusive simulation, which yields a renormalized mixing rate coefficient for the diffusive simulation in agreement with experiment.
Notch
signaling and the determination of appendage identity
Authors: Kurata Shoichiro , Go Masahiro J. , Artavanis-Tsakonas Spyros , Gehring Walter J.
Published: July 2002

The
Notch
signaling pathway defines an evolutionarily conserved cell–cell interaction mechanism that throughout development controls the ability of precursor cells to respond to developmental signals. Here we show that
Notch
signaling regulates the expression of the master control genes
eyeless
,
vestigial
, and
Distal-less
, which in combination with homeotic genes induce the formation of eyes, wings, antennae, and legs. Therefore,
Notch
is involved in a common regulatory pathway for the determination of the various
Drosophila
appendages.
On the interpretation of quantitative structure–function activity relationship data for lactate oxidase
Authors: Yorita Kazuko , Misaki Hideo , Palfey Bruce A. , Massey Vincent
Published: July 2002

The native flavin, FMN, has been removed from the
l
-lactate oxidase of
Aerococcus viridans
, and
the apoprotein reconstituted with 12 FMN derivatives with various
substituents at the flavin 6- and 8-positions. Impressive linear
relationships are exhibited between the sum of the Hammett
σ
para
and σ
ortho
parameters and the redox
potentials of the free flavins, and between the redox potentials of the
free and enzyme-bound flavins. Rapid reaction kinetics studies of the
reconstituted enzymes with the substrates
l
-lactate and
l
-mandelate show an increase in the reduction rate constant
with increasing redox potential, except that, with lactate, a limiting
rate constant of ≈700 s
−1
is obtained with flavins of
high potential. Similar breakpoints are found in plots of the rate
constants for flavin N5-sulfite adduct formation and for the reaction
of the reduced enzymes with molecular oxygen. These results are
interpreted in terms of a two-step equilibrium preceding the chemical
reaction step, in which the second equilibrium step provides an upper
limit to the rate with which the particular substrate or ligand is
positioned with the flavin in the correct fashion for the observed
chemical reaction to occur. The relationship of rate constants for
flavin reduction and N5-sulfite adduct formation with flavin redox
potential below the observed breakpoint indicate development of
significant negative charge in the transition states of the reactions.
In the case of reduction by substrate, the results are consistent
either with a hydride transfer mechanism or with the so called
“carbanion” mechanism, in which the substrate α-proton is
abstracted by an enzyme base protected from exchange with solvent.
These conclusions are supported by substrate α-deuterium isotope
effects and by solvent viscosity effects on sulfite binding.
Polar residues drive association of polyleucine transmembrane helices
Authors: Zhou Fang Xiao , Merianos Helen J. , Brunger Axel T. , Engelman Donald M.
Published: July 2002

Although many polar residues are directly involved in transmembrane
protein functions, the extent to which they contribute to more general
structural features is still unclear. Previous studies have
demonstrated that asparagine residues can drive transmembrane helix
association through interhelical hydrogen bonding [Choma, C.,
Gratkowski, H., Lear, J. D. & DeGrado, W. F. (2000)
Nat. Struct. Biol.
7, 161–166; and Zhou, F. X.,
Cocco, M. J., Russ, W. P., Brunger, A. T. & Engelman,
D. M. (2000)
Nat. Struct. Biol.
7, 154–160]. We have
studied the ability of other polar residues to promote helix
association in detergent micelles and in biological membranes. Our
results show that polyleucine sequences with Asn, Asp, Gln, Glu, and
His, residues capable of being simultaneously hydrogen bond donors and
acceptors, form homo- or heterooligomers. In contrast, polyleucine
sequences with Ser, Thr, and Tyr do not associate more than the
polyleucine sequence alone. The results therefore provide experimental
evidence that interactions between polar residues in the helices of
transmembrane proteins may serve to provide structural stability and
oligomerization specificity. Furthermore, such interactions can allow
structural flexibility required for the function of some membrane
proteins.
Tonic nicotinic modulation of serotoninergic transmission in the spinal cord
Authors: Cordero-Erausquin Matilde , Changeux Jean-Pierre
Published: July 2002

The spinal serotoninergic projection from the raphe magnus has been
shown to modulate nociceptive inputs, and activation of this projection
mediates nicotine-elicited analgesia. Here, we investigate the
interactions between cholinergic and serotoninergic systems in the
spinal cord, by conducting serotonin [5-hydroxytryptamine (5-HT)]
efflux experiments on mouse spinal slices. At least three spinal
populations of nicotinic receptors are distinguished that affect 5-HT
release. The first could be directly located on serotoninergic
terminals, is insensitive to nanomolar concentrations of
methyllicaconitine (MLA), and may be subjected to a basal (not maximal)
cholinergic tone. The second is tonically and maximally activated by
endogenous acetylcholine, insensitive to nanomolar
concentrations of MLA, and present on inhibitory neurons. The last is
also present on inhibitory neurons but is sensitive to nanomolar
concentrations of MLA and not tonically activated by acetylcholine.
Multiple nicotinic acetylcholine receptor populations thus
differentially exert tonic or not tonic control on 5-HT transmission in
the spinal cord. These receptors may be major targets for nicotine
effects on antinociception. In addition, the presence of a tonic
nicotinic modulation of 5-HT release indicates that
endogenous acetylcholine plays a role in the physiological
regulation of descending 5-HT pathways to the spinal cord.
Gene elements that affect the longevity of
rbcL
sequence-containing transcripts in
Chlamydomonas reinhardtii
chloroplasts
Authors: Singh Mahipal , Boutanaev Alexander , Zucchi Paola , Bogorad Lawrence
Published: July 2002

