bluuebunny/crossref_metadata_2025 huggingface dataset:
Wake potentials in nonuniform dusty magnetoplasmas
Authors: Salimullah M. , Shukla P. K.
Published: July 2002

It is shown that the presence of equilibrium density gradients in a magnetized dusty plasma can introduce new types of drift-like waves. These give rise to oscillatory wake-potentials, which can focus ions and attract dust grains of similar polarity. Using a test particle approach, typical profiles for wake-potentials are obtained in several interesting cases. It is found that the effective attraction length is independent of the magnetic field strength for high density dusty plasmas, whereas it is larger for larger magnetic fields in a low-density dusty plasma. Furthermore, the dust attraction takes place predominantly in the inhomogeneous region of the dusty plasma having smaller lattice periodicity for smaller scale length of the density inhomogeneity.
A simple time-dependent analytic model of the P2 asymmetry in cylindrical hohlraums
Authors: Landen O. L. , Amendt P. A. , Suter L. J. , Turner R. E. , Glendinning S. G. , Haan S. W. , Pollaine S. M. , Hammel B. A. , Tabak M. , Rosen M. D. , Lindl J. D.
Published: July 2002

A simple time-dependent analytic view factor model for the P2 asymmetry in cylindrical hohlraums is presented. The model includes the effects of hohlraum wall motion, time-varying hohlraum wall albedo and time-varying hohlraum-to-capsule diameter. The results of the model compare well with predictions from two-dimensional radiation hydrodynamics computer simulations and with time-resolved hohlraum symmetry data obtained at the Nova laser facility [J. T. Hunt and D. R. Speck, Opt. Eng. 28, 461 (1989)] and Omega laser facility [J. M. Sources, R. L. McCrory, C. P. Verdon et al., Phys. Plasmas 3, 2108 (1996)]. The model is also extended to the more complicated but relevant case of a multiple-ring cylindrical hohlraum illumination geometry, the baseline scheme for the National Ignition Facility [J. A. Paisner, J. D. Boyes, S. A. Kumpan, and M. Sorem, ICF Quart. 5, 110 (1995)].
Structure of ultrathin silicon dioxide films
Authors: Boyd Ian W. , Wilson John I. B.
Published: July 2002

Differential infrared spectroscopy has been used to study the silicon-oxygen stretching band of thin silicon dioxide films thermally grown on single-crystal silicon. We consistently observe an asymmetry in the spectra of films thicker than about 100 Å, of about 9 cm−1. The peak position, width, and degree of asymmetry are also found to be sensitively dependent upon film thickness below 100–150 Å, while above this level these features are only very weakly dependent upon film thickness, indicating the presence of a thin layer of different structural or bonding properties. Our interpretation suggests that the infrared spectra of layers up to 100 Å thick are significantly affected by strain originating at the silicon-oxide interface, in agreement with recent observations.
Photorefractive properties of doped cadmium telluride
Authors: Bylsma R. B. , Bridenbaugh P. M. , Olson D. H. , Glass A. M.
Published: July 2002

The first study of the photorefractive properties of doped CdTe has demonstrated high sensitivity for optical processing applications. Of the binary II-VI and III-V semiconductors, CdTe has the highest electro-optic coefficient r41 in the infrared, some three times larger than that of GaAs and InP. Deep levels introduced into CdTe exhibit appropriate absorption and photoconductivity at 1.06 μm by doping with V and Ti impurities. Photorefractive beam coupling experiments in CdTe:V gave small signal gains of 0.7 cm−1, and diffraction efficiencies with no applied electrical field of 0.7%. Thus, CdTe appears to be superior to previously studied III-V semiconductors, in the near-infrared spectrum. Optimization of doping and trap densities is expected to result in gain which exceeds the absorption loss, thereby allowing phase conjugation with infrared injection lasers.
Picosecond gain dynamics in KrF amplifiers
Authors: Taylor A. J. , Gibson R. B. , Roberts J. P.
Published: July 2002

The steady-state and dynamic gain characteristics in a KrF amplifier are studied with the use of 1 ps pulses. A saturation energy density in the range 1.8–2.0±0.2 mJ/cm2 is measured. The recovery of the small-signal gain coefficient exhibits two recovery processes with time constants of 67±10 ps and 2.0±0.2 ns.
Y-Ba-Cu-O films prepared by a paint-on method
Authors: Shih I. , Qiu C. X.
Published: July 2002

Polycrystalline films of Y-Ba-Cu-O with a thickness of about 20–40 μm have been prepared on alumina substrates using a paint-on method. The liquid source used was obtained by mixing powder of Y2O3, BaCO3, and CuO in liquid triethanolamine. Several Y-Ba-Cu-O films with an onset temperature of about 100 K and a zero resistance temperature of 85 K have been obtained after a short heat treatment at 1000 °C in flowing O2.
Correlation of ‘‘twins’’ observed by optical microscopy and transmission electron microscopy in YBa2Cu3O7−x superconductors
Authors: Hoff H. A. , Singh A. K. , Pande C. S.
Published: July 2002

By using transmission electron microscopy and optical microscopy on the same specimens, the patterns of light- and dark-contrast lines seen in reflected polarized light were shown to be an interference pattern due to the variable spacing of suboptical microtwins. These microtwins are mostly [110] reflection twins. The [110] twinning was observed to be cyclic and occasionally pseudotetragonal because of the progressive cycling of contact twin domains. Within a domain, and occasionally in a whole grain, the [110] reflection twins occurred as polysynthetic lamellae. The morphology of the domain structure can be explained from the theory of martensitic transformation.
Y-Ba-Cu-O superconducting film on oxidized silicon
Authors: Gupta Ram P. , Khokle W. S. , Dubey R. C. , Singhal Seema , Nagpal K. C. , Rao G. S. T. , Jain J. D.
Published: July 2002

