bluuebunny/crossref_metadata_2025 huggingface dataset:
Autoregulation of a bacterial σ factor explored by using segmental isotopic labeling and NMR
Authors: Camarero Julio A. , Shekhtman Alexander , Campbell Elizabeth A. , Chlenov Mark , Gruber Tanja M. , Bryant Donald A. , Darst Seth A. , Cowburn David , Muir Tom W.
Published: July 2002

Bacterial σ factors combine with the catalytic core RNA polymerase to direct the process of transcription initiation through sequence-specific interactions with the −10 and −35 elements of promoter DNA. In the absence of core RNA polymerase, the DNA-binding function of σ is autoinhibited by its own N-terminal 90 amino acids (region 1.1), putatively by a direct interaction with conserved region 4.2, which binds the −35 promoter element. In the present work, this mechanism of autoinhibition was studied by using a combination of NMR spectroscopy and segmental isotopic labeling of a σ
70
-like subunit from
Thermotoga maritima
. Our data argue strongly against a high-affinity interaction between these two domains. Instead we suggest that autoinhibition of DNA binding occurs through an indirect steric and/or electrostatic mechanism. More generally, the present work illustrates the power of segmental isotopic labeling for probing molecular interactions in large proteins by NMR.
Inosine induces axonal rewiring and improves behavioral outcome after stroke
Authors: Chen Peng , Goldberg David E. , Kolb Bryan , Lanser Marc , Benowitz Larry I.
Published: July 2002

Cerebral infarct (stroke) often causes devastating and irreversible losses of function, in part because of the brain's limited capacity for anatomical reorganization. The purine nucleoside inosine has previously been shown to induce neurons to express a set of growth-associated proteins and to extend axons in culture and
in vivo
. We show here that in adult rats with unilateral cortical infarcts, inosine stimulated neurons on the undamaged side of the brain to extend new projections to denervated areas of the midbrain and spinal cord. This growth was paralleled by improved performance on several behavioral measures.
CD25+CD4+ T cells contribute to the control of memory CD8+ T cells
Authors: Murakami Masaaki , Sakamoto Akemi , Bender Jeremy , Kappler John , Marrack Philippa
Published: July 2002

Previously we demonstrated that IL-15 and IL-2 control the number of memory CD8+ T cells in mice. IL-15 induces, and IL-2 suppresses the division of these cells. Here we show that CD25+CD4+ regulatory T cells play an important role in the IL-2-mediated control of memory phenotype CD8+ T cell number. In animals, the numbers of CD25+CD4+ T cells were inversely correlated with the numbers of memory phenotype CD8+ T cells with age. Treatment with anti-IL-2 caused CD25+CD4+ T cells to disappear and, concurrently, increased the numbers of memory phenotype CD8+ T cells. This increase in the numbers of CD8+ memory phenotype T cells was not manifest in animals lacking CD4+ cells. Importantly, adoptive transfer of CD25+CD4+ T cells significantly reduced division of memory phenotype CD8+ T cells. Thus, we conclude that CD25+CD4+ T cells are involved in the IL-2-mediated inhibition of memory CD8+ T cell division and that IL-2 controls memory phenotype CD8+ T cell numbers at least in part through maintenance of the CD25+CD4+ T cell population.
A molecular mechanism of action of theophylline: Induction of histone deacetylase activity to decrease inflammatory gene expression
Authors: Ito Kazuhiro , Lim Sam , Caramori Gaetano , Cosio Borja , Chung K. Fan , Adcock Ian M. , Barnes Peter J.
Published: July 2002

The molecular mechanism for the anti-inflammatory action of theophylline is currently unknown, but low-dose theophylline is an effective add-on therapy to corticosteroids in controlling asthma. Corticosteroids act, at least in part, by recruitment of histone deacetylases (HDACs) to the site of active inflammatory gene transcription. They thereby inhibit the acetylation of core histones that is necessary for inflammatory gene transcription. We show bothin vitroandin vivothat low-dose theophylline enhances HDAC activity in epithelial cells and macrophages. This increased HDAC activity is then available for corticosteroid recruitment and predicts a cooperative interaction between corticosteroids and theophylline. This mechanism occurs at therapeutic concentrations of theophylline and is dissociated from phosphodiesterase inhibition (the mechanism of bronchodilation) or the blockade of adenosine receptors, which are partially responsible for its side effects. Thus we have shown that low-dose theophylline exerts an anti-asthma effect through increasing activation of HDAC which is subsequently recruited by corticosteroids to suppress inflammatory genes.
Cluster analysis of gene expression dynamics
Authors: Ramoni Marco F. , Sebastiani Paola , Kohane Isaac S.
Published: July 2002

This article presents a Bayesian method for model-based clustering of gene expression dynamics. The method represents gene-expression dynamics as autoregressive equations and uses an agglomerative procedure to search for the most probable set of clusters given the available data. The main contributions of this approach are the ability to take into account the dynamic nature of gene expression time series during clustering and a principled way to identify the number of distinct clusters. As the number of possible clustering models grows exponentially with the number of observed time series, we have devised a distance-based heuristic search procedure able to render the search process feasible. In this way, the method retains the important visualization capability of traditional distance-based clustering and acquires an independent, principled measure to decide when two series are different enough to belong to different clusters. The reliance of this method on an explicit statistical representation of gene expression dynamics makes it possible to use standard statistical techniques to assess the goodness of fit of the resulting model and validate the underlying assumptions. A set of gene-expression time series, collected to study the response of human fibroblasts to serum, is used to identify the properties of the method.
Gene activation by interaction of an inhibitor with a cytoplasmic signaling protein
Authors: Peng Gang , Hopper James E.
Published: July 2002

