bluuebunny/crossref_metadata_2025 huggingface dataset:
IL-18 contributes to the spontaneous development of atopic dermatitis-like inflammatory skin lesion independently of IgE/stat6 under specific pathogen-free conditions
Authors: Konishi Hiroshi , Tsutsui Hiroko , Murakami Takaaki , Yumikura-Futatsugi Shizue , Yamanaka Kei-ichi , Tanaka Minoru , Iwakura Yohichiro , Suzuki Noboru , Takeda Kiyoshi , Akira Shizuo , Nakanishi Kenji , Mizutani Hitoshi
Published: September 2002

Atopic dermatitis (AD) is a pruritic inflammatory skin disease. Because IL-18 directly stimulates T cells and mast cells to release AD-associated molecules, Th2 cytokines, and histamine, we investigated the capacity of IL-18 to induce AD-like inflammatory skin disease by analyzing KIL-18Tg and KCASP1Tg, which skin-specifically overexpress IL-18 and caspase-1, respectively. They spontaneously developed relapsing dermatitis with mastocytosis and Th2 cytokine accumulation accompanied by systemic elevation of IgE and histamine. Stat6-deficient KCASP1Tg displayed undetectable levels of IgE but manifested the same degree of cutaneous changes, whereas IL-18-deficient KCASP1Tg evaded the dermatitis, suggesting that IL-18 causes the skin changes in the absence of IgE/stat6. KIL-18Tg and IL-1-deficient KCASP1Tg took longer to display the lesion than KCASP1Tg. Thus, AD-like inflammation is initiated by overrelease of IL-18 and accelerated by IL-1. Our present study might provide insight into understanding the pathogenesis of and establishing therapeutics for chronic inflammatory skin diseases including AD.
α-Synucleinopathy and selective dopaminergic neuron loss in a rat lentiviral-based model of Parkinson's disease
Authors: Lo Bianco C. , Ridet J-L. , Schneider B. L. , Déglon N. , Aebischer P.
Published: September 2002

Parkinson's disease (PD) is characterized by the progressive loss of substantia nigra dopaminergic neurons and the presence of cytoplasmic inclusions named Lewy bodies. Two missense mutations of the α-synuclein (α-syn; A30P and A53T) have been described in several families with an autosomal dominant form of PD. α-Syn also constitutes one of the main components of Lewy bodies in sporadic cases of PD. To develop an animal model of PD, lentiviral vectors expressing different human or rat forms of α-syn were injected into the substantia nigra of rats. In contrast to transgenic mice models, a selective loss of nigral dopaminergic neurons associated with a dopaminergic denervation of the striatum was observed in animals expressing either wild-type or mutant forms of human α-syn. This neuronal degeneration correlates with the appearance of abundant α-syn-positive inclusions and extensive neuritic pathology detected with both α-syn and silver staining. Lentiviral-mediated expression of wild-type or mutated forms of human α-syn recapitulates the essential neuropathological features of PD. Rat α-syn similarly leads to protein aggregation but without cell loss, suggesting that inclusions are not the primary cause of cell degeneration in PD. Viral-mediated genetic models may contribute to elucidate the mechanism of α-syn-induced cell death and allow the screening of candidate therapeutic molecules.
A unified model of NMDA receptor-dependent bidirectional synaptic plasticity
Authors: Shouval Harel Z. , Bear Mark F. , Cooper Leon N
Published: September 2002

Synapses in the brain are bidirectionally modifiable, but the routes of induction are diverse. In various experimental paradigms,
N
-methyl-
d
-aspartate receptor-dependent long-term depression and long-term potentiation have been induced selectively by varying the membrane potential of the postsynaptic neurons during presynaptic stimulation of a constant frequency, the rate of presynaptic stimulation, and the timing of pre- and postsynaptic action potentials. In this paper, we present a mathematical embodiment of bidirectional synaptic plasticity that is able to explain diverse induction protocols with a fixed set of parameters. The key assumptions and consequences of the model can be tested experimentally; further, the model provides the foundation for a unified theory of
N
-methyl-
d
-aspartate receptor-dependent synaptic plasticity.
Extinction rates under nonrandom patterns of habitat loss
Authors: Seabloom Eric W. , Dobson Andy P. , Stoms David M.
Published: September 2002