The chloroplast gene
rbcL
encodes the large
subunit of the CO
2
-fixing enzyme ribulose-bisphosphate
carboxylase. In previous work a target for photo-accelerated
degradation of
Chlamydomonas reinhardtii rbcL
transcripts
in vivo
was found to lie within the first 63
nucleotides, and a sequence element required for increasing the
longevity of transcripts of
rbcL
-reporter genes was
found to occur between nucleotides 170 and 350. Photo-accelerated
degradation of
rbcL
transcripts has been found to
require nucleotides 21 to 41. Transcript nucleotides lying between 329
and 334 and between 14 and 27 are essential for stabilizing transcripts
in vivo
; mutations in either region reduce the longevity
of transcripts. It is postulated that the effectiveness of
photo-accelerated endonuclease attacks on the nucleotide 21 to 41
region is reduced by physical blockage or distortion of the target
sequence by interacting proteins that associate with nucleotides in the
14 to 27 and 329 to 334 regions of the transcripts. Both the nucleotide
+329 to +334 stabilizing sequence of
rbcL
and a
transcription enhancing sequence that lies between +126 and +170 encode
well conserved (cyanobacteria through angiosperms) amino acid
sequences; the evolution of expression control elements within the
protein coding sequence of
rbcL
is considered.
Supernatant protein factor, which stimulates the conversion of squalene to lanosterol, is a cytosolic squalene transfer protein and enhances cholesterol biosynthesis
Authors: Shibata Norihito , Arita Makoto , Misaki Yuko , Dohmae Naoshi , Takio Koji , Ono Teruo , Inoue Keizo , Arai Hiroyuki
Published: July 2002

Squalene epoxidase, a membrane-associated enzyme that converts
squalene to squalene 2,3-oxide, plays an important role in the
maintenance of cholesterol homeostasis. In 1957, Bloch and colleagues
identified a factor from rat liver cytosol termed “supernatant
protein factor (SPF),” which promotes the squalene epoxidation
catalyzed by rat liver microsomes with oxygen, NADPH, FAD, and
phospholipid [Tchen, T. T. & Bloch, K. (1957)
J.
Biol. Chem.
226, 921–930]. Although purification of SPF by
11,000-fold was reported, no information is so far available on the
primary structure or biological function of SPF. Here we report the
cDNA cloning and expression of SPF from rat and human. The encoded
protein of 403 amino acids belongs to a family of cytosolic
lipid-binding/transfer proteins such as α-tocopherol transfer
protein, cellular retinal binding protein, yeast phosphatidylinositol
transfer protein (Sec14p), and squid retinal binding protein.
Recombinant SPF produced in
Escherichia coli
enhances
microsomal squalene epoxidase activity and promotes intermembrane
transfer of squalene
in vitro
. SPF mRNA is expressed
abundantly in the liver and small intestine, both of which are
important sites of cholesterol biosynthesis. SPF is expressed
significantly in isolated hepatocytes, but the expression level was
markedly decreased after 48 h of
in vitro
culture.
Moreover, SPF was not detectable in most of the cell lines tested,
including HepG2 and McARH7777 hepatomas. Transfection of SPF cDNA
in McARH7777 significantly stimulated
de novo
cholesterol biosynthesis. These data suggest that SPF is a
cytosolic squalene transfer protein capable of regulating
cholesterol biosynthesis.
Erythropoietin prevents motor neuron apoptosis and neurologic disability in experimental spinal cord ischemic injury
Authors: Celik Murat , Gökmen Necati , Erbayraktar Serhat , Akhisaroglu Mustafa , Konakç Selman , Ulukus Cagnur , Genc Sermin , Genc Kursad , Sagiroglu Emel , Cerami Anthony , Brines Michael
Published: July 2002

The cytokine erythropoietin (EPO) possesses potent neuroprotective activity against a variety of potential brain injuries, including transient ischemia and reperfusion. It is currently unknown whether EPO will also ameliorate spinal cord injury. Immunocytochemistry performed using human spinal cord sections showed abundant EPO receptor immunoreactivity of capillaries, especially in white matter, and motor neurons within the ventral horn. We used a transient global spinal ischemia model in rabbits to test whether exogenous EPO can cross the blood–spinal cord barrier and protect these motor neurons. Spinal cord ischemia was produced in rabbits by occlusion of the abdominal aorta for 20 min, followed by saline or recombinant human (rHu)-EPO (350, 800, or 1,000 units/kg of body weight) administered intravenously immediately after the onset of reperfusion. The functional neurological status of animals was better for rHu-EPO-treated animals 1 h after recovery from anesthesia, and improved dramatically over the next 48 h. In contrast, saline-treated animals exhibited a poorer neurological score at 1 h and did not significantly improve. Histopathological examination of the affected spinal cord revealed widespread motor neuron injury associated with positive terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling in control but not in rHu-EPO-treated animals. These observations suggest both an acute as well as a delayed beneficial action of rHu-EPO in ischemic spinal cord injury. Because rHu-EPO is currently used widely with an excellent safety profile, clinical trials evaluating its potential to prevent motor neuron apoptosis and the neurological deficits that occur as a consequence of ischemic injury are warranted.
Ion channel gating: A first-passage time analysis of the Kramers type
Authors: Goychuk Igor , Hänggi Peter
Published: July 2002