We report thick superconducting films of Y-Ba-Cu-O on oxidized silicon substrates. The critical temperatures for onset and zero resistance are 96 and 77 K, respectively. X-ray diffraction analysis predicts 1, 2, 3 composition and orthorhombic phase of the film.
Liquid-sodium thermoacoustic engine
Authors: Migliori A. , Swift G. W.
Published: July 2002

We have constructed a thermoacoustic engine that uses liquid sodium as its working substance. The engine generates acoustic power using heat flowing from a high-temperature source to a low-temperature sink. The measured performance of this engine disagrees significantly with numerical calculations based on our theory of thermoacoustic engines. The efficiency of the engine is a substantial fraction of Carnot’s efficiency, and its power density is comparable to that of the conventional heat engines in widespread use. Thus we expect this type of engine to be of practical, economic importance.
Dynamics and orientation of N
+
(CD
3
)
3
-bromoacetylcholine bound to its binding site on the nicotinic acetylcholine receptor
Authors: Williamson P. T. F. , Watts J. A. , Addona G. H. , Miller K. W. , Watts A.
Published: July 2002

Dynamic and structural information has been obtained for an
analogue of acetylcholine while bound to the agonist binding site on
the nicotinic acetylcholine receptor (nAcChoR), using wide-line
deuterium solid-state NMR. Analysis of the deuterium lineshape obtained
at various temperatures from unoriented nAcChoR membranes labeled with
deuterated bromoacetylcholine (BAC) showed that the quaternary ammonium
group of the ligand is well constrained within the agonist binding site
when compared with the dynamics observed in the crystalline solids.
This motional restriction would suggest that a high degree of
complementarity exists between the quaternary ammonium group of the
ligand and the protein within the agonist binding site. nAcChoR
membranes were uniaxially oriented by isopotential centrifugation as
determined by phosphorous NMR of the membrane phospholipids. Analysis
of the deuterium NMR lineshape of these oriented membranes enriched
with the nAcChoR labeled with
N
+
(CD
3
)
3
-BAC has enabled us to
determine that the angle formed between the quaternary ammonium group
of the BAC and the membrane normal is 42° in the desensitized form of
the receptor. This measurement allows us to orient in part the bound
ligand within the proposed receptor binding site.
Cortical correlates of learning in monkeys adapting to a new dynamical environment
Authors: Gandolfo F. , Li C.-S. R. , Benda B. J. , Schioppa C. Padoa , Bizzi E.
Published: July 2002

In this paper, we describe the neural changes observed in the primary motor cortex of two monkeys while they learned a new motor skill. The monkeys had to adapt their reaching movements to external forces that interfered with the execution of their arm movements. We found a sizable population of cells that changed their tuning properties during exposure to the force field. These cells took on the properties of neurons that are involved in the control of movement. Furthermore, the cells maintained the acquired activity as the monkey readapted to the no-force condition. Recent imaging studies in humans have reported the effects of motor learning in the primary motor cortex. Our results are consistent with the findings of these studies and provide evidence for single-cell plasticity in the primary motor cortex of primates.
Active site mutant transgene confers tolerance to human β-glucuronidase without affecting the phenotype of MPS VII mice
Authors: Sly William S. , Vogler Carole , Grubb Jeffrey H. , Zhou Mi , Jiang Jinxing , Zhou Xiao Yan , Tomatsu Shunji , Bi Yanhua , Snella Elizabeth M.
Published: July 2002

Mucopolysaccharidosis type VII (MPS VII; Sly syndrome) is an
autosomal recessive lysosomal storage disorder due to an inherited
deficiency of β-glucuronidase. A naturally occurring mouse model for
this disease was discovered at The Jackson Laboratory and shown to be
due to homozygosity for a 1-bp deletion in exon 10 of the
gus
gene. The murine model MPS VII
(
gus
mps/mps
) has been very well
characterized and used extensively to evaluate experimental strategies
for lysosomal storage diseases, including bone marrow transplantation,
enzyme replacement therapy, and gene therapy. To enhance the value of
this model for enzyme and gene therapy, we produced a transgenic mouse
expressing the human β-glucuronidase cDNA with an amino acid
substitution at the active site nucleophile (E540A) and bred it onto
the MPS VII (
gus
mps/mps
) background. We
demonstrate here that the mutant mice bearing the active site mutant
human transgene retain the clinical, morphological, biochemical, and
histopathological characteristics of the original MPS VII
(
gus
mps/mps
) mouse. However, they are now
tolerant to immune challenge with human β-glucuronidase. This
“tolerant MPS VII mouse model” should be useful for preclinical
trials evaluating the effectiveness of enzyme and/or gene therapy
with the human gene products likely to be administered to human
patients with MPS VII.
Visual habit formation in monkeys with neurotoxic lesions of the ventrocaudal neostriatum
Authors: Fernandez-Ruiz Juan , Wang Jin , Aigner Thomas G. , Mishkin Mortimer
Published: July 2002

Visual habit formation in monkeys, assessed by concurrent visual
discrimination learning with 24-h intertrial intervals (ITI), was found
earlier to be impaired by removal of the inferior temporal visual area
(TE) but not by removal of either the medial temporal lobe or inferior
prefrontal convexity, two of TE's major projection targets. To assess
the role in this form of learning of another pair of structures to
which TE projects, namely the rostral portion of the tail of the
caudate nucleus and the overlying ventrocaudal putamen, we injected a
neurotoxin into this neostriatal region of several monkeys and tested
them on the 24-h ITI task as well as on a test of visual recognition
memory. Compared with unoperated monkeys, the experimental animals were
unaffected on the recognition test but showed an impairment on the 24-h
ITI task that was highly correlated with the extent of their
neostriatal damage. The findings suggest that TE and its projection
areas in the ventrocaudal neostriatum form part of a circuit that
selectively mediates visual habit formation.
On the absorbance changes in the photocycle of the photoactive yellow protein: A quantum-chemical analysis
Authors: Molina Vicent , Merchán Manuela
Published: July 2002