Most models that examine the effects of habitat conversion on species extinctions assume that habitat conversion occurs at random. This assumption allows predictions about extinction rates based on the species–area relationship. We show that the spatially aggregated nature of habitat conversion introduces a significant bias that may lead species-loss rates to exceed those predicted by species–area curves. Correlations between human activity and major compositional gradients, or species richness, also alter predicted species extinction rates. We illustrate the consequences of nonrandom patterns of habitat conversion by using a data set that combines the distribution of native vascular plants with human activity patterns in California.
Dimerization of cotton fiber cellulose synthase catalytic subunits occurs via oxidation of the zinc-binding domains
Authors: Kurek Isaac , Kawagoe Yasushi , Jacob-Wilk Deborah , Doblin Monika , Delmer Deborah
Published: September 2002

Cellulose synthase (CesA) proteins are components of CesA complexes (rosettes) and are thought to catalyze the chain elongation step in glucan polymerization. Little is understood about rosette assembly, including how CesAs interact with each other or with other components within the complexes. The first conserved region at the N terminus of plant CesA proteins contains two putative zinc fingers that show high homology to the RING-finger motif. We show that this domain in GhCesA1 can bind two atoms of Zn
2+
, as predicted by its structure. Analysis in the yeast two-hybrid system indicates that the N-terminal portions of cotton fiber GhCesA1 and GhCesA2 containing these domains can interact to form homo- or heterodimers. Although Zn
2+
binding occurs only when the protein is in the reduced form, biochemical analyses show that under oxidative conditions, the GhCesA1 zinc-finger domain and also the full-length protein dimerize via intermolecular disulfide bonds, indicating CesA dimerization can be regulated by redox state. We also provide evidence that the herbicide CGA 325′615 (Syngenta, Basel), which inhibits synthesis of crystalline cellulose and leads to a disruption of rosette architecture, may affect the oxidative state of the zinc-finger domain that is necessary for rosette stability. Taken together, these results support a model in which at least part of the process of rosette assembly and function may involve oxidative dimerization between CesA subunits.
Can patient self-management help explain the SES health gradient?
Authors: Goldman Dana P. , Smith James P.
Published: September 2002

There are large differences in health outcomes by socioeconomic status (SES) that cannot be explained fully by traditional arguments, such as access to care or poor health behaviors. We consider a different explanation—better self-management of disease by the more educated. We examine differences by education in treatment adherence among patients with two illnesses, diabetes and HIV, and then assess the subsequent impact of differential adherence on health status. One unique component of this research is that for diabetes we combine two different surveys—one cohort study and one randomized clinical trial—that are usually used exclusively by either biomedical or/and social scientists separately. For both illnesses, we find significant effects of adherence that are much stronger among patients with high SES. After controlling for other factors, more educated HIV+ patients are more likely to adhere to therapy, and this adherence made them experience improvements in their self-reported general health. Similarly, among diabetics, the less educated were much more likely to switch treatment, which led to worsening general health. In the randomized trial setting, intensive treatment regimens that compensated for poor adherence led to better improvements in glycemic control for the less educated. Among two distinct chronic illnesses, the ability to maintain a better health regimen is an important independent determinant of subsequent health outcomes. This finding is robust across clinical trial and population-based settings. Because this ability varies by schooling, self-maintenance is an important reason for the steep SES gradient in health outcomes.
Designability of α-helical proteins
Authors: Emberly Eldon G. , Wingreen Ned S. , Tang Chao
Published: September 2002