The opening rate of voltage-gated potassium ion channels exhibits a characteristic knee-like turnover where the common exponential voltage dependence changes suddenly into a linear one. An explanation of this puzzling crossover is put forward in terms of a stochastic first passage time analysis. The theory predicts that the exponential voltage dependence correlates with the exponential distribution of closed residence times. This feature occurs at large negative voltages when the channel is predominantly closed. In contrast, the linear part of voltage dependence emerges together with a nonexponential distribution of closed dwelling times with increasing voltage, yielding a large opening rate. Depending on the parameter set, the closed-time distribution displays a power law behavior that extends over several decades.
Conformational flexibility of bacterial RNA polymerase
Authors: Darst Seth A. , Opalka Natacha , Chacon Pablo , Polyakov Andrey , Richter Catherine , Zhang Gongyi , Wriggers Willy
Published: July 2002

The structure of
Escherichia coli
core RNA polymerase (RNAP) was determined by cryo-electron microscopy and image processing of helical crystals to a nominal resolution of 15 Å. Because of the high sequence conservation between the core RNAP subunits, we were able to interpret the
E. coli
structure in relation to the high-resolution x-ray structure of
Thermus aquaticus
core RNAP. A very large conformational change of the
T. aquaticus
RNAP x-ray structure, corresponding to opening of the main DNA/RNA channel by nearly 25 Å, was required to fit the
E. coli
map. This finding reveals, at least partially, the range of conformational flexibility of the RNAP, which is likely to have functional implications for the initiation of transcription, where the DNA template must be loaded into the channel.
Vipp1
deletion mutant of
Synechocystis
: A connection between bacterial phage shock and thylakoid biogenesis?
Authors: Westphal Sabine , Heins Lisa , Soll Jürgen , Vothknecht Ute C.
Published: July 2002

Plant chloroplasts originated from an endosymbiotic event by which
an ancestor of contemporary cyanobacteria was engulfed by an early
eukaryotic cell and then transformed into an organelle. Oxygenic
photosynthesis is the specific feature of cyanobacteria and
chloroplasts, and the photosynthetic machinery resides in an internal
membrane system, the thylakoids. The origin and genesis of thylakoid
membranes, which are essential for oxygenic photosynthesis, are still
an enigma. Vipp1 (vesicle-inducing protein in plastids 1) is a protein
located in both the inner envelope and the thylakoids of
Pisum
sativum
and
Arabidopsis thaliana
. In
Arabidopsis
disruption of the
VIPP1
gene
severely affects the plant's ability to form properly structured
thylakoids and as a consequence to carry out photosynthesis. In
contrast, Vipp1 in
Synechocystis
appears to be located
exclusively in the plasma membrane. Yet, as in higher plants,
disruption of the
VIPP1
gene locus leads to the complete
loss of thylakoid formation. So far
VIPP1
genes are
found only in organisms carrying out oxygenic photosynthesis. They
share sequence homology with a subunit encoded by the bacterial phage
shock operon (PspA) but differ from PspA by a C-terminal extension of
about 30 amino acids. In two cyanobacteria,
Synechocystis
and
Anabaena,
both a
VIPP1
and a
pspA
gene are present, and
phylogenetic analysis indicates that
VIPP1
originated
from a gene duplication of the latter and thereafter acquired its new
function. It also appears that the C-terminal extension that
discriminates VIPP1 proteins from PspA is important for its function in
thylakoid formation.
A model of a turbulent boundary layer with a nonzero pressure gradient
Authors: Barenblatt G. I. , Chorin A. J. , Prostokishin V. M.
Published: July 2002

According to a model of the turbulent boundary layer that we propose, in the absence of external turbulence the intermediate region between the viscous sublayer and the external flow consists of two sharply separated self-similar structures. The velocity distribution in these structures is described by two different scaling laws. The mean velocity
u
in the region adjacent to the viscous sublayer is described by the previously obtained Reynolds-number-dependent scaling law
φ = u
/
u
*
=
A
η
α
,
A
= 1/
\documentclass[12pt]{minimal}
\usepackage{amsmath}
\usepackage{wasysym}
\usepackage{amsfonts}
\usepackage{amssymb}
\usepackage{amsbsy}
\usepackage{mathrsfs}
\setlength{\oddsidemargin}{-69pt}
\begin{document}
\begin{equation*}\sqrt{3}\end{equation*}\end{document}
ln
Re
Λ
+ 5/2, α = 3/2 In
Re
Λ
, η = u
*
y
/ν. (Here
u
*
is the dynamic or friction velocity,
y
is the distance from the wall, ν the kinematic viscosity of the fluid, and the Reynolds number
Re
Λ
is well defined by the data.) In the region adjacent to the external flow, the scaling law is different: φ =
B
η
β
. The power β for zero-pressure-gradient boundary layers was found by processing various experimental data and is close (with some scatter) to 0.2. We show here that for nonzero-pressure-gradient boundary layers, the power β is larger than 0.2 in the case of an adverse pressure gradient and less than 0.2 for a favorable pressure gradient. Similarity analysis suggests that both the coefficient
B
and the power β depend on
Re
Λ
and on a new dimensionless parameter
P
proportional to the pressure gradient. Recent experimental data of Perry, Marušić, and Jones were analyzed, and the results are in agreement with the model we propose.
Tracking memory's trace
Authors: Horn Gabriel , Nicol Alister U. , Brown Malcolm W.
Published: July 2002