Galactose-inducible genes (
GAL
genes) in yeast
Saccharomyces cerevisiae
are efficiently transcribed only when the sequence-specific transcription activator Gal4p is activated. Activation of Gal4p requires the interaction between the Gal4p inhibitory protein Gal80p and the galactokinase paralog, Gal3p. It has been proposed that Gal3p binds to a Gal80p-Gal4p complex in the nucleus to activate Gal4p. Here, we present evidence that the Gal3p–Gal80p interaction occurs in the cytoplasm, and concurrently, Gal80p is removed from Gal4p at the
GAL
gene promoter. We also show that
GAL
gene expression can be activated by heterologous protein–protein interaction in the cytoplasm that is independent of galactose and Gal3p function. These results indicate that galactose-triggered Gal3p-Gal80p association in the cytoplasm activates Gal4p in the nucleus.
Gene order evolution and paleopolyploidy in hemiascomycete yeasts
Authors: Wong Simon , Butler Geraldine , Wolfe Kenneth H.
Published: July 2002

The wealth of comparative genomics data from yeast species allows the molecular evolution of these eukaryotes to be studied in great detail. We used “proximity plots” to visually compare chromosomal gene order information from 14 hemiascomycetes, including the recent Génolevures survey, to
Saccharomyces cerevisiae
. Contrary to the original reports, we find that the Génolevures data strongly support the hypothesis that
S. cerevisiae
is a degenerate polyploid. Using gene order information alone, 70% of the
S. cerevisiae
genome can be mapped into “sister” regions that tile together with almost no overlap. This map confirms and extends the map of sister regions that we constructed previously by using duplicated genes, an independent source of information. Combining gene order and gene duplication data assigns essentially the whole genome into sister regions, the largest gap being only 36 genes long. The 16 centromere regions of
S. cerevisiae
form eight pairs, indicating that an ancestor with eight chromosomes underwent complete doubling; alternatives such as segmental duplications can be ruled out. Gene arrangements in
Kluyveromyces lactis
and four other species agree quantitatively with what would be expected if they diverged from
S. cerevisiae
before its polyploidization. In contrast,
Saccharomyces exiguus
,
Saccharomyces servazzii
, and
Candida glabrata
show higher levels of gene adjacency conservation, and more cases of imperfect conservation, suggesting that they split from the
S. cerevisiae
lineage after polyploidization. This finding is confirmed by sequences around the
C. glabrata TRP1
and
IPP1
loci, which show that it contains sister regions derived from the same duplication event as that of
S. cerevisiae
.
Evidence for
S
-nitrosothiol-dependent changes in fibrinogen that do not involve transnitrosation or thiolation
Authors: Akhter Shirin , Vignini Arianna , Wen Zhong , English Ann , Wang Peng G. , Mutus Bulent
Published: July 2002

S
-nitrosoglutathione (GSNO, 50 μM) inhibited the initial rate of thrombin-catalyzed human and bovine fibrinogen polymerization by ≈50% to 68% respectively. Inhibition was also observed with other structurally varied
S
-nitrosothiols (RSNOs) including sugar derivatives of
S
-nitroso-
N
-acetylpenicillamine (SNAP). The fact that the same concentration of GSNO had no effect on thrombin-dependent hydrolysis of tosylglycylprolylarginine-4-nitroanilide acetate suggested that this inhibition was due to GSNO-induced changes in fibrinogen structure. This result was confirmed by CD spectroscopy where GSNO or
S
-nitrosohomocysteine increased the α-helical content of fibrinogen by ≈15% and 11%, respectively.
S-
carboxymethylamido derivatives of glutathione or homocysteine had no effect on the fibrinogen secondary structure. The GSNO-dependent secondary structural effects were reversed on gel filtration chromatography, suggesting that the effects were allosteric. Further evidence for fibrinogen–GSNO interactions was obtained from GSNO-dependent quenching of the intrinsic fibrinogen Trp fluorescence and the perturbation of the GSNO circular dichroic absorbance as a function of [fibrinogen]. The
K
d
s of 3 to 10 μM for fibrinogen–GSNO interactions with a stoichiometry of 2:1 (GSNO:fibrinogen) were estimated from isothermal titration calorimetry and fluorescence quenching, respectively. These results suggest that RSNOs induce changes to fibrinogen structure by interacting at specific aromatic rich domains. Three such putative RSNO-binding domains have been identified in the unordered, aromatic residue-rich C-termini of the α-chains of fibrinogen.
HIV-1 matrix protein p17 increases the production of proinflammatory cytokines and counteracts IL-4 activity by binding to a cellular receptor
Authors: De Francesco Maria A. , Baronio Manuela , Fiorentini Simona , Signorini Costantino , Bonfanti Carlo , Poiesi Claudio , Popovic Mikulas , Grassi Manuela , Garrafa Emirena , Bozzo Luisa , Lewis George K. , Licenziati Stefano , Gallo Robert C. , Caruso Arnaldo
Published: July 2002