A typical protein structure is a compact packing of connected α-helices and/or β-strands. We have developed a method for generating the ensemble of compact structures a given set of helices and strands can form. The method is tested on structures composed of four α-helices connected by short turns. All such natural four-helix bundles that are connected by short turns seen in nature are reproduced to closer than 3.6 Å per residue within the ensemble. Because structures with no natural counterpart may be targets for
ab initio
structure design, the designability of each structure in the ensemble—defined as the number of sequences with that structure as their lowest-energy state—is evaluated using a hydrophobic energy. For the case of four α-helices, a small set of highly designable structures emerges, most of which have an analog among the known four-helix fold families; however, several packings and topologies with no analogs in protein database are identified.
Transitions between dynamical states of differing stability in the human brain
Authors: Meyer-Lindenberg Andreas , Ziemann Ulf , Hajak Göran , Cohen Leonardo , Berman Karen Faith
Published: September 2002

What mechanisms underlie the flexible formation, adaptation, synchronization, and dissolution of large-scale neural assemblies from the 10
10
densely interconnected, continuously active neurons of the human brain? Nonlinear dynamics provides a unifying perspective on self-organization. It shows that the emergence of patterns in open, nonequilibrium systems is governed by their stability in response to small disturbances and predicts macroscopic transitions between patterns of differing stability. Here, we directly demonstrate that such transitions can be elicited in the human brain by interference at the neural level. As a probe, we used a classic motor coordination paradigm exhibiting well described movement states of differing stability. Functional neuroimaging identified premotor (PMA) and supplementary motor (SMA) cortices as having neural activity linked to the degree of behavioral instability. These regions then were transiently disturbed with graded transcranial magnetic stimulation, which caused sustained and macroscopic behavioral transitions from the less stable out-of-phase to the stable in-phase movement, whereas the stable pattern could not be affected. Moreover, the strength of the disturbance needed (a measure of neural stability) was linked to the degree of behavioral stability, demonstrating the applicability of nonlinear system theory as a powerful predictor of the dynamical repertoire of the human brain.
Peptide blockade of HIFα degradation modulates cellular metabolism and angiogenesis
Authors: Willam Carsten , Masson Norma , Tian Ya-Min , Mahmood S. Aleema , Wilson Michael I. , Bicknell Roy , Eckardt Kai-Uwe , Maxwell Patrick H. , Ratcliffe Peter J. , Pugh Christopher W.
Published: September 2002

Hypoxia-inducible factor-1 (HIF) is a transcription factor central to oxygen homeostasis. It is regulated via its α isoforms. In normoxia they are ubiquitinated by the von Hippel-Lindau E3 ligase complex and destroyed by the proteasome, thereby preventing the formation of an active transcriptional complex. Oxygen-dependent enzymatic hydroxylation of either of two critical prolyl residues in each HIFα chain has recently been identified as the modification necessary for targeting by the von Hippel-Lindau E3 ligase complex. Here we demonstrate that polypeptides bearing either of these prolyl residues interfere with the degradative pathway, resulting in stabilization of endogenous HIFα chains and consequent up-regulation of HIF target genes. Similar peptides in which the prolyl residues are mutated are inactive. Induction of peptide expression in cell cultures affects physiologically important functions such as glucose transport and leads cocultured endothelial cells to form tubules. Coupling of these HIFα sequences to the HIV
tat
translocation domain allows delivery of recombinant peptide to cells with resultant induction of HIF-dependent genes. Injection of
tat
-HIF polypeptides in a murine sponge angiogenesis assay causes a markedly accelerated local angiogenic response and induction of glucose transporter-1 gene expression. These results demonstrate the feasibility of using these polypeptides to enhance HIF activity, opening additional therapeutic avenues for ischemic diseases.
High-resolution SNP mapping by denaturing HPLC
Authors: Nairz Knud , Stocker Hugo , Schindelholz Benno , Hafen Ernst
Published: September 2002