There is strong converging evidence that the intermediate and
medial part of the hyperstriatum ventrale of the chick brain is a
memory store for information acquired through the learning process of
imprinting. Neurons in this memory system come, through imprinting, to
respond selectively to the imprinting stimulus (IS) neurons and so
possess the properties of a memory trace. Therefore, the responses of
the intermediate and medial part of the hyperstriatum ventrale neurons
to a visual imprinting stimulus were determined before, during, and
after training. Of the total recorded population, the proportions of IS
neurons shortly after each of two 1-h training sessions were
significantly higher (approximately 2 times) than the pretraining
proportion. However, ≈4.5 h later this proportion had fallen
significantly and did not differ significantly from the pretraining
proportion. Nevertheless, ≈21.5 h after the end of training, the
proportion of IS neurons was at its highest (approximately 3 times the
pretraining level). No significant fluctuations occurred in the
proportions of neurons responding to the alternative stimulus. In
addition, nonmonotonic changes were found commonly in the activity of
230 of the neurons tracked individually from before training to shortly
after the end of training. Thus the pattern of change in responsiveness
both at the population level and at the level of individual neurons was
highly nonmonotonic. Such a pattern of change is not consistent with
simple models of memory based on synaptic strengthening to asymptote. A
model is proposed that accounts for the changes in the population
responses to the imprinting stimulus in terms of changes in the
responses of individual neurons.
Molecular dynamics of a grid-mounted molecular dipolar rotor in a rotating electric field
Authors: Vacek Jaroslav , Michl Josef
Published: July 2002

Classical molecular dynamics is applied to the rotation of a
dipolar molecular rotor mounted on a square grid and driven by rotating
electric field
E
(ν) at
T
≃ 150
K. The rotor is a complex of Re with two substituted
o
-phenanthrolines, one positively and one negatively
charged, attached to an axial position of Rh

\documentclass[12pt]{minimal}
\usepackage{amsmath}
\usepackage{wasysym}
\usepackage{amsfonts}
\usepackage{amssymb}
\usepackage{amsbsy}
\usepackage{mathrsfs}
\setlength{\oddsidemargin}{-69pt}
\begin{document}
\begin{equation*}{\mathrm{_{2}^{4+}}}\end{equation*}\end{document}

in a
[2]staffanedicarboxylate grid through
2-(3-cyanobicyclo[1.1.1]pent-1-yl)malonic dialdehyde. Four regimes
are characterized by
a
, the average lag per turn:
(
i
) synchronous (
a
<
1/
e
) at
E
(ν) =
|
E
(ν)| >
E
c
(ν)
[
E
c
(ν) is the critical field strength],
(
ii
) asynchronous (1/
e
<
a
< 1) at
E
c
(ν)
>
E
(ν) >
E
bo
(ν) >
kT
/μ, [
E
bo
(ν)
is the break-off field strength], (
iii
) random driven
(
a
≃ 1) at
E
bo
(ν) >
E
(ν)
>
kT
/μ, and (
iv
) random
thermal (
a
≃ 1) at
kT
/μ >
E
(ν). A fifth
regime, (v) strongly hindered,
W
>
kT
,
Eμ
, (
W
is the
rotational barrier), has not been examined. We find
E
bo
(ν)/kVcm
−1
≃
(
kT
/μ)/kVcm
−1
+
0.13(ν/GHz)
1.9
and
E
c
(ν)/kVcm
−1
≃
(2.3
kT
/μ)/kVcm
−1
+
0.87(ν/GHz)
1.6
. For ν > 40 GHz, the rotor
behaves as a macroscopic body with a friction constant proportional to
frequency, η/eVps ≃ 1.14 ν/THz, and for ν < 20
GHz, it exhibits a uniquely molecular behavior.
Paper-like electronic displays: Large-area rubber-stamped plastic sheets of electronics and microencapsulated electrophoretic inks
Authors: Rogers John A. , Bao Zhenan , Baldwin Kirk , Dodabalapur Ananth , Crone Brian , Raju V. R. , Kuck Valerie , Katz Howard , Amundson Karl , Ewing Jay , Drzaic Paul
Published: July 2002

Electronic systems that use rugged lightweight plastics
potentially offer attractive characteristics (low-cost processing,
mechanical flexibility, large area coverage, etc.) that are not easily
achieved with established silicon technologies. This paper summarizes
work that demonstrates many of these characteristics in a realistic
system: organic active matrix backplane circuits (256 transistors) for
large (≈5 × 5-inch) mechanically flexible sheets of electronic
paper, an emerging type of display. The success of this effort relies
on new or improved processing techniques and materials for plastic
electronics, including methods for (
i
) rubber stamping
(microcontact printing) high-resolution (≈1 μm) circuits with low
levels of defects and good registration over large areas,
(
ii
) achieving low leakage with thin dielectrics
deposited onto surfaces with relief, (
iii
) constructing
high-performance organic transistors with bottom contact geometries,
(
iv
) encapsulating these transistors, (
v
)
depositing, in a repeatable way, organic semiconductors with uniform
electrical characteristics over large areas, and (
vi
)
low-temperature (≈100°C) annealing to increase the on/off ratios
of the transistors and to improve the uniformity of their
characteristics. The sophistication and flexibility of the patterning
procedures, high level of integration on plastic substrates, large area
coverage, and good performance of the transistors are all important
features of this work. We successfully integrate these circuits with
microencapsulated electrophoretic “inks” to form sheets of
electronic paper.
T cell activation up-regulates cyclic nucleotide phosphodiesterases 8A1 and 7A3
Authors: Glavas Natalie A. , Ostenson Cari , Schaefer Jonathan B. , Vasta Valeria , Beavo Joseph A.
Published: July 2002