Purified recombinant HIV-1 p17 matrix protein significantly increased HIV-1 replication in preactivated peripheral blood mononuclear cell cultures obtained from healthy donors. Because HIV-1 infection and replication is related to cell activation and differentiation status, in the present study, we investigated the role played by p17 during the process of T cell stimulation. Using freshly isolated peripheral blood mononuclear cells, we demonstrate that p17 was able to enhance levels of tumor necrosis factor α and IFN-γ released from cells stimulated by IL-2. IL-4 was found to down-regulate IFN-γ and tumor necrosis factor α, and p17 restored the ability of cells to produce both cytokines. The property of p17 to increase production of proinflammatory cytokines could be a mechanism exploited by the virus to create a more suitable environment for HIV-1 infection and replication. Our data show that p17 exerts its biological activity after binding to a specific cellular receptor expressed on activated T lymphocytes. The functional p17 epitope involved in receptor binding was found to be located at the NH2-terminal region of viral protein. Immunization of BALB/c mice with a 14-aa synthetic peptide representative of the HIV-1 p17 functional region (SGGELDRWEKIRLR) resulted in the development of p17 neutralizing antibodies capable of blocking the interaction between p17 and its cellular receptor. Our results define a role for p17 in HIV-1 pathogenesis and contribute to our understanding of the molecular mechanism of HIV-1 infection and the development of additional antiviral therapeutic strategies.
GATA-3 transcriptional imprinting in Th2 lymphocytes: A mathematical model
Authors: Höfer Thomas , Nathansen Holger , Löhning Max , Radbruch Andreas , Heinrich Reinhart
Published: July 2002

Immunological memory involves the fast recall of cytokine expression by T helper (Th) lymphocytes. Two distinct profiles of cytokine expression, Th1 and Th2, can be induced by antigen and polarizing signals during activation of naive Th cells and can subsequently be reexpressed on stimulation by antigen alone. The transcription factor GATA-3 induces Th2 development. GATA-3 is activated by the Th2-polarizing stimulus, IL-4, and has recently been observed to autoactivate its transcription. Based on these experimental data, we developed a mathematical model of GATA-3 expression that assumes independent activation of GATA-3 transcription by IL-4 and by GATA-3. Cooperativity of GATA-3 transcriptional activation is shown to create a threshold for autoactivation, resulting in the coexistence of two distinct GATA-3 expression states: a state of basal expression and a state of high expression sustained by autoactivation. Suprathreshold IL-4 signals induce a transition from basal to high GATA-3 expression. Thus, GATA-3 autoactivation creates a bistable system that can memorize a transient inductive signal. The model further predicts conditions under which the state of high GATA-3 expression can be abolished, which may extinguish the Th2 cytokine memory.
Kinetic analysis of estrogen receptor/ligand interactions
Authors: Rich Rebecca L. , Hoth Lise R. , Geoghegan Kieran F. , Brown Thomas A. , LeMotte Peter K. , Simons Samuel P. , Hensley Preston , Myszka David G.
Published: July 2002

Surface plasmon resonance biosensor technology was used to directly measure the binding interactions of small molecules to the ligand-binding domain of human estrogen receptor. In a screening mode, specific ligands of the receptor were easily discerned from nonligands. In a high-resolution mode, the association and dissociation phase binding responses were shown to be reproducible and could be fit globally to a simple interaction model to extract reaction rate constants. On average, antagonist ligands (such as tamoxifen and nafoxidine) were observed to bind to the receptor with association rates that were 500-fold slower than agonists (such as estriol and β-estradiol). This finding is consistent with these antagonists binding to an altered conformation of the receptor. The biosensor assay also could identify subtle differences in how the same ligand interacted with two different isoforms of the receptor (α and β). The biosensor's ability to determine kinetic rate constants for small molecule/protein interactions provides unique opportunities to understand the mechanisms associated with complex formation as well as new information to drive the optimization of drug candidates.
Promoter-proximal tethering elements regulate enhancer-promoter specificity in the
Drosophila
Antennapedia
complex
Authors: Calhoun Vincent C. , Stathopoulos Angelike , Levine Michael
Published: July 2002

Insulator DNAs and promoter competition regulate enhancer–promoter interactions within complex genetic loci. Here we provide evidence for a third mechanism: promoter-proximal tethering elements. The
Scr
-
ftz
region of the
Antennapedia
gene complex includes two known enhancers, AE1 and T1. AE1 selectively interacts with the
ftz
promoter to maintain pair-rule stripes of
ftz
expression during gastrulation and germ-band elongation. The T1 enhancer, located 3′ of the
ftz
gene and ≈25 kb 5′ of the
Scr
promoter, selectively activates
Scr
expression in the prothorax and posterior head segments. A variety of
P
element minigenes were examined in transgenic embryos to determine the basis for specific AE1-
ftz
and T1-
Scr
interactions. A 450-bp DNA fragment located ≈100 bp 5′ of the
Scr
transcription start site is essential for T1-
Scr
interactions and can mediate long-range activation of a
ftz
/
lacZ
reporter gene when placed 5′ of the
ftz
promoter. We suggest that the
Scr
450 fragment contains tethering elements that selectively recruit T1 to the
Scr
promoter. Tethering elements might regulate enhancer–promoter interactions at other complex genetic loci.
Molecular dynamic study of orotidine-5′-monophosphate decarboxylase in ground state and in intermediate state: A role of the 203–218 loop dynamics
Authors: Hur Sun , Bruice Thomas C.
Published: July 2002