Spectral changes in the photocycle of the photoactive yellow protein (PYP) are investigated by usingab initiomulticonfigurational second-order perturbation theory at the available structures experimentally determined. Using the dark ground-state crystal structure [Genick, U. K., Soltis, S. M., Kuhn, P., Canestrelli, I. L. & Getzoff, E. D. (1998)Nature (London)392, 206–209], the ππ* transition to the lowest excited state is related to the typical blue-light absorption observed at 446 nm. The different nature of the second excited state (nπ*) is consistent with the alternative route detected at 395-nm excitation. The results suggest the low-temperature photoproduct PYPHLas the most plausible candidate for the assignment of the cryogenically trapped early intermediate (Genicket al.). We cannot establish, however, a successful correspondence between the theoretical spectrum for the nanosecond time-resolved x-ray structure [Perman, B., Šrajer, V., Ren, Z., Teng, T., Pradervand, C.,et al. (1998)Science279, 1946–1950] and any of the spectroscopic photoproducts known up to date. It is fully confirmed that the colorless light-activated intermediate recorded by millisecond time-resolved crystallography [Genick, U. K., Borgstahl, G. E. O., Ng, K., Ren, Z., Pradervand, C.,et al. (1997)Science275, 1471–1475] is protonated, nicely matching the spectroscopic features of the photoproduct PYPM. The overall contribution demonstrates that a combined analysis of high-level theoretical results and experimental data can be of great value to perform assignments of detected intermediates in a photocycle.
Cyclin D3 activates Caspase 2, connecting cell proliferation with cell death
Authors: Mendelsohn Andrew R. , Hamer Janice D. , Wang Zhaoyuan Bill , Brent Roger
Published: July 2002

Precancerous cells that enter S phase without appropriate growth and viability factors undergo programmed cell death, suggesting that apoptosis may help guarantee organismic integrity [Evan, G. & Littlewood, T. (1998)
Science
281, 1317–1322]. However, the connection between proliferation and cell death has remained unclear. Here, we show that the positive cell cycle regulator cyclin D3 [Matsushime H., Roussel M. F., Ashmun, R. A. & Sherr, C. J. (1991)
Cell
65, 701–713] interacts with the death enzyme Caspase 2 [Wang, L., Miura, M., Bergeron, L., Zhu, H. & Yuan, J. (1994)
Cell
78, 739–750]. Directed expression of cyclin D3 and Caspase 2 in human cells potentiated apoptosis compared with expression of Caspase 2 alone. Cyclin D3 expression increased the amount of cleaved (active) Caspase 2. We describe a PCR mutagenesis/ligation/two-hybrid/green fluorescent protein approach that facilitates the isolation of missense mutant proteins defective in interaction with particular partners absent other phenotypes or knowledge of the system. We used this approach to isolate Caspase 2 mutants that did not bind cyclin D3 (noninteractors). Noninteractors were sensitive to apoptosis-dependent proteolysis, but did not potentiate apoptosis. Noninteractors did not block apoptosis caused by wild-type Caspase 2. Our results are consistent with the idea that an interaction with cyclin D3 may stabilize Caspase 2, and suggest that a physical interaction between cyclin D3 and Caspase 2 connects the genetic networks that govern cell-cycle progression with those that govern cell death.
Epithelial and endothelial expression of the green fluorescent protein reporter gene under the control of bovine prion protein (PrP) gene regulatory sequences in transgenic mice
Authors: Lemaire-Vieille Catherine , Schulze Tobias , Podevin-Dimster Valérie , Follet Jérome , Bailly Yannick , Blanquet-Grossard Françoise , Decavel Jean-Pierre , Heinen Ernst , Cesbron Jean-Yves
Published: July 2002

The expression of the cellular form of the prion protein (PrP
c
) gene is required for prion replication and neuroinvasion in transmissible spongiform encephalopathies. The identification of the cell types expressing PrP
c
is necessary to understanding how the agent replicates and spreads from peripheral sites to the central nervous system. To determine the nature of the cell types expressing PrP
c
, a green fluorescent protein reporter gene was expressed in transgenic mice under the control of 6.9 kb of the bovine PrP gene regulatory sequences. It was shown that the bovine PrP gene is expressed as two populations of mRNA differing by alternative splicing of one 115-bp 5′ untranslated exon in 17 different bovine tissues. The analysis of transgenic mice showed reporter gene expression in some cells that have been identified as expressing PrP, such as cerebellar Purkinje cells, lymphocytes, and keratinocytes. In addition, expression of green fluorescent protein was observed in the plexus of the enteric nervous system and in a restricted subset of cells not yet clearly identified as expressing PrP: the epithelial cells of the thymic medullary and the endothelial cells of both the mucosal capillaries of the intestine and the renal capillaries. These data provide valuable information on the distribution of PrP
c
at the cellular level and argue for roles of the epithelial and endothelial cells in the spread of infection from the periphery to the brain. Moreover, the transgenic mice described in this paper provide a model that will allow for the study of the transcriptional activity of the PrP gene promoter in response to scrapie infection.
Diacylglycerol kinase ɛ regulates seizure susceptibility and long-term potentiation through arachidonoyl– inositol lipid signaling
Authors: Rodriguez de Turco Elena B. , Tang Wen , Topham Matthew K. , Sakane Fumio , Marcheselli Victor L. , Chen Chu , Taketomi Akinobu , Prescott Stephen M. , Bazan Nicolas G.
Published: July 2002