Agents that increase intracellular cAMP inhibit the activation and function of T cells and can lead to cell death. Recently, it has been postulated that cAMP inhibits T cell function in large part by acting as a brake on the T cell receptor and costimulatory receptor pathways. Therefore, for full activation of the T cell to occur, this inhibitory influence must be removed. One likely mechanism for accomplishing this is by up-regulation and/or activation of specific cyclic nucleotide phosphodiesterases (PDEs), and such a mechanism for one phosphodiesterase, PDE7A1, has been reported. In this paper, we extend this mechanism to another isozyme variant of the same PDE family, PDE7A3. We also report the full-length sequence of human PDE8A1 and show that it also is induced in response to a combination of T cell receptor and costimulatory receptor pathway activation. However, the time course for induction of PDE8A1 is slower than that of PDE7A1. The basal level measured and, therefore, the apparent fold induction of PDE7A1 mRNA and protein depend in large part on the method of isolation of the T cells. On the other hand, regardless of the isolation method, the basal levels of PDE7A3 and PDE8A1 are very low and fold activation is much higher. Constitutively expressed PDE8A1 and PDE7A3 also have been isolated from a human T cell line, Hut78.
Insight into the molecular basis of pathogen abundance: Group AStreptococcusinhibitor of complement inhibits bacterial adherence and internalization into human cells
Authors: Hoe Nancy P. , Ireland Robin M. , DeLeo Frank R. , Gowen Brian B. , Dorward David W. , Voyich Jovanka M. , Liu Mengyao , Burns Eugene H. , Culnan Derek M. , Bretscher Anthony , Musser James M.
Published: July 2002

Streptococcal inhibitor of complement (Sic) is a secreted protein made predominantly by serotype M1 Group AStreptococcus(GAS), which contributes to persistence in the mammalian upper respiratory tract and epidemics of human disease. Unexpectedly, an isogenicsic-negative mutant adhered to human epithelial cells significantly better than the wild-type parental strain. Purified Sic inhibited the adherence of asicnegative serotype M1 mutant and of non-Sic-producing GAS strains to human epithelial cells. Sic was rapidly internalized by human epithelial cells, inducing cell flattening and loss of microvilli. Ezrin and moesin, human proteins that functionally link the cytoskeleton to the plasma membrane, were identified as Sic-binding proteins by affinity chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis. Sic colocalized with ezrin inside epithelial cells and bound to the F-actin-binding site region located in the carboxyl terminus of ezrin and moesin. Synthetic peptides corresponding to two regions of Sic had GAS adherence-inhibitory activity equivalent to mature Sic and inhibited binding of Sic to ezrin. In addition, thesicmutant was phagocytosed and killed by human polymorphonuclear leukocytes significantly better than the wild-type strain, and Sic colocalized with ezrin in discrete regions of polymorphonuclear leukocytes. The data suggest that binding of Sic to ezrin alters cellular processes critical for efficient GAS contact, internalization, and killing. Sic enhances bacterial survival by enabling the pathogen to avoid the intracellular environment. This process contributes to the abundance of M1 GAS in human infections and their ability to cause epidemics.
X-linked adrenal hypoplasia congenita is caused by abnormal nuclear localization of the DAX-1 protein
Authors: Lehmann Sylvia G. , Lalli Enzo , Sassone-Corsi Paolo
Published: July 2002

Mutations in the
DAX-1
[dosage-sensitive sex reversal–adrenal hypoplasia congenita (AHC) critical region on the X chromosome; NR0B1] gene cause X-linked AHC associated with hypogonadotropic hypogonadism.
DAX-1
encodes an unusual orphan member of the nuclear hormone receptor superfamily, acting as a transcriptional repressor of genes involved in the steroidogenic pathway. All
DAX-1
mutations found in AHC patients alter the protein C terminus, which shares similarity to the ligand binding domain of nuclear hormone receptors and bears transcriptional repressor activity. This property is invariably impaired in DAX-1 AHC mutants. Here we show that the localization of DAX-1 AHC mutant proteins is drastically shifted toward the cytoplasm, even if their nuclear localization signal, which resides in the N terminal of the protein, is intact. Cytoplasmic localization of DAX-1 AHC mutants correlates with an impairment in their transcriptional repression activity. These results reveal a critical role of an intact C terminus in determining DAX-1 subcellular localization and constitute an important example of a defect in human organogenesis caused by impaired nuclear localization of a transcription factor.
A role for ASIC3 in the modulation of high-intensity pain stimuli
Authors: Chen Chih-Cheng , Zimmer Anne , Sun Wei-Hsin , Hall Jennifer , Brownstein Michael J. , Zimmer Andreas
Published: July 2002