Molecular dynamics simulations have been used to derive the structures of ground (orotidine-5′-monophosphate decarboxylase⋅orotidine 5′-monophosphate; ODC⋅OMP) and intermediate (ODC⋅intermediate; ODC⋅I
−
) states in the ODC-catalyzed decarboxylation of OMP. For comparison, a molecular dynamics simulation of the conformers of OMP dissolved in water was also studied. This structural information is unavailable from present crystal structures. The electrostatic network in the active site around the carboxylate moiety of OMP exhibits remarkable stability. The conformation of enzyme-bound OMP is very similar to the conformation of OMP in water. Thus, the proposed Circe effect mechanism for ODC catalysis is unlikely. Comparison of ground state and intermediate state structures shows that on decarboxylation C6 takes the position of the carboxylate O8. This significant movement of the ligand is accompanied by a placement of the C6 carbanion in the vicinity of the protonated Lys-93 and is enforced by a change of the 203–218 loop from an unstructured form to an ordered β-hairpin. Previously proposed mechanisms involving protonation at O2, O4, or C5 have in common internal stabilization of the anionic intermediate by conjugation with positive charge on the pyrimidine ring. These mechanisms are not supported because there are no proton sources near O2, O4, and C5. We propose that the stabilization of intermediate ODC⋅I
−
is achieved by movement of the carbanion toward the external cation Lys-93 on decarboxylation and organization of the 203–218 loop. Because the intermediate and transition state are energetically similar, stabilization of the former decreases the free energy content of the latter.
Cytotoxic T lymphocytes directed against a tumor-specific mutated antigen display similar HLA tetramer binding but distinct functional avidity and tissue distribution
Authors: Echchakir Hamid , Dorothée Guillaume , Vergnon Isabelle , Menez Jeanne , Chouaib Salem , Mami-Chouaib Fathia
Published: July 2002

We have previously identified an antigen (Ag) recognized on a human large cell carcinoma of the lung by a tumor-specific cytotoxic T lymphocyte clone derived from autologous tumor infiltrating lymphocytes (TILs). The antigenic peptide is presented by HLA-A2 molecules and is encoded by a mutated α-actinin-4(ACTN4) gene. In the present report, we have isolated two anti-α-actinin-4 T cell clones from the same patient TIL and from his peripheral blood lymphocytes (PBLs) by using tetramers of soluble HLA-A2 molecules loaded with the mutated peptide. Although all of the clones displayed similar tetramer labeling, those isolated from PBL showed lower avidity of Ag recognition and killed the specific target much less efficiently, indicating that tetramer staining does not correlate with clone avidity/tumor reactivity. T cell receptor (TCR) analysis revealed that α-actinin-4-reactive clones used distinct α and β chain rearrangements, demonstrating TCR repertoire diversity. Interestingly, TCRβ chain gene usage indicated that only Ag-specific clones with high functional avidity were expanded at the tumor site, whereas a low-avidity clone was exclusively amplified in patient peripheral blood. Our results point to the existence of distinct but overlapping antitumor TCR repertoires in TIL and PBL and suggest a selectivein situexpansion of tumor-specific cytotoxic T lymphocyte with high avidity/tumor reactivity.
Atomic force microscopy of Precambrian microscopic fossils
Authors: Kempe André , Schopf J. William , Altermann Wladyslaw , Kudryavtsev Anatoliy B. , Heckl Wolfgang M.
Published: July 2002

Atomic force microscopy (AFM) is a technique used routinely in material science to image substances at a submicron (including nm) scale. We apply this technique to analysis of the fine structure of organic-walled Precambrian fossils, microscopic sphaeromorph acritarchs (cysts of planktonic unicellular protists) permineralized in ≈650-million-year-old cherts of the Chichkan Formation of southern Kazakhstan. AFM images, backed by laser-Raman spectroscopic analysis of individual specimens, demonstrate that the walls of these petrified fossils are composed of stacked arrays of ≈200-nm-sized angular platelets of polycyclic aromatic kerogen. Together, AFM and laser-Raman spectroscopy provide means by which to elucidate the submicron-scale structure of individual microscopic fossils, investigate the geochemical maturation of ancient organic matter, and, potentially, distinguish true fossils from pseudofossils and probe the mechanisms of fossil preservation by silica permineralization.
Trehalose synthesis is induced upon exposure of
Escherichia coli
to cold and is essential for viability at low temperatures
Authors: Kandror Olga , DeLeon Ann , Goldberg Alfred L.
Published: July 2002

Trehalose accumulates dramatically in microorganisms during heat shock and osmotic stress and helps protect cells against thermal injury and oxygen radicals. Here we demonstrate an important role of this sugar in cold-adaptation of bacteria. A mutant
Escherichia coli
strain unable to produce trehalose died much faster than the wild type at 4°C. Transformation of the mutant with the

otsA

/
otsB
genes, responsible for trehalose synthesis, restored trehalose content and cell viability at 4°C. After temperature downshift from 37°C to 16°C (“cold shock”), trehalose levels in wild-type cells increased up to 8-fold. Although this accumulation of trehalose did not influence growth at 16°C, it enhanced cell viability when the temperature fell further to 4°C. Before the trehalose build-up, levels of mRNA encoding OtsA/OtsB increased markedly. This induction required the σ factor, RpoS, but was independent of the major cold-shock protein, CspA.
otsA
/
B
mRNA was much more stable at 16°C than at 37°C and contained a “downstream box,” characteristic of cold-inducible mRNAs. Thus,
otsA
/
otsB
induction and trehalose synthesis are activated during cold shock (as well as during heat shock) and play an important role in resistance of
E. coli
(and probably other organisms) to low temperatures.
Cell surface expression and secretion of heparanase markedly promote tumor angiogenesis and metastasis
Authors: Goldshmidt Orit , Zcharia Eyal , Abramovitch Rinat , Metzger Shula , Aingorn Helena , Friedmann Yael , Schirrmacher Volker , Mitrani Eduardo , Vlodavsky Israel
Published: July 2002