With the availability of complete genome sequences, new rapid and reliable strategies for positional cloning become possible. Single-nucleotide polymorphisms (SNPs) permit the mapping of mutations at a resolution not amenable to classical genetics. Here we describe a SNP mapping procedure that relies on resolving polymorphisms by denaturing HPLC without the necessity of determining the nature of the SNPs. With the example of mapping mutations to the
Drosophila
nicastrin
locus, we discuss the benefits of this method, evaluate the frequency of closely linked and potentially misleading second site mutations, and demonstrate the use of denaturing high-performance liquid chromatography to identify mutations in the candidate genes and to fine-map chromosomal breakpoints. Furthermore, we show that recombination events are not uniformly dispersed over the investigated region but rather occur at hot spots.
Closure of the Venus flytrap module of mGlu8 receptor and the activation process: Insights from mutations converting antagonists into agonists
Authors: Bessis Anne-Sophie , Rondard Philippe , Gaven Florence , Brabet Isabelle , Triballeau Nicolas , Prézeau Laurent , Acher Francine , Pin Jean-Philippe
Published: September 2002

Ca
2+
, pheromones, sweet taste compounds, and the main neurotransmitters glutamate and γ-aminobutyric acid activate G protein-coupled receptors (GPCRs) that constitute the GPCR family 3. These receptors are dimers, and each subunit has a large extracellular domain called a Venus flytrap module (VFTM), where agonists bind. This module is connected to a heptahelical domain that activates G proteins. Recently, the structure of the dimer of mGlu1 VFTMs revealed two important conformational changes resulting from glutamate binding. First, agonists can stabilize a closed state of at least one VFTM in the dimer. Second, the relative orientation of the two VFTMs in the dimer is different in the presence of glutamate, such that their C-terminal ends (which are connected to the G protein-activating heptahelical domain) become closer by more than 20 Å. This latter change in orientation has been proposed to play a key role in receptor activation. To elucidate the respective role of VFTM closure and the change in orientation of the VFTMs in family 3 GPCR activation, we analyzed the mechanism of action of the mGlu8 receptor antagonists ACPT-II and MAP4. Molecular modeling studies suggest that these two compounds prevent the closure of the mGlu8 VFTM because of ionic and steric hindrance, respectively. We show here that the replacement of the residues responsible for these hindrances (Asp-309 and Tyr-227, respectively) by Ala allows ACPT-II or MAP4 to fully activate the receptors. These data are consistent with the requirement of the VFTM closure for family 3 GPCR activation.
Y genetic data support the Neolithic demic diffusion model
Authors: Chikhi Lounès , Nichols Richard A. , Barbujani Guido , Beaumont Mark A.
Published: September 2002

There still is no general agreement on the origins of the European gene pool, even though Europe has been more thoroughly investigated than any other continent. In particular, there is continuing controversy about the relative contributions of European Palaeolithic hunter-gatherers and of migrant Near Eastern Neolithic farmers, who brought agriculture to Europe. Here, we apply a statistical framework that we have developed to obtain direct estimates of the contribution of these two groups at the time they met. We analyze a large dataset of 22 binary markers from the non-recombining region of the Y chromosome (NRY), by using a genealogical likelihood-based approach. The results reveal a significantly larger genetic contribution from Neolithic farmers than did previous indirect approaches based on the distribution of haplotypes selected by using post hoc criteria. We detect a significant decrease in admixture across the entire range between the Near East and Western Europe. We also argue that local hunter-gatherers contributed less than 30% in the original settlements. This finding leads us to reject a predominantly cultural transmission of agriculture. Instead, we argue that the demic diffusion model introduced by Ammerman and Cavalli-Sforza [Ammerman, A. J. & Cavalli-Sforza, L. L. (1984)
The Neolithic Transition and the Genetics of Populations in Europe
(Princeton Univ. Press, Princeton)] captures the major features of this dramatic episode in European prehistory.
Mycobacterial heparin-binding hemagglutinin and laminin-binding protein share antigenic methyllysines that confer resistance to proteolysis
Authors: Pethe Kevin , Bifani Pablo , Drobecq Hervé , Sergheraert Christian , Debrie Anne-Sophie , Locht Camille , Menozzi Franco D.
Published: September 2002