Acid-sensing ion channel 3 (ASIC3), a proton-gated ion channel of the degenerins/epithelial sodium channel (DEG/ENaC) receptor family is expressed predominantly in sensory neurons including nociceptive neurons responding to protons. To study the role of ASIC3 in pain signaling, we generated ASIC3 knockout mice. Mutant animals were healthy and responded normally to most sensory stimuli. However, in behavioral assays for pain responses, ASIC3 null mutant mice displayed a reduced latency to the onset of pain responses, or more pain-related behaviors, when stimuli of moderate to high intensity were used. This unexpected effect seemed independent of the modality of the stimulus and was observed in the acetic acid-induced writhing test (0.6 vs. 0.1–0.5%), in the hot-plate test (52.5 and 55 vs. 50°C), and in tests for mechanically induced pain (tail-pinch vs. von Frey filaments). We postulate that ASIC3 is involved in modulating moderate- to high-intensity pain sensation.
Mechanistic coupling of protease signaling and initiation of coagulation by tissue factor
Authors: Riewald Matthias , Ruf Wolfram
Published: July 2002

The crucial role of cell signaling in hemostasis is clearly established by the action of the downstream coagulation protease thrombin that cleaves platelet-expressed G-protein-coupled protease activated receptors (PARs). Certain PARs are cleaved by the upstream coagulation proteases factor Xa (Xa) and the tissue factor (TF)–factor VIIa (VIIa) complex, but these enzymes are required at high nonphysiological concentrations and show limited recognition specificity for the scissile bond of target PARs. However, defining a physiological mechanism of PAR activation by upstream proteases is highly relevant because of the potent anti-inflammatory
in vivo
effects of inhibitors of the TF initiation complex. Activation of substrate factor X (X) by the TF–VIIa complex is here shown to produce enhanced cell signaling in comparison to the TF–VIIa complex alone, free Xa, or Xa that is generated
in situ
by the intrinsic activation complex. Macromolecular assembly of X into a ternary complex of TF–VIIa–X is required for proteolytic conversion to Xa, and product Xa remains transiently associated in a TF–VIIa–Xa complex. By trapping this complex with a unique inhibitor that preserves Xa activity, we directly show that Xa in this ternary complex efficiently activates PAR-1 and -2. These experiments support the concept that proinflammatory upstream coagulation protease signaling is mechanistically coupled and thus an integrated part of the TF–VIIa-initiated coagulation pathway, rather than a late event during excessive activation of coagulation and systemic generation of proteolytic activity.
Acridine and phenothiazine derivatives as pharmacotherapeutics for prion disease
Authors: Korth Carsten , May Barnaby C. H. , Cohen Fred E. , Prusiner Stanley B.
Published: July 2002

Prion diseases in humans and animals are invariably fatal. Prions
are composed of a disease-causing isoform (PrP
Sc
) of the
normal host prion protein (PrP
C
) and replicate by
stimulating the conversion of PrP
C
into nascent
PrP
Sc
. We report here that tricyclic derivatives of
acridine and phenothiazine exhibit half-maximal inhibition of
PrP
Sc
formation at effective concentrations
(EC
50
) between 0.3 μM and 3 μM in cultured cells
chronically infected with prions. The EC
50
for
chlorpromazine was 3 μM, whereas quinacrine was 10 times more potent.
A variety of 9-substituted, acridine-based analogues of quinacrine were
synthesized, which demonstrated variable antiprion potencies similar to
those of chlorpromazine and emphasized the importance of the side chain
in mediating the inhibition of PrP
Sc
formation. Thus, our
studies show that tricyclic compounds with an aliphatic side chain at
the middle ring moiety constitute a new class of antiprion reagents.
Because quinacrine and chlorpromazine have been used in humans for many
years as antimalarial and antipsychotic drugs, respectively, and are
known to pass the blood–brain barrier, we suggest that they are
immediate candidates for the treatment of Creutzfeldt–Jakob disease
and other prion diseases.
Rac1 mediates STAT3 activation by autocrine IL-6
Authors: Faruqi Tatjana R. , Gomez Dolores , Bustelo Xosé R. , Bar-Sagi Dafna , Reich Nancy C.
Published: July 2002

The activity of the small GTPase, Rac1, plays a role in various cellular processes including cytoskeletal rearrangement, gene transcription, and malignant transformation. In this report constitutively active Rac1 (Rac V12) is shown to stimulate the activation of STAT3, a member of the family of signal transducers and activators of transcription (STATs). The activity of Rac1 leads to STAT3 translocation to the nucleus coincident with STAT3-dependent gene expression. The expression of Vav (Δ1–187), a constitutively active guanine nucleotide exchange factor for the Rho GTPases, or activated forms of Ras or Rho family members, leads to STAT3-specific activation. The activation of STAT3 requires tyrosine phosphorylation at residue 705, but is not dependent on phosphorylation of Ser-727. Our studies indicate that Rac1 induces STAT3 activation through an indirect mechanism that involves the autocrine production and action of IL-6, a known mediator of STAT3 response. Rac V12 expression results in the induction of the IL-6 and IL-6 receptor genes and neutralizing antibodies directed against the IL-6 receptor block Rac1-induced STAT3 activation. Furthermore, inhibition of the nuclear factor-κB activation or disruption of IL-6-mediated signaling through the expression of IκBα S32AS36A and suppressor of cytokine signaling 3 , respectively, blocks Rac1-induced STAT3 activation. These findings elucidate a mechanism dependent on the induction of an autocrine IL-6 activation loop through which Rac1 mediates STAT3 activation establishing a link between oncogenic GTPase activity and Janus kinase/STAT signaling.
Cyclin-dependent kinases as a therapeutic target for stroke
Authors: Osuga Hitoshi , Osuga Sachiko , Wang Fuhu , Fetni Raouf , Hogan Matthew J. , Slack Ruth S. , Hakim Antoine M. , Ikeda Joh-E , Park David S.
Published: July 2002