Arachidonoyldiacylglycerol (20:4-DAG) is a
second messenger derived from phosphatidylinositol 4,5-bisphosphate and
generated by stimulation of glutamate metabotropic receptors linked to
G proteins and activation of phospholipase C. 20:4-DAG signaling is
terminated by its phosphorylation to phosphatidic acid, catalyzed by
diacylglycerol kinase (DGK). We have cloned the murine
DGKɛ gene that showed, when expressed in COS-7 cells, selectivity for
20:4-DAG. The significance of DGKɛ in synaptic function was
investigated in mice with targeted disruption of the DGKɛ.
DGKɛ
−/−
mice showed a higher resistance to
eletroconvulsive shock with shorter tonic seizures and faster recovery
than DGKɛ
+/+
mice. The phosphatidylinositol
4,5-bisphosphate-signaling pathway in cerebral cortex was greatly
affected, leading to lower accumulation of 20:4-DAG and free 20:4.
Also, long-term potentiation was attenuated in perforant path–dentate
granular cell synapses. We propose that DGKɛ contributes to modulate
neuronal signaling pathways linked to synaptic activity, neuronal
plasticity, and epileptogenesis.
Endosomal localization and function of sorting nexin 1
Authors: Zhong Qi , Lazar Cheri S. , Tronchère Hélène , Sato Trey , Meerloo Timo , Yeo Michele , Songyang Zhou , Emr Scott D. , Gill Gordon N.
Published: July 2002

There are 17 human members of the sorting nexin (SNX) family of proteins that contain Phox (PX) domains. Yeast orthologs function in vesicular trafficking and mammalian proteins have been implicated in endocytic trafficking of cell surface receptors. The first member of this family, SNX1, was identified via interaction with the epidermal growth factor receptor. The present studies indicate that SNX1 and SNX2 are colocalized to tubulovesicular endosomal membranes and this localization depends on PI 3-kinase activity. Point mutations in the PX domain that abolish recognition of phosphorylated phosphatidylinositol (PtdIns)
in vitro
abolish vesicle localization
in vivo
indicating that lipid binding by the PX domain is necessary for localization to vesicle membranes. Deletion of a predicted coiled-coil region in the COOH terminus of SNX1 also abolished vesicle localization, indicating that this helical domain, too, is necessary for SNX1 localization. Thus, both PX domain recognition of PtdIns and COOH terminal helical domains are necessary for localization of SNX1 with neither alone being sufficient. Regulated overexpression of the NH
2
terminus of SNX1 containing the PX domain decreased the rate of ligand-induced epidermal growth factor receptor degradation, an effect consistent with inhibition of endogenous SNX1 function in the endosome compartment. SNX1 thus functions in regulating trafficking in the endosome compartment via PX domain recognition of phosphorylated PtdIns and via interaction with other protein components.
Nonsaturation of AMPA and NMDA receptors at hippocampal synapses
Authors: McAllister A. Kimberley , Stevens Charles F.
Published: July 2002

An important issue in synaptic physiology is the extent to which postsynaptic receptors are saturated by the neurotransmitter released from a single synaptic vesicle. Although the bulk of evidence supports receptor saturation, recent studies have started to reveal that α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) andN-methyl-d-aspartate (NMDA) receptors may not be saturated by a single vesicle of glutamate. Here, we address this question through a study of putative single synapses, made by hippocampal neurons in culture, that are identified by FM1–43 staining. An analysis of the sources of variability in the amplitudes of miniature excitatory postsynaptic currents at single synapses reveals that this variability must arise presynaptically, from variations in the quantity of agonist released. Thus, glutamate receptors at hippocampal synapses are not generally saturated by quantal release.
μ-Opioid receptors and limbic responses to aversive emotional stimuli
Authors: Liberzon Israel , Zubieta Jon Kar , Fig Lorraine M. , Phan K. Luan , Koeppe Robert A. , Taylor Stephan F.
Published: July 2002

Functional neuroimaging studies implicate limbic and paralimbic activity in emotional responses, but few studies have sought to understand neurochemical mechanisms which modulate these responses. We have used positron emission tomography to measure μ-opioid receptor binding, and cerebral blood flow in the same subjects, and demonstrated that the baseline binding potential and the regional cerebral blood flow in the left inferior temporal pole are functionally related. Higher baseline μ-opioid receptor binding potential was associated with lower regional cerebral blood flow in this region during presentation of emotionally salient stimuli. This is consistent with an inhibitory/anxiolytic role of the endogenous opioid system in limbic regions of the temporal lobe and basal forebrain.
Coactivator/corepressor ratios modulate PR-mediated transcription by the selective receptor modulator RU486
Authors: Liu Zheng , Auboeuf Didier , Wong Jiemin , Chen J. Don , Tsai Sophia Y. , Tsai Ming-Jer , O'Malley Bert W.
Published: July 2002