Mycobacterium tuberculosis
and
Mycobacterium
bovis
bacillus Calmette–Guérin produce a heparin-binding hemagglutinin adhesin (HBHA) required for extrapulmonary dissemination and a laminin-binding protein (LBP) involved in cytoadherence through laminin recognition. These adhesins bear posttranslational modifications that are not present when the proteins are produced in a recombinant (r) form in
Escherichia coli
. Mass spectrometry analysis of HBHA revealed that the posttranslational modifications are borne by the C-terminal moiety, which comprises the heparin-binding domain made of repeated lysine-rich motifs. Amino acid sequencing showed that these modifications consist of mono- and dimethyllysines within these motifs. The methyllysine-containing repeats were recognized by mAb 4057D2 and were also detected in LBP, which is equally recognized by mAb 4057D2. This Ab does not recognize the recombinant forms of these proteins. However, when rHBHA and rLBP were subjected to NaBH
4
and formalin treatment to induce lysine methylation, reactivity with mAb 4057D2 was recovered. Methylated rHBHA displayed enhanced resistance to proteolysis compared with rHBHA, as previously observed for native HBHA.
S
-adenosylmethionine-dependent HBHA methyltransferase activity was detected in the cell-wall fractions of
M. bovis
bacillus Calmette–Guérin and of
Mycobacterium
smegmatis
, a species that produces LBP but naturally lacks
hbhA
, suggesting that the same enzyme(s) methylate(s) both LBP and HBHA. This hypothesis was confirmed by the fact that HBHA produced by recombinant
M. smegmatis
was also methylated. These results show that mycobacteria use enzymatic methylation of lysines to ensure greater stability of their adhesins.
Identification of G protein-coupled receptors for
Drosophila
PRXamide peptides, CCAP, corazonin, and AKH supports a theory of ligand-receptor coevolution
Authors: Park Yoonseong , Kim Young-Joon , Adams Michael E.
Published: September 2002

G-protein coupled receptors (GPCRs) are ancient, ubiquitous sensors vital to environmental and physiological signaling throughout organismal life. With the publication of the
Drosophila
genome, numerous “orphan” GPCRs have become available for functional analysis. Here we characterize two groups of GPCRs predicted as receptors for peptides with a C-terminal amino acid sequence motif consisting of −PRXamide (PRXa). Assuming ligand-receptor coevolution, two alternative hypotheses were constructed and tested. The insect PRXa peptides are evolutionarily related to the vertebrate peptide neuromedin U (NMU), or are related to arginine vasopressin (AVP), both of which have PRXa motifs. Seven
Drosophila
GPCRs related to receptors for NMU and AVP were cloned and expressed in
Xenopus
oocytes for functional analysis. Four
Drosophila
GPCRs in the NMU group (CG11475, CG8795, CG9918, CG8784) are activated by insect PRXa pyrokinins, (−FXPRXamide), Cap2b-like peptides (−FPRXamide), or ecdysis triggering hormones (−PRXamide). Three
Drosophila
GPCRs in the vasopressin receptor group respond to crustacean cardioactive peptide (CCAP), corazonin, or adipokinetic hormone (AKH), none of which are PRXa peptides. These findings support a theory of coevolution for NMU and
Drosophila
PRXa peptides and their respective receptors.
Emergency response to a smallpox attack: The case for mass vaccination
Authors: Kaplan Edward H. , Craft David L. , Wein Lawrence M.
Published: September 2002

In the event of a smallpox bioterrorist attack in a large U.S. city, the interim response policy is to isolate symptomatic cases, trace and vaccinate their contacts, quarantine febrile contacts, but vaccinate more broadly if the outbreak cannot be contained by these measures. We embed this traced vaccination policy in a smallpox disease transmission model to estimate the number of cases and deaths that would result from an attack in a large urban area. Comparing the results to mass vaccination from the moment an attack is recognized, we find that mass vaccination results in both far fewer deaths and much faster epidemic eradication over a wide range of disease and intervention policy parameters, including those believed most likely, and that mass vaccination similarly outperforms the existing policy of starting with traced vaccination and switching to mass vaccination only if required.
Massive cross-modal cortical plasticity and the emergence of a new cortical area in developmentally blind mammals
Authors: Kahn Dianna M. , Krubitzer Leah
Published: September 2002