Cyclin-dependent kinases (CDKs) are commonly known to regulate cell proliferation. However, previous reports suggest that in cultured postmitotic neurons, activation of CDKs is a signal for death rather than cell division. We determined whether CDK activation occurs in mature adult neurons during focal stroke
in vivo
and whether this signal was required for neuronal death after reperfusion injury. Cdk4/cyclin D1 levels and phosphorylation of its substrate retinoblastoma protein (pRb) increase after stroke. Deregulated levels of E2F1, a transcription factor regulated by pRb, are also observed. Administration of a CDK inhibitor blocks pRb phosphorylation and the increase in E2F1 levels and dramatically reduces neuronal death by 80%. These results indicate that CDKs are an important therapeutic target for the treatment of reperfusion injury after ischemia.
Molecular determinants for CC-chemokine recognition by a poxvirus CC-chemokine inhibitor
Authors: Seet Bruce T. , Singh Rajkumari , Paavola Chad , Lau Elaine K. , Handel Tracy M. , McFadden Grant
Published: July 2002

Poxviruses express a family of secreted proteins that bind with high affinity to chemokines and antagonize the interaction with their cognate G protein-coupled receptors (GPCRs). These viral inhibitors are novel in structure and, unlike cellular chemokine receptors, are able to specifically interact with most, if not all, CC-chemokines. We therefore sought to define the structural features of CC-chemokines that facilitate this broad-spectrum interaction. Here, we identify the residues present on human monocyte chemoattractant protein-1 (MCP-1) that are required for high-affinity interaction with the vaccinia virus 35-kDa CC-chemokine binding protein (VV-35kDa). Not only do these residues correspond to those required for interaction with the cognate receptor CCR2b but they are also conserved among many CC-chemokines. Thus, the results provide a structural basis for the ability of VV-35kDa to promiscuously recognize CC-chemokines and block binding to their receptors.
The 2.0-Å crystal structure of tachylectin 5A provides evidence for the common origin of the innate immunity and the blood coagulation systems
Authors: Kairies Norman , Beisel Hans-Georg , Fuentes-Prior Pablo , Tsuda Ryoko , Muta Tatsushi , Iwanaga Sadaaki , Bode Wolfram , Huber Robert , Kawabata Shun-ichiro
Published: July 2002

Because invertebrates lack an adaptive immune system, they had to evolve effective intrinsic defense strategies against a variety of microbial pathogens. This ancient form of host defense, the innate immunity, is present in all multicellular organisms including humans. The innate immune system of the Japanese horseshoe crab
Tachypleus tridentatus
, serving as a model organism, includes a hemolymph coagulation system, which participates both in defense against microbes and in hemostasis. Early work on the evolution of vertebrate fibrinogen suggested a common origin of the arthropod hemolymph coagulation and the vertebrate blood coagulation systems. However, this conjecture could not be verified by comparing the structures of coagulogen, the clotting protein of the horseshoe crab, and of mammalian fibrinogen. Here we report the crystal structure of tachylectin 5A (TL5A), a nonself-recognizing lectin from the hemolymph plasma of
T. tridentatus
. TL5A shares not only a common fold but also related functional sites with the γ fragment of mammalian fibrinogen. Our observations provide the first structural evidence of a common ancestor for the innate immunity and the blood coagulation systems.
Functional cooperation between T helper cell determinants
Authors: Gerloni Mara , Xiong Sidong , Mukerjee Sonjoy , Schoenberger Stephen P. , Croft Michael , Zanetti Maurizio
Published: July 2002

The immune response to T helper (Th) cell determinants of a variety of antigens is often poor and limits severely the potential efficacy of current therapeutic measures through vaccination. Here, we report that an immunologically silent tumor determinant can be rendered immunogenic if linked with a dominant determinant of a parasite antigen, suggesting the existence of functional Th–Th cooperationin vivo. This phenomenon could be mimicked in part by signaling either through CD40 to the antigen-presenting cells or through OX40 to the tumor-determinant reactive T cells, with maximal effects obtained by combined anti-CD40 and anti-OX40 treatmentin vivo. The data suggest that CD4 T cells reactive with a dominant determinant provide help to other CD4 T cells through up-regulating the costimulatory ability of antigen-presenting cells, in much the same way as help for CD8 cells. CD4 help for CD4 T cells represents a new immunological principle and offers new practical solutions for vaccine therapy against cancer and other diseases in which antigenic help is limiting.
Viruses from extreme thermal environments
Authors: Rice George , Stedman Kenneth , Snyder Jamie , Wiedenheft Blake , Willits Debbie , Brumfield Susan , McDermott Timothy , Young Mark J.
Published: July 2002

Viruses of extreme thermophiles are of great interest because they serve as model systems for understanding the biochemistry and molecular biology required for life at high temperatures. In this work, we report the discovery, isolation, and preliminary characterization of viruses and virus-like particles from extreme thermal acidic environments (70–92°C, pH 1.0–4.5) found in Yellowstone National Park. Six unique particle morphologies were found in
Sulfolobus
enrichment cultures. Three of the particle morphologies are similar to viruses previously isolated from
Sulfolobus
species from Iceland and/or Japan. Sequence analysis of their viral genomes suggests that they are related to the Icelandic and Japanese isolates. In addition, three virus particle morphologies that had not been previously observed from thermal environments were found. These viruses appear to be completely novel in nature.
Pax8 has a key role in thyroid cell differentiation
Authors: Pasca di Magliano Marina , Di Lauro Roberto , Zannini Mariastella
Published: July 2002