Selective receptor modulators, such as the antiprogestin RU486, are known to exhibit partial agonist activities in a cell-type-dependent manner. Employing an
in vitro
chromatin transcription system that recapitulates progesterone receptor (PR)-mediated transcription
in vivo
, we have investigated the molecular basis by which the antiprogestin RU486 regulates transcription in a cell-type-specific manner. We have compared the effects of RU486 on PR-dependent transcription
in vitro
using T47D and HeLa cell nuclear extracts. RU486 exhibits a differential ability to activate transcription within these two cell types. The differential effect on transcription correlates with different ratios of endogenous coactivators/corepressors in these cells. Unlike agonist-bound PR that interacts only with coactivators such as steroid receptor coactivator-1 (SRC-1), RU486-bound PR binds to both coactivator SRC-1 and corepressor silencing mediator for retinoid and thyroid hormone receptor (SMRT)
in vitro
. Both SRC-1 and SMRT have the capacity to modulate RU486-dependent activity. Moreover, a change in the relative levels of SRC-1 and SMRT contained in our chromatin transcription system modulates agonist/antagonist effects of RU486 on transcription by PR. Our data indicate that the ability of RU486 to activate transcription is modulated by the ratio of coactivators to corepressors and substantiate the important roles of coregulators in the regulation of steroid receptor mediated transactivation in response to selective receptor modulators.
Adaptive evolution of small heat shock protein/ αB-crystallin promoter activity of the blind subterranean mole rat,Spalax ehrenbergi
Authors: Hough R. B. , Avivi A. , Davis J. , Joel A. , Nevo E. , Piatigorsky J.
Published: July 2002

Blind mole rats have degenerated subcutaneous eyes that are visually nonfunctional. In this investigation, we have compared the tissue specificity of the small heat shock protein (shsp)/αB-crystallin promoter of the mole rat superspecies,Spalax ehrenbergi, with that of the mouse. Earlier experiments showed that mouse shsp/αB-crystallin promoter/enhancer activity is high in the lens and moderate in the heart and skeletal muscle of transgenic mice. Here, we show in transgenic mouse experiments using the firefly luciferase reporter gene that, despite relatively few changes in sequence, the mole rat shsp/αB-crystallin promoter/enhancer has selectively lost lens activity after 13.5 days of embryogenesis (E13.5). The ratios of mole rat/mouse promoter activity were 0.01 for lens, 1.7 for heart, and 13.6 for skeletal muscle in 8-wk-old transgenic mice. Our data indicate that the shsp/αB-crystallin promoter/enhancer has undergone adaptive changes corresponding to the subterranean evolution of the blind mole rat. We speculate that selective pressures on metabolic economy may have contributed to these tissue-specific modifications of promoter/enhancer function during adaptation to life underground.
From hematopoiesis to neuropoiesis: Evidence of overlapping genetic programs
Authors: Terskikh Alexey V. , Easterday Mathew C. , Li Linheng , Hood Leroy , Kornblum Harley I. , Geschwind Daniel H. , Weissman Irving L.
Published: July 2002

It is reasonable to propose that gene expression profiles of purified stem cells could give clues for the molecular mechanisms of stem cell behavior. We took advantage of cDNA subtraction to identify a set of genes selectively expressed in mouse adult hematopoietic stem cells (HSC) as opposed to bone marrow (BM). Analysis of HSC-enriched genes revealed several key regulatory gene candidates, including two novel seven transmembrane (7TM) receptors. Furthermore, by using cDNA microarray techniques we found a large set of HSC-enriched genes that are expressed in mouse neurospheres (a population greatly enriched for neural progenitor cells), but not present in terminally differentiated neural cells.In situhybridization demonstrated that many of them, including one HSC-enriched 7TM receptor, were selectively expressed in the germinal zones of fetal and adult brain, the regions harboring mouse neural stem cells. We propose that at least some of the transcripts that are selectively and commonly expressed in two or more types of stem cells define a functionally conserved group of genes evolved to participate in basic stem cell functions, including stem cell self-renewal.
γ-Secretase inhibitors repress thymocyte development
Authors: Hadland Brandon K. , Manley Nancy R. , Su Dong-ming , Longmore Gregory D. , Moore Chad L. , Wolfe Michael S. , Schroeter Eric H. , Kopan Raphael
Published: July 2002

A major therapeutic target in the search for a cure to the devastating Alzheimer's disease is γ-secretase. This activity resides in a multiprotein enzyme complex responsible for the generation of Aβ42 peptides, precipitates of which are thought to cause the disease. γ-Secretase is also a critical component of the Notch signal transduction pathway; Notch signals regulate development and differentiation of adult self-renewing cells. This has led to the hypothesis that therapeutic inhibition of γ-secretase may interfere with Notch-related processes in adults, most alarmingly in hematopoiesis. Here, we show that application of γ-secretase inhibitors to fetal thymus organ cultures interferes with T cell development in a manner consistent with loss or reduction of Notch1 function. Progression from an immature CD4−/CD8−state to an intermediate CD4+/CD8+double-positive state was repressed. Furthermore, treatment beginning later at the double-positive stage specifically inhibited CD8+single-positive maturation but did not affect CD4+single-positive cells. These results demonstrate that pharmacological γ-secretase inhibition recapitulates Notch1 loss in a vertebrate tissue and present a system in which rapid evaluation of γ-secretase-targeted pharmaceuticals for their ability to inhibit Notch activity can be performed in a relevant context.
Thermodynamic characterization of the interaction between TRAF2 and tumor necrosis factor receptor peptides by isothermal titration calorimetry
Authors: Ye Hong , Wu Hao
Published: July 2002