The present study emphasizes the importance of cell surface expression and secretion of heparanase (endo-β-d-glucuronidase) in tumor angiogenesis and metastasis. For this purpose, nonmetastatic Eb mouse lymphoma cells were transfected with the predominantly intracellular human heparanase or with a readily secreted chimeric construct composed of the human enzyme and the chicken heparanase signal peptide. Eb cells overexpressing the secreted heparanase invaded a reconstituted basement membrane to a much higher extent than cells overexpressing the intracellular enzyme. Cell invasion was inhibited in the presence of laminaran sulfate, a potent inhibitor of heparanase activity and experimental metastasis. The increased invasivenessin vitrowas reflectedin vivoby rapid and massive liver colonization and accelerated mortality. In fact, mice inoculated with cells expressing the secreted enzyme succumb because of liver metastasis and dysfunction, as early as 10 days after s.c. inoculation of the cells, when their tumor burden did not exceed 1% of body weight. Cell surface localization and secretion of heparanase markedly stimulated tumor angiogenesis, as demonstrated by a 4–6-fold increase in vessel density and functionality evaluated by MRI of tumors produced by cells expressing the secreted vs. the nonsecreted heparanase, consistent with actual counting of blood vessels. Altogether, our results indicate that the potent proangoigenic and prometastatic properties of heparanase are tightly regulated by its cellular localization and secretion. The increased potency of the secreted enzyme makes it a promising target for anticancer drug development.
The CcmE protein of the
c
-type cytochrome biogenesis system: Unusual
in vitro
heme incorporation into apo-CcmE and transfer from holo-CcmE to apocytochrome
Authors: Daltrop Oliver , Stevens Julie M. , Higham Christopher W. , Ferguson Stuart J.
Published: July 2002

Three key steps of cytochrome
c
biogenesis in many Gram-negative bacteria, the uptake of heme by the heme chaperone CcmE, the covalent attachment of heme to CcmE, and its subsequent release from CcmE to an apocytochrome
c
, have been achieved
in vitro
. apo-CcmE from
Escherichia coli
preferentially bound to ferric, with high affinity (
K
d
, 200 nM), rather than ferrous heme. The preference for ferric heme was confirmed by competition with 8-anilino-1-naphthalenesulfonate, which bound to a hydrophobic pocket in apo-CcmE. Reduction under certain conditions of the ferric heme–CcmE complex, which has characteristics of a
b
-type cytochrome, resulted in covalent attachment of heme to the protein. The resulting
in vitro
-produced holo-CcmE was identical to the
in vivo
-produced holo-CcmE, proving that unmodified Fe-protoporphyrin IX is incorporated into CcmE. Only noncovalent binding of mesoheme to CcmE was observed, thus implicating at least one vinyl group in covalent binding of heme to CcmE. Heme transferred
in vitro
from holo-CcmE to apocytochrome
c
, provided the heme was reduced. The necessity for reduced holo-CcmE might explain the role of the heme chaperone, i.e., prevention of reaction of ferric heme with apocytochrome and thus avoidance of incorrect side products. In addition, an AXXAH mutant of the CXXCH binding motif in the apocytochrome
c
was unable to accept heme from holo-CcmE. These
in vitro
results mimic, and thus have implications for, the molecular pathway of heme transfer during
c
-type cytochrome maturation in many species of bacteria
in vivo
.
Virtual migration in tethered flying monarch butterflies reveals their orientation mechanisms
Authors: Mouritsen Henrik , Frost Barrie J.
Published: July 2002

A newly developed flight simulator allows monarch butterflies to fly actively for up to several hours in any horizontal direction while their fall migratory flight direction can be continuously recorded. From these data, long segments of virtual flight paths of tethered, flying, migratory monarch butterflies were reconstructed, and by advancing or retarding the butterflies' circadian clocks, we have shown that they possess a time-compensated sun compass. Control monarchs on local time fly approximately southwest, those 6-h time-advanced fly southeast, and 6-h time-delayed butterflies fly in northwesterly directions. Moreover, butterflies flown in the same apparatus under simulated overcast in natural magnetic fields were randomly oriented and did not change direction when magnetic fields were rotated. Therefore, these experiments do not provide any evidence that monarch butterflies use a magnetic compass during migration.
Crystal structure of human cytomegalovirus IL-10 bound to soluble human IL-10R1
Authors: Jones Brandi C. , Logsdon Naomi J. , Josephson Kristopher , Cook Jennifer , Barry Peter A. , Walter Mark R.
Published: July 2002