In the current investigation, the neurophysiological organization of the neocortex was examined in adult animals that were bilaterally enucleated very early in life, before the retino-geniculo-cortical pathway was established. Our results indicate that some aspects of development of cortical fields are not mediated by specific sensory inputs. However, the current study also demonstrates that peripheral innervation plays a large role in the organization of the neocortex, as cortical territories normally involved in visual processing are completely captured by the auditory and somatosensory system. Thus, a large degree of phenotypic variability in cortical organization can be accomplished solely by removing or modifying sensory inputs.
High diversity and rapid changeover of expressed
var
genes during the acute phase of
Plasmodium falciparum
infections in human volunteers
Authors: Peters Jennifer , Fowler Elizabeth , Gatton Michelle , Chen Nanhua , Saul Allan , Cheng Qin
Published: September 2002

Plasmodium falciparum
erythrocyte membrane protein 1 (PfEMP1) proteins expressed on the surface of
P. falciparum
-infected erythrocytes undergo antigenic variation by switching the gene expressed within a repertoire of approximately 50
var
genes per haploid genome. The switching of PfEMP1 plays an important role in the survival and pathogenesis of the parasite. To understand how a parasite switches its
var
gene expression in human infections, we investigated the composition and change of
var
gene transcripts during the acute phase of well-defined laboratory-induced
P. falciparum
infections in naïve human hosts. Multiple
var
transcripts, with the same dominant transcript, were identified in samples collected after three to four asexual-parasite cycles in two volunteers infected with cloned 3D7
P. falciparum
via mosquito bites. A major change in composition and frequency of
var
gene transcripts was observed between the culture used to infect the mosquitoes and the parasites recovered from the infected volunteers. A further change was seen when infected blood from a mosquito-infected volunteer was either passaged to other volunteers or cultured
in vitro
. The diversity of
var
transcripts did not increase with time. The results suggest that the switch of
var
gene expression is reinitiated after mosquito transmission and that
var
genes may rapidly switch from the first gene expressed after liver stage, but subsequent switching occurs at a much lower rate.
Mistargeting hippocampal axons by expression of a truncated Eph receptor
Authors: Yue Yong , Chen Zhi-Yong , Gale Nick W. , Blair-Flynn Jan , Hu Tian-Jing , Yue Xin , Cooper Margaret , Crockett David P. , Yancopoulos George D. , Tessarollo Lino , Zhou Renping
Published: September 2002

Topographic mapping of axon terminals is a general principle of neural architecture that underlies the interconnections among many neural structures. The Eph family tyrosine kinase receptors and their ligands, the ephrins, have been implicated in the formation of topographic projection maps. We show that multiple Eph receptors and ligands are expressed in the hippocampus and its major subcortical projection target, the lateral septum, and that expression of a truncated Eph receptor in the mouse brain results in a pronounced alteration of the hippocamposeptal topographic map. Our observations provide strong support for a critical role of Eph family guidance factors in regulating ontogeny of hippocampal projections.
Sonic hedgehog rescues cranial neural crest from cell death induced by ethanol exposure
Authors: Ahlgren Sara C. , Thakur Vijaya , Bronner-Fraser Marianne
Published: September 2002