Transformation of rat thyroid cells with polyoma virus middle T
antigen results in loss of the thyroid-differentiated phenotype,
measured as the expression of the thyroglobulin (
Tg
),
thyroperoxidase (
TPO
), and sodium/iodide symporter
(
NIS
) genes. Among the transcription factors involved in
the regulation of these genes, TTF-1 and TTF-2 were still detected at
nearly wild-type levels, while a specific loss of the paired domain
transcription factor Pax8 was observed. In this study, we used the PCPy
cell line as a model system to study the role of Pax8 in thyroid
differentiation. We demonstrate that the reintroduction of Pax8 in PCPy
cells is sufficient to activate expression of the
endogenous genes encoding thyroglobulin, thyroperoxidase,
and sodium/iodide symporter. Thus, this cell system provides direct
evidence for the ability of Pax8 to activate transcription of
thyroid-specific genes at their chromosomal locus and strongly suggests
a fundamental role of this transcription factor in the maintenance of
functional differentiation in thyroid cells. Moreover, we show that
Pax8 and TTF-1 cooperate in the activation of the thyroglobulin
promoter and that additional thyroid-specific mechanism(s) are involved
in such a cooperation. To identify the Pax8 domain able to mediate the
specific activation of the thyroglobulin promoter, we transfected in
PCPy cells three different Pax8 isoforms. The results of such
experiments indicate that for the transcriptional activation of
thyroid-specific genes, Pax8 uses an as yet unidentified functional
domain.
Insight into hepatocellular carcinogenesis at transcriptome level by comparing gene expression profiles of hepatocellular carcinoma with those of corresponding noncancerous liver
Authors: Xu Xiang-Ru , Huang Jian , Xu Zhi-Gang , Qian Bin-Zhi , Zhu Zhi-Dong , Yan Qing , Cai Ting , Zhang Xin , Xiao Hua-Sheng , Qu Jian , Liu Feng , Huang Qiu-Hua , Cheng Zhi-Hong , Li Neng-Gan , Du Jian-Jun , Hu Wei , Shen Kun-Tang , Lu Gang , Fu Gang , Zhong Ming , Xu Shu-Hua , Gu Wen-Yi , Huang Wei , Zhao Xin-Tai , Hu Geng-Xi , Gu Jian-Ren , Chen Zhu , Han Ze-Guang
Published: July 2002

Human hepatocellular carcinoma (HCC) is one of the most
common cancers worldwide. In this work, we report on a comprehensive
characterization of gene expression profiles of hepatitis B
virus-positive HCC through the generation of a large set of 5′-read
expressed sequence tag (EST) clusters (11,065 in total) from HCC and
noncancerous liver samples, which then were applied to a cDNA
microarray system containing 12,393 genes/ESTs and to comparison with
a public database. The commercial cDNA microarray, which contains 1,176
known genes related to oncogenesis, was used also for profiling gene
expression. Integrated data from the above approaches identified 2,253
genes/ESTs as candidates with differential expression. A number of
genes related to oncogenesis and hepatic function/differentiation
were selected for further semiquantitative reverse transcriptase–PCR
analysis in 29 paired HCC/noncancerous liver samples. Many genes
involved in cell cycle regulation such as cyclins, cyclin-dependent
kinases, and cell cycle negative regulators were deregulated in most
patients with HCC. Aberrant expression of the Wnt-β-catenin pathway
and enzymes for DNA replication also could contribute to the
pathogenesis of HCC. The alteration of transcription levels was noted
in a large number of genes implicated in metabolism, whereas a profile
change of others might represent a status of dedifferentiation of the
malignant hepatocytes, both considered as potential markers of
diagnostic value. Notably, the altered transcriptome profiles in HCC
could be correlated to a number of chromosome regions with
amplification or loss of heterozygosity, providing one of the
underlying causes of the transcription anomaly of HCC.
An alternative cytokinin biosynthesis pathway
Authors: Åstot Crister , Dolezal Karel , Nordström Anders , Wang Qun , Kunkel Tim , Moritz Thomas , Chua Nam-Hai , Sandberg Göran
Published: July 2002

Studies of
de novo
cytokinin biosynthesis in
isopentenyltransferase (
ipt
)-transformed
Arabidopsis thaliana
, involving
in vivo
deuterium labeling and mass spectrometry, showed that the biosynthetic
rate of zeatinriboside-5′-monophosphate was around 66-fold higher than
that of isopentenyladenosine-5′-monophosphate (iPMP), the proposed
primary product of the
Agrobacterium
ipt. Double tracer
analysis, using [
2
H
6
] isopentenyladenosine
and deuterium oxide, provided evidence for an alternative,
iPMP-independent, biosynthetic pathway for zeatin-type cytokinins,
present in both
ipt
-expressing and wild-type
Arabidopsis thaliana
. Reduction of the biosynthetic flux
in the alternative pathway by use of mevastatin, an inhibitor for
3-hydroxy-3-methylglutaryl CoA reductase, indicated a terpenoid origin
for the side-chain precursor of the iPMP independent pathway.