The tumor necrosis factor receptor (TNFR) superfamily can induce diverse biological effects, including cell survival, proliferation, differentiation, and apoptosis. The major signal transducers for TNFRs are the family of TNF receptor associated factors (TRAFs). The direct interaction between TRAFs and the intracellular tails of TNFRs is the first step of this signal relay process. Structural studies have revealed a trimeric nature of TRAF2 and a symmetrical mode of receptor binding, suggesting the involvement of trivalent TNFR2-receptor interaction in the signal transduction. In this study, using isothermal titration calorimetry (ITC), we report thermodynamic characterization of the interaction between TRAF2 and monomeric peptide sequences from TNFR members, including TNFR2, CD40, CD30, Ox40, and 4-1BB, and the Epstein–Barr virus (EBV)-transforming protein, latent infection membrane protein-1 (LMP1). The dissociation constants of the interaction were shown to range between 40 μM and 1.9 mM, which are substantially weaker than most protein–peptide interactions. The interaction is entirely driven by exothermic enthalpy, consistent with the abundance of polar contacts. The enthalpy of the interaction has a significant temperature dependence (ΔCp = −245 cal/mol⋅K). The unfavorable entropy in the interaction and the comparison with structural energetics calculations suggest the involvement of conformational rearrangement in the interaction. The low affinity of TRAF2 to monomeric receptor peptides further supports the importance of avidity contribution in TRAF2 recruitment by these receptors upon ligand-induced trimerization or higher order oligomerization.
Myotubularin, a protein tyrosine phosphatase mutated in myotubular myopathy, dephosphorylates the lipid second messenger, phosphatidylinositol 3-phosphate
Authors: Taylor Gregory S. , Maehama Tomohiko , Dixon Jack E.
Published: July 2002

The lipid second messenger phosphatidylinositol 3-phosphate [PI(3)P] plays a crucial role in intracellular membrane trafficking. We report here that myotubularin, a protein tyrosine phosphatase required for muscle cell differentiation, is a potent PI(3)P phosphatase. Recombinant human myotubularin specifically dephosphorylates PI(3)P
in vitro
. Overexpression of a catalytically inactive substrate-trapping myotubularin mutant (C375S) in human 293 cells increases PI(3)P levels relative to that of cells overexpressing the wild-type enzyme, demonstrating that PI(3)P is a substrate for myotubularin
in vivo
. In addition, a
Saccharomyces cerevisiae
strain in which the myotubularin-like gene (YJR110w) is disrupted also exhibits increased PI(3)P levels. Both the recombinant yeast enzyme and a human myotubularin-related protein (KIAA0371) are able to dephosphorylate PI(3)P
in vitro
, suggesting that this activity is intrinsic to all myotubularin family members. Mutations in the
MTM1
gene that cause human myotubular myopathy dramatically reduce the ability of the phosphatase to dephosphorylate PI(3)P. Our findings provide evidence that myotubularin exerts its effects during myogenesis by regulating cellular levels of the inositol lipid PI(3)P.
Speeding molecular recognition by using the folding funnel: The fly-casting mechanism
Authors: Shoemaker Benjamin A. , Portman John J. , Wolynes Peter G.
Published: July 2002

Protein folding and binding are kindred processes. Many proteins in the cell are unfolded, so folding and function are coupled. This paper investigates how binding kinetics is influenced by the folding of a protein. We find that a relatively unstructured protein molecule can have a greater capture radius for a specific binding site than the folded state with its restricted conformational freedom. In this scenario of binding, the unfolded state binds weakly at a relatively large distance followed by folding as the protein approaches the binding site: the “fly-casting mechanism.” We illustrate this scenario with the hypothetical kinetics of binding a single repressor molecule to a DNA site and find that the binding rate can be significantly enhanced over the rate of binding of a fully folded protein.
The coreceptor mutation CCR5Δ32 influences the dynamics of HIV epidemics and is selected for by HIV
Authors: Sullivan Amy D. , Wigginton Janis , Kirschner Denise
Published: July 2002

We explore the impact of a host genetic factor on heterosexual HIV epidemics by using a deterministic mathematical model. A protective allele unequally distributed across populations is exemplified in our models by the 32-bp deletion in the host-cell chemokine receptor CCR5, CCR5Δ32. Individuals homozygous for CCR5Δ32 are protected against HIV infection whereas those heterozygous for CCR5Δ32 have lower pre-AIDS viral loads and delayed progression to AIDS. CCR5Δ32 may limit HIV spread by decreasing the probability of both risk of infection and infectiousness. In this work, we characterize epidemic HIV within three dynamic subpopulations: CCR5/CCR5 (homozygous, wild type), CCR5/CCR5Δ32 (heterozygous), and CCR5Δ32/CCR5Δ32 (homozygous, mutant). Our results indicate that prevalence of HIV/AIDS is greater in populations lacking the CCR5Δ32 alleles (homozygous wild types only) as compared with populations that include people heterozygous or homozygous for CCR5Δ32. Also, we show that HIV can provide selective pressure for CCR5Δ32, increasing the frequency of this allele.
Mutational inactivation of the proapoptotic gene
BAX
confers selective advantage during tumor clonal evolution
Authors: Ionov Yurij , Yamamoto Hiroyuki , Krajewski Stanislaw , Reed John C. , Perucho Manuel
Published: July 2002

A remarkable instability at simple repeated sequences characterizes
gastrointestinal cancer of the microsatellite mutator phenotype (MMP).
Mutations in the DNA mismatch repair gene family underlie the MMP, a
landmark for hereditary nonpolyposis colorectal cancer. These tumors
define a distinctive pathway for carcinogenesis because they display a
particular spectrum of mutated cancer genes containing target repeats
for mismatch repair deficiency. One such gene is
BAX
, a
proapoptotic member of the Bcl-2 family of proteins, which
plays a key role in programmed cell death. More than half of colon and
gastric cancers of the MMP contain
BAX
frameshifts in a
(G)
8
mononucleotide tract. However, the functional
significance of these mutations in tumor progression has not been
established. Here we show that inactivation of the wild-type
BAX
allele by
de novo
frameshift
mutations confers a strong advantage during tumor clonal evolution.
Tumor subclones with only mutant alleles frequently appeared after
inoculation into nude mice of single-cell clones of colon tumor cell
lines with normal alleles. In contrast, no clones of BAX-expressing
cells were found after inoculation of homozygous cell clones without
wild-type
BAX
. These results support the interpretation
that
BAX
inactivation contributes to tumor progression
by providing a survival advantage. In this context, survival analyses
show that
BAX
mutations are indicators of poor prognosis
for both colon and gastric cancer of the MMP.
Cardiomyocytes induce endothelial cells to trans-differentiate into cardiac muscle: Implications for myocardium regeneration
Authors: Condorelli G. , Borello U. , De Angelis L. , Latronico M. , Sirabella D. , Coletta M. , Galli R. , Balconi G. , Follenzi A. , Frati G. , Cusella De Angelis M. G. , Gioglio L. , Amuchastegui S. , Adorini L. , Naldini L. , Vescovi A. , Dejana E. , Cossu G.
Published: July 2002