Human IL-10 (hIL-10) modulates critical immune and inflammatory responses by way of interactions with its high- (IL-10R1) and low-affinity (IL-10R2) cell surface receptors. Human cytomegalovirus exploits the IL-10 signaling pathway by expressing a functional viral IL-10 homolog (cmvIL-10), which shares only 27% sequence identity with hIL-10 yet signals through IL-10R1 and IL-10R2. To define the molecular basis of this virus–host interaction, we determined the 2.7-Å crystal structure of cmvIL-10 bound to the extracellular fragment of IL-10R1 (sIL-10R1). The structure reveals cmvIL-10 forms a disulfide-linked homodimer that binds two sIL-10R1 molecules. Although cmvIL-10 and hIL-10 share similar intertwined topologies and sIL-10R1 binding sites, their respective interdomain angles differ by ∼40°. This difference results in a striking re-organization of the IL-10R1s in the putative cell surface complex. Solution binding studies show cmvIL-10 and hIL-10 share essentially identical affinities for sIL-10R1 whereas the Epstein–Barr virus IL-10 homolog (ebvIL-10), whose structure is highly similar to hIL-10, exhibits a ∼20-fold reduction in sIL-10R1 affinity. Our results suggest cmvIL-10 and ebvIL-10 have evolved different molecular mechanisms to engage the IL-10 receptors that ultimately enhance the respective ability of their virus to escape immune detection.
Interaction of intracellular β amyloid peptide with chaperone proteins
Authors: Fonte Virginia , Kapulkin Wadim Jan , Taft Andrew , Fluet Amy , Friedman David , Link Christopher D.
Published: July 2002

Expression of the human β amyloid peptide (Aβ) in transgenic
Caenorhabditis elegans
animals can lead to the formation of intracellular immunoreactive deposits as well as the formation of intracellular amyloid. We have used this model to identify proteins that interact with intracellular Aβ
in vivo
. Mass spectrometry analysis of proteins that specifically coimmunoprecipitate with Aβ has identified six likely chaperone proteins: two members of the HSP70 family, three αB-crystallin-related small heat shock proteins (HSP-16s), and a putative ortholog of a mammalian small glutamine-rich tetratricopeptide repeat-containing protein proposed to regulate HSP70 function. Quantitative reverse transcription–PCR analysis shows that the small heat shock proteins are also transcriptionally induced by Aβ expression. Immunohistochemistry demonstrates that HSP-16 protein closely colocalizes with intracellular Aβ in this model. Transgenic animals expressing a nonaggregating Aβ variant, a single-chain Aβ dimer, show an altered pattern of coimmunoprecipitating proteins and an altered cellular distribution of HSP-16. Double-stranded RNA inhibition of R05F9.10, the putative
C. elegans
ortholog of the human small glutamine-rich tetratricopeptide-repeat-containing protein (SGT), results in suppression of toxicity associated with Aβ expression. These results suggest that chaperone function can play a role in modulating intracellular Aβ metabolism and toxicity.
Internal IgH class switch region deletions are position-independent and enhanced by AID expression
Authors: Dudley Darryll D. , Manis John P. , Zarrin Ali A. , Kaylor Lianne , Tian Ming , Alt Frederick W.
Published: July 2002

Ig heavy chain class switch recombination (CSR) involves a recombination/deletion mechanism that exchanges the expressed CHgene with a downstream CHgene. CSR is mediated by highly repetitive switch (S) region sequences and requires the activation-induced deaminase (AID). The S region 5′ of the Cμ gene (Sμ) can undergo high-frequency internal deletions in normal B cells and B cell lines activated for CSR, although the relationship of these deletions and CSR has not been elucidated. In this study, we introduced constitutively transcribed Sμ or Sγ2b regions into a pro-B cell line that can be activated for AID expression, CSR, and endogenous Sμ deletions. We find that randomly integrated S region transcription units in these cells also undergo increased levels of internal rearrangements after cellular activation, indicating that the deletion process is independent of location within the Ig heavy chain locus and potentially AID-promoted. To test the latter issue, we generated hybridomas from wild-type and AID-deficient activated B cells and assayed them for internal Sμ deletions and S region mutations. These studies demonstrated that efficient intra-S region recombination depends on AID expression and that internal S region deletions are accompanied by frequent mutations, indicating that most S region deletions occur by the same mechanism as CSR.
Intercellular calcium signaling mediated by point-source burst release of ATP
Authors: Arcuino Gregory , Lin Jane H.-C. , Takano Takahiro , Liu Collins , Jiang Li , Gao Qun , Kang Jian , Nedergaard Maiken
Published: July 2002

Calcium signaling, manifested as intercellular waves of rising cytosolic calcium, is, in many cell types, the result of calcium-induced secretion of ATP and activation of purinergic receptors. The mechanism by which ATP is released has hitherto not been established. Here, we show by real-time bioluminescence imaging that ATP efflux is not uniform across a field of cells but is restricted to brief, abrupt point-source bursts. The ATP bursts emanate from single cells and manifest the transient opening of nonselective membrane channels, which admits fluorescent indicators of ≤1.5 kDa. These observations challenge the existence of regenerative ATP release, because ATP efflux is finite and restricted to a point source. Transient efflux of cytosolic nucleotides from a subset of cells may represent a conserved pathway for coordinating local activity of electrically nonexcitable cells, because identical patterns of ATP release were identified in human astrocytes, endothelial cells, and bronchial epithelial cells.
Defective T cell development and function incalcineurinAβ-deficient mice
Authors: Bueno Orlando F. , Brandt Eric B. , Rothenberg Marc E. , Molkentin Jeffery D.
Published: July 2002