Alcohol is a teratogen that induces a variety of abnormalities including brain and facial defects [Jones, K. & Smith, D. (1973)
Lancet
2, 999-1001], with the exact nature of the deficit depending on the time and magnitude of the dose of ethanol to which developing fetuses are exposed. In addition to abnormal facial structures, ethanol-treated embryos exhibit a highly characteristic pattern of cell death. Dying cells are observed in the premigratory and migratory neural crest cells that normally populate most facial structures. The observation that blocking Sonic hedgehog (Shh) signaling results in similar craniofacial abnormalities prompted us to examine whether there was a link between this aspect of fetal alcohol syndrome and loss of
Shh
. We demonstrate that administration of ethanol to chick embryos results in a dramatic loss of
Shh
, as well as a loss of transcripts involved in Shh signaling pathways. In contrast, other signaling molecules examined do not demonstrate such dramatic changes. Furthermore, we demonstrate that both the ethanol-induced cranial neural crest cell death and the associated craniofacial growth defect can be rescued by application of Shh. These data suggest that craniofacial anomalies resulting from fetal alcohol exposure are caused at least partially by loss of
Shh
and subsequent neural crest cell death.
In TH2 cells theIl4gene has a series of accessibility states associated with distinctive probabilities of IL-4 production
Authors: Guo Liying , Hu-Li Jane , Zhu Jinfang , Watson Cynthia J. , Difilippantonio Michael J. , Pannetier Christophe , Paul William E.
Published: September 2002

TH2 clones may produce very variable amounts of IL-4. Among six TH2 clones prepared from homozygous or heterozygous mice in whichGfpreplaced the first exon ofIl4,a range of patterns of CpG methylation in theIl4/Il13locus was observed correlating with the degree of expression of IL-4 or green fluorescence protein. Patterns of histone acetylation also showed differences between “high” and “low” TH2 clones. These results indicate that in TH2 cells theIl4locus may display variable patterns of chromatin accessibility associated with distinct degrees of IL-4 expression. This finding suggests a regulation of IL-4 expression keyed to the function of this cytokine in cell/cell interactions and in the regulation of threshold responses.
The crystal structure of human CD21: Implications for Epstein–Barr virus and C3d binding
Authors: Prota Andrea E. , Sage David R. , Stehle Thilo , Fingeroth Joyce D.
Published: September 2002

Human complement receptor type 2 (CD21) is the cellular receptor for Epstein–Barr virus (EBV), a human tumor virus. The N-terminal two short consensus repeats (SCR1–SCR2) of the receptor interact with the EBV glycoprotein gp350/220 and also with the natural CD21 ligand C3d. Here we present the crystal structure of the CD21 SCR1–SCR2 fragment in the absence of ligand and demonstrate that it is able to bind EBV. Based on a functional analysis of wild-type and mutant CD21 and molecular modeling, we identify a likely region for EBV attachment and demonstrate that this region is not involved in the interaction with C3d. A comparison with the previously determined structure of CD21 SCR1–SCR2 in complex with C3d shows that, in both cases, CD21 assumes compact V-shaped conformations. However, our analysis reveals a surprising degree of flexibility at the SCR1–SCR2 interface, suggesting interactions between the two domains are not specific. We present evidence that the V-shaped conformation is induced by deglycosylation of the protein, and that physiologic glycosylation of CD21 would result in a more extended conformation, perhaps with additional epitopes for C3d binding.
The multidrug transporter, P-glycoprotein, actively mediates cholesterol redistribution in the cell membrane
Authors: Garrigues Alexia , Escargueil Alexandre E. , Orlowski Stéphane
Published: September 2002

P-glycoprotein (P-gp) is a plasma membrane ATP-binding cassette transporter, responsible for multidrug resistance in tumor cells. P-gp catalyzes the ATP hydrolysis-dependent efflux of numerous amphiphilic compounds of unrelated chemical structures. In the absence of any identified substrate, P-gp exhibits an apparently futile, basal ATPase activity. By using native membrane vesicles containing high amounts of P-gp, we show here that (
i
) this basal ATPase activity is tightly dependent on the presence of cholesterol in the membrane; (
ii
) the stimulation of P-gp ATPase activity by drugs transported by P-gp is higher in the absence than in the presence of cholesterol and, conversely, the stimulation of P-gp ATPase activity by cholesterol is higher in the absence than in the presence of known P-gp substrates; (
iii
) P-gp mediates the ATP-dependent relocation of cholesterol from the cytosolic leaflet to the exoplasmic leaflet of the plasma membrane; and (
iv
) the decrease of the cholesterol dependence of P-gp ATPase activity induced by known P-gp substrates is correlated with the inhibition of the ATP-dependent cholesterol redistribution within the membrane. These data are highly evocative of a coupling between the basal ATPase activity of P-gp and its intramembrane cholesterol-redistribution function, and they are fully consistent with the possibility that P-gp may actively translocate cholesterol in the membrane. Finally, this P-gp-mediated cholesterol redistribution in the cell membrane makes it likely that P-gp contributes in stabilizing the cholesterol-rich microdomains, rafts and caveolae, and that it is involved in the regulation of cholesterol trafficking in cells.
Deletion of the M
5
muscarinic acetylcholine receptor attenuates morphine reinforcement and withdrawal but not morphine analgesia
Authors: Basile Anthony S. , Fedorova Irina , Zapata Agustin , Liu Xiaoguang , Shippenberg Toni , Duttaroy Alokesh , Yamada Masahisa , Wess Jürgen
Published: September 2002