The concept of tissue-restricted differentiation of postnatal stem cells has been challenged by recent evidence showing pluripotency for hematopoietic, mesenchymal, and neural stem cells. Furthermore, rare but well documented examples exist of already differentiated cells in developing mammals that change fate and trans-differentiate into another cell type. Here, we report that endothelial cells, either freshly isolated from embryonic vessels or established as homogenous cells in culture, differentiate into beating cardiomyocytes and express cardiac markers when cocultured with neonatal rat cardiomyocytes or when injected into postischemic adult mouse heart. Human umbilical vein endothelial cells also differentiate into cardiomyocytes under similar experimental conditions and transiently coexpress von Willebrand factor and sarcomeric myosin. In contrast, neural stem cells, which efficiently differentiate into skeletal muscle, differentiate into cardiomyocytes at a low rate. Fibroblast growth factor 2 and bone morphogenetic protein 4, which activate cardiac differentiation in embryonic cells, do not activate cardiogenesis in endothelial cells or stimulate trans-differentiation in coculture, suggesting that different signaling molecules are responsible for cardiac induction during embryogenesis and in successive periods of development. The fact that endothelial cells can generate cardiomyocytes sheds additional light on the plasticity of endothelial cells during development and opens perspectives for cell autologous replacement therapies.
Dominant localization of prostaglandin D receptors on arachnoid trabecular cells in mouse basal forebrain and their involvement in the regulation of non-rapid eye movement sleep
Authors: Mizoguchi Akira , Eguchi Naomi , Kimura Kazushi , Kiyohara Yoshimoto , Qu Wei-Min , Huang Zhi-Li , Mochizuki Takatoshi , Lazarus Michael , Kobayashi Takuya , Kaneko Takeshi , Narumiya Shuh , Urade Yoshihiro , Hayaishi Osamu
Published: July 2002

Infusion of prostaglandin (PG) D
2
into the lateral
ventricle of the brain induced an increase in the amount of non-rapid
eye movement sleep in wild-type (WT) mice but not in mice deficient in
the PGD receptor (DP). Immunofluorescence staining of WT mouse brain
revealed that DP immunoreactivity was dominantly localized in the
leptomeninges (LM) of the basal forebrain but that PGD synthase
immunoreactivity was widely distributed in the LM of the entire brain.
Electron microscopic observation indicated that DP-immunoreactive
particles were predominantly located on the plasma membranes of
arachnoid trabecular cells of the LM. The region with the highest DP
immunoreactivity was clearly defined as bilateral wings in the LM of
the basal forebrain located lateral to the optic chiasm in the
proximity of the ventrolateral preoptic area, one of the putative sleep
centers, and the tuberomammillary nucleus, one of the putative wake
centers. The LM of this region contained DP mRNA 70-fold higher than
that in the cortex as judged from the results of quantitative reverse
transcription–PCR. PGD
2
infusion into the subarachnoid
space of this region increased the extracellular adenosine level more
than 2-fold in WT mice but not in the DP-deficient mice. These results
indicate that DPs in the arachnoid trabecular cells of the basal
forebrain mediate an increase in the extracellular adenosine level and
sleep induction by PGD
2
.
Comparison of mechanisms of action of luteinizing hormone-releasing hormone (LHRH) antagonist cetrorelix and LHRH agonist triptorelin on the gene expression of pituitary LHRH receptors in rats
Authors: Kovacs Magdolna , Schally Andrew V.
Published: July 2002

The mechanisms through which luteinizing hormone (LH)-releasing hormone (LHRH) antagonists suppress pituitary gonadotroph functions and LHRH-receptor (LHRH-R) expression are incompletely understood. Consequently, we investigated the direct effect of LHRH antagonist cetrorelix
in vitro
on the expression of the pituitary LHRH-R gene and its ability to counteract the exogenous LHRH and the agonist triptorelin in the regulation of this gene. We also compared the effects of chronic administration of cetrorelix and triptorelin on the LHRH-R mRNA level and gonadotropin secretion in ovariectomized (OVX) and normal female rats. The exposure of pituitary cells
in vitro
to 3-min pulses of 1 nM LHRH or 0.1 nM triptorelin for 5 h increased the LHRH-R mRNA level by 77–88%. Continuous perfusion of the cells with 50 nM cetrorelix did not cause any significant changes, but prevented the stimulatory effect of LHRH pulses on the receptor mRNA expression. In OVX rats, 10 days after administration of a depot formulation of cetrorelix, releasing 100 μg of peptide daily, the elevated LHRH-R mRNA level was decreased by 73%, whereas daily injection of 100 μg of triptorelin caused a 41% suppression. In normal female rats, cetrorelix treatment suppressed the LHRH-R mRNA level by 33%, but triptorelin increased it by 150%. The highly elevated serum LH levels in OVX rats and the normal LH concentration of cycling rats were rapidly and completely suppressed by cetrorelix. Triptorelin decreased the serum LH in OVX rats to the precastration level, but had no effect on basal LH in normal rats. Our results confirm that LHRH antagonists, such as cetrorelix, inhibit the gene expression of pituitary LHRH-R indirectly, by counteracting the stimulatory effect of LHRH. A rapid suppression of serum LH by LHRH antagonists would be advantageous in the treatment of sex hormone-dependent tumors and other conditions.
Toward detecting and identifying macromolecules in a cellular context: Template matching applied to electron tomograms
Authors: Böhm Jochen , Frangakis Achilleas S. , Hegerl Reiner , Nickell Stephan , Typke Dieter , Baumeister Wolfgang
Published: July 2002