The calcium-dependent phosphatase calcineurin and its downstream transcriptional effector nuclear factor of activated T cells (NFAT) are important regulators of inducible gene expression in multiple cell types. In T cells, calcineurin-NFAT signaling represents a critical event for mediating cellular activation and the immune response. The widely used immunosuppressant agents cyclosporin and FK506 are thought to antagonize the immune response by directly inhibiting calcineurin-NFAT signal transduction in lymphocytes. To unequivocally establish the importance of calcineurin signaling as a mediator of the immune response, we deleted the gene encoding the predominant calcineurin isoform expressed in lymphocytes,calcineurin Aβ(CnAβ).CnAβ−/−mice were viable as adults, but displayed defective T cell development characterized by fewer total CD3 cells and reduced CD4 and CD8 single positive cells. Total peripheral T cell numbers were significantly reduced inCnAβ−/−mice and were defective in proliferative capacity and IL-2 production in response to PMA/ionomycin and T cell receptor cross-linking.CnAβ−/−mice also were permissive to allogeneic tumor-cell transplantationin vivo, similar to cyclosporin-treated wild-type mice. A mechanism for the compromised immune response is suggested by the observation thatCnAβ−/−T cells are defective in stimulation-induced NFATc1, NFATc2, and NFATc3 activation. These results establish a critical role for CnAβ signaling in regulating T cell development and activationin vivo.
Accountability in action?: the case of a database purchasing decision 1
Authors: Neyland Daniel , Woolgar Steve
Published: July 2002

The increasing prevalence of audit in university settings has raised concerns about the potentially adverse effects of invasive measures of performance upon the conduct of research and generation of knowledge. What sustains the current commitment to audit? It is argued that in order to address this question we need to understand how and to what extent notions of accountability are played out in practice. This is illustrated through the analysis of materials from an ethnographic study of ‘good management practice’ in the deployment of technologies in university settings. The paper examines the ways in which ideas of accountability — involving considerations such as ‘value for money’— inform the practical processes of deciding about the purchase of a new database technology.
Japanese body image: Structure and esteem scores in a cross‐cultural perspective
Authors: Kowner Rotem
Published: July 2002

Culture appears to affect body image in general and body esteem in particular, yet do cultural differences in these constructs concern with factorial structure or merely the magnitude of their manifestation? This study examined what body parts and functions the body esteem of young Japanese adults consists of, and assessed its relation with several other construals of the self. A secondary goal was to compare scores of body esteem in Japan with data previously obtained for people of similar age and background in diverse cultures, such as the USA, Hong Kong, and Israel. The primary contention of this study was that body image might vary notably across different cultures. Because of cultural differences in the self, and indirectly also due to physiognomic variation, members of various cultures may differ in the way they conceive their own body, have divergent body ideals, and ultimately experience different feelings toward their body. The subjects were 569 Japanese undergraduates who filled in the Body Esteem Scale (BES) as well as measures of self‐esteem, body consciousness, and social anxiety. Findings show fairly similar structure of body esteem to that found in the USA, but lower ratings of body esteem among Japanese than among their American, Chinese, and Israeli counterparts. In addition, findings indicate a large gender difference on the total BES score: Men expressed higher body esteem than women. Among both genders, however, body esteem correlated positively with self‐esteem and body consciousness, but negatively with social anxiety. It is suggested that the structure similarity between Japanese and American body esteem is the result of universal human mating patterns as well as similar personality structure. The reason for the lower Japanese body esteem may involve general tendency for self‐effacement and social anxiety. Further causes for the lower scores, which may concern broader cultural and historical perspectives, are briefly discussed.
Prayers, Pilgrimages and Petitions: The Secret Church and the Growth of Civil Society in Slovakia
Authors: Doellinger David
Published: July 2002

A photograph of Pope John Paul II shaking hands with Ján Čarnogurský, First Deputy Prime Minister of Czechoslovakia, at the Vatican appeared in full color on the cover of the February 1990 issue ofRodinné spolocčenstvo.Čarnogurský symbolizes the speed of Czechoslovakia's political revolution and the important role that individuals who had gained political experience as dissidents played in Czechoslovakia's post-Communist government. Just 2 months before meeting with the Pope, Čarnogurský, a Roman Catholic activist in Slovakia, had been awaiting trial in Bratislava for editing the Slovak secret church's most politically-orientedsamizdatperiodical. Hundreds of demonstrators, organized by the Slovak secret church, had already been protesting his arrest for several weeks when the Velvet Revolution began in Prague on 17 November 1989.
Paap and Noel revisited: Individual differences in independent word recognition tasks in relation to the dual-task paradigm
Authors: Hayes Maureen , Masterson Jackie
Published: July 2002

Paap and Noel (1991) found that participants' pronunciation latencies were faster for low-frequency irregular words when named under a concurrent high digit memory load than when named under a low load. The effects reported by Paap and Noel have proved difficult to replicate in subsequent studies. The present research suggests that individual differences in word recognition skill relate to who will or will not show these effects. In two experiments, participants were allocated to skill groups on the basis of latency in tasks tapping lexical and sub-lexical reading processes. In both studies, only one group (the “intermediate skilled”), showed evidence of such effects. A combined analysis of the data from Experiments 1 and 2 was carried out. This confirmed and extended the analyses of the individual experiments. The results are discussed in relation to contemporary models of word recognition.
How predictable is spelling? Developing and testing metrics of phoneme-grapheme contingency
Authors: Perry Conrad , Ziegler Johannes C. , Coltheart Max
Published: July 2002

Prior research has purported to show that words with infrequent phoneme-grapheme correspondences are more difficult to spell than words with frequent phoneme-grapheme correspondences. Defining exactly what a phoneme-grapheme relationship is, however, is not necessarily straightforward. There are a number of different assumptions that can be made. In this study, we developed four metrics of sound-spelling contingency based on all monosyllabic English words, including those with complex morphology. These metrics differed in the extent to which they included assumptions about spelling. The psychological reality of these different metrics was evaluated against data from a large-scale study of skilled adult spelling and a nonword spelling experiment. The results suggest that when spelling, people are sensitive to positional information, morphological status, vowel type, and a number of more idiosyncratic constraints.
Comments on the Roundtable Discussion: The Progress of Heterodox Economics
Authors: Fusfeld Daniel R.
Published: July 2002

Heterodox economics today is alive and well, and will always be with us. It is generated by opposition to mainstream orthodoxy and performs the important function of preventing the mainstream from stagnating. Orthodoxy and heterodoxy feed off each other, and both are transformed in the process.
The Problem of Liberty in the thought of Adam Smith
Authors: Harpham Edward J.
Published: July 2002

In this paper I propose to investigate the problem of liberty in Adam Smith's work. Suggesting that there is a “problem” may strike some as strange. After all, is not Smith simply the great defender of the system of natural liberty, a set of economic proposals that would remove the State from the business of directing the economy? Does he not maintain unequivocally that individuals are the best judges of their own self-interest and argue that they should be allowed to commit their labor and capital to those enterprises they deem most useful? Is Smith not one of the great defenders of the concept of negative liberty in modern liberal thought?
Vocational education and labour markets in deregulated, flexibly coordinated and planned societies
Authors: Szydlik Marc
Published: July 2002

This article deals with consequences of different educational and labour market regimes for the matching of people to jobs. It differentiates between flexibly coordinated, planned and deregulated societies, focusing on the Federal Republic of Germany, the German Democratic Republic and the United States of America. The empirical analyses are based on the German Socio-Economic Panel (GSOEP) and the Panel Study of Income Dynamics (PSID). The analyses confirm most of the hypotheses. In all three countries, many employees have to deal with a mismatch between their qualification and the requirements of their jobs. The number of employees with a mismatch is considerably higher in the United States. However, this applies especially to young employees. In Germany, young people have a much better chance of using most of their qualifications in their first jobs, but their position in the labour market will worsen as they grow older. As expected, overqualified people earn considerably less than their colleagues who are working in adequate jobs. However, education is still worthwhile even if one is overqualified.
Transmission of cell stress from endoplasmic reticulum to mitochondria
Authors: Hori Osamu , Ichinoda Fusae , Tamatani Takashi , Yamaguchi Atsushi , Sato Naoya , Ozawa Kentaro , Kitao Yasuko , Miyazaki Mayuki , Harding Heather P. , Ron David , Tohyama Masaya , M Stern David , Ogawa Satoshi
Published: July 2002

The rat homologue of a mitochondrial ATP-dependent protease Lon was cloned from cultured astrocytes exposed to hypoxia. Expression of Lon was enhanced in vitro by hypoxia or ER stress, and in vivo by brain ischemia. These observations suggested that changes in nuclear gene expression (Lon) triggered by ER stress had the potential to impact important mitochondrial processes such as assembly and/or degradation of cytochrome c oxidase (COX). In fact, steady-state levels of nuclear-encoded COX IV and V were reduced, and mitochondrial-encoded subunit II was rapidly degraded under ER stress. Treatment of cells with cycloheximide caused a similar imbalance in the accumulation of COX subunits, and enhanced mRNA for Lon and Yme1, the latter another mitochondrial ATP-dependent protease. Furthermore, induction of Lon or GRP75/mtHSP70 by ER stress was inhibited in PERK (−/−) cells. Transfection studies revealed that overexpression of wild-type or proteolytically inactive Lon promoted assembly of COX II into a COX I–containing complex, and partially prevented mitochondrial dysfunction caused by brefeldin A or hypoxia. These observations demonstrated that suppression of protein synthesis due to ER stress has a complex effect on the synthesis of mitochondrial-associated proteins, both COX subunits and ATP-dependent proteases and/or chaperones contributing to assembly of the COX complex.
Functional binding interaction identified between the axonal CAM L1 and members of the ERM family
Authors: Dickson Tracey C. , Mintz C. David , Benson Deanna L. , Salton Stephen R.J.
Published: July 2002

Ayeast two-hybrid library was screened using the cytoplasmic domain of the axonal cell adhesion molecule L1 to identify binding partners that may be involved in the regulation of L1 function. The intracellular domain of L1 bound to ezrin, a member of the ezrin, radixin, and moesin (ERM) family of membrane–cytoskeleton linking proteins, at a site overlapping that for AP2, a clathrin adaptor. Binding of bacterial fusion proteins confirmed this interaction. To determine whether ERM proteins interact with L1 in vivo, extracellular antibodies to L1 were used to force cluster the protein on cultured hippocampal neurons and PC12 cells, which were then immunolabeled for ERM proteins. Confocal analysis revealed a precise pattern of codistribution between ERMs and L1 clusters in axons and PC12 neurites, whereas ERMs in dendrites and spectrin labeling remained evenly distributed. Transfection of hippocampal neurons grown on an L1 substrate with a dominant negative ERM construct resulted in extensive and abnormal elaboration of membrane protrusions and an increase in axon branching, highlighting the importance of the ERM–actin interaction in axon development. Together, our data indicate that L1 binds directly to members of the ERM family and suggest this association may coordinate aspects of axonal morphogenesis.