Little is known about the physiological roles of the M
5
muscarinic receptor, the last member of the muscarinic receptor family (M
1
–M
5
) to be cloned. In the brain, the M
5
receptor subtype is preferentially expressed by dopaminergic neurons of the substantia nigra and the ventral tegmental area. Dopaminergic neurons located in the ventral tegmental area are known to play important roles in mediating both the rewarding effects of opiates and other drugs of abuse and the manifestations of opiate/drug withdrawal symptoms. We therefore speculated that acetylcholine-dependent activation of M
5
receptors might modulate the manifestations of opiate reward and withdrawal. This hypothesis was tested in a series of behavioral, biochemical, and neurochemical studies using M
5
receptor-deficient mice (M
5
−/−
mice) as novel experimental tools. We found that the rewarding effects of morphine, as measured in the conditioned place preference paradigm, were substantially reduced in M
5
−/−
mice. Furthermore, both the somatic and affective components of naloxone-induced morphine withdrawal symptoms were significantly attenuated in M
5
−/−
mice. In contrast, the analgesic efficacy of morphine and the development of tolerance to the analgesic effects of morphine remained unaltered by the lack of M
5
receptors. The finding that M
5
receptor activity modulates both morphine reward and withdrawal processes suggests that M
5
receptors may represent a novel target for the treatment of opiate addiction.
NusA-stimulated RNA polymerase pausing and termination participates in the
Bacillus subtilis trp
operon attenuation mechanism
in
vitro
Authors: Yakhnin Alexander V. , Babitzke Paul
Published: September 2002

The
trp
RNA-binding attenuation protein (TRAP) regulates expression of the
Bacillus subtilis trpEDCFBA
operon by transcription attenuation and translation control mechanisms. Both mechanisms require the binding of tryptophan-activated TRAP to the 11 (G/U)AG-repeat segment in the
trp
leader transcript. To promote termination, TRAP must bind to the nascent RNA before the antiterminator structure forms. Because only 20 nucleotides separate the TRAP-binding site from the 3′ end of the antiterminator, TRAP has a short time frame to control this regulatory decision. Synchronization of factor binding and/or RNA folding with the RNA polymerase position is a major challenge in all attenuation mechanisms. Because RNA polymerase pausing allows this synchronization in many attenuation mechanisms, we performed experiments
in vitro
to determine whether pausing participates in the
B. subtilis trp
attenuation mechanism. We identified two NusA-stimulated pause sites in the
trp
leader region. Formation of pause hairpins participates in pausing at both positions. The first pause occurred at the nucleotide just preceding the critical overlap between the alternative antiterminator and terminator structures. TRAP binding to transcripts containing preexisting pause complexes releases RNA polymerase, suggesting that pausing provides additional time for TRAP to bind and promote termination. The second pause is downstream from the
trp
leader termination point, raising the possibility that this pause event participates in the
trpE
translation control mechanism. NusA also increases the efficiency of termination in the
trp
leader region and shifts termination one nucleotide upstream. Finally, NusA-stimulated termination is cooperative, suggesting that binding of multiple NusA molecules influences termination.
Disruption