Electron tomography is the only technique available that allows us
to visualize the three-dimensional structure of unfixed and unstained
cells currently with a resolution of 6–8 nm, but with the prospect to
reach 2–4 nm. This raises the possibility of detecting and identifying
specific macromolecular complexes within their cellular context by
virtue of their structural signature. Templates derived from the
high-resolution structure of the molecule under scrutiny are used to
search the reconstructed volume. Here we outline and test a
computationally feasible two-step procedure: In a first step,
mean-curvature motion is used for segmentation, yielding subvolumes
that contain with a high probability macromolecules in the expected
size range. Subsequently, the particles contained in the subvolumes are
identified by cross-correlation, using a set of three-dimensional
templates. With simulated and real tomographic data we demonstrate that
such an approach is feasible and we explore the detection limits. Even
structurally similar particles, such as the thermosome, GroEL, and the
20S proteasome can be identified with high fidelity. This opens up
exciting prospects for mapping the territorial distribution of
macromolecules and for analyzing molecular interactions
in
situ
.
SNAP-29: A general SNARE protein that inhibits SNARE disassembly and is implicated in synaptic transmission
Authors: Su Qingning , Mochida Sumiko , Tian Jin-Hua , Mehta Rashi , Sheng Zu-Hang
Published: July 2002

Using the yeast two-hybrid system with syntaxin-1A as bait, we isolated soluble NSF attachment protein (SNAP)-29 from a human brain cDNA library. Synaptosomal fractionation and immunocytochemical staining of hippocampal neurons in culture showed that SNAP-29 is present at synapses and is predominantly associated with synaptic vesicles. The interaction of SNAP-29 with syntaxin-1 was further confirmed with immunoprecipitation analysis. Binding competition studies with SNAP-29 demonstrated that it could compete with α-SNAP for binding to synaptic SNAP receptors (SNAREs) and consequently inhibit disassembly of the SNARE complex. Introduction of SNAP-29 into presynaptic superior cervical ganglion neurons in culture significantly inhibited synaptic transmission in an activity-dependent manner. Although SNAP-29 has been suggested to be a general SNARE component in membrane trafficking, our findings suggest that it may function as a regulator of SNARE complex disassembly and modulate the process of postfusion recycling of the SNARE components.
The γ-secretase-cleaved C-terminal fragment of amyloid precursor protein mediates signaling to the nucleus
Authors: Gao Yuehua , Pimplikar Sanjay W.
Published: July 2002

Sequential processing of the amyloid precursor protein
(APP) by β- and γ-secretases generates the Aβ peptide, a
major constituent of the senile plaques observed in Alzheimer's
disease. The cleavage by γ-secretase also results in the cytoplasmic
release of a 59- or 57-residue-long C-terminal fragment (Cγ). This
processing resembles regulated intramembrane proteolysis of
transmembrane proteins such as Notch, where the released cytoplasmic
fragments enter the nucleus and modulate gene expression. Here, we
examined whether the analogous Cγ fragments of APP also exert effects
in the nucleus. We find that ectopically expressed Cγ is present both
in the cytoplasm and in the nucleus. Interestingly, expression of
Cγ59 causes disappearance of PAT1, a protein that interacts with the
APP cytoplasmic domain, from the nucleus and induces its proteosomal
degradation. Treatment of cells with lactacystin prevents PAT1
degradation and retains its nuclear localization. By contrast, Cγ57,
a minor product of γ-cleavage, is only marginally effective in PAT1
degradation. Furthermore, Cγ59 but not Cγ57 potently represses
retinoic acid-responsive gene expression. Thus, our studies provide the
evidence that, as predicted by the regulated intramembrane
proteolysis mechanism, Cγ seems to function in the nucleus.
Fluorescent light-induced chromatid breaks distinguish Alzheimer disease cells from normal cells in tissue culture.
Authors: Parshad R P , Sanford K K , Price F M , Melnick L K , Nee L E , Schapiro M B , Tarone R E , Robbins J H
Published: July 2002

The neurodegeneration and amyloid deposition of sporadic Alzheimer disease (AD) also occur in familial AD and in all trisomy-21 Down syndrome (DS) patients, suggesting a common pathogenetic mechanism. We investigated whether defective processing of damaged DNA might be that mechanism, as postulated for the neurodegeneration in xeroderma pigmentosum, a disease with defective repair not only of UV radiation-induced, but also of some oxygen free radical-induced, DNA lesions. We irradiated AD and DS skin fibroblasts or blood lymphocytes with fluorescent light, which is known to cause free radical-induced DNA damage. The cells were then treated with either beta-cytosine arabinoside (araC) or caffeine, and chromatid breaks were quantified. At least 28 of 31 normal donors and 10 of 11 donors with nonamyloid neurodegenerations gave normal test results. All 12 DS, 11 sporadic AD, and 16 familial AD patients tested had abnormal araC and caffeine tests, as did XP-A cells. In one of our four AD families, an abnormal caffeine test was found in all 10 afflicted individuals (including 3 asymptomatic when their skin biopsies were obtained) and in 8 of 11 offspring at a 50% risk for AD. Our tests could prove useful in predicting inheritance of familial AD and in supporting, or rendering unlikely, the diagnosis of sporadic AD in patients suspected of having the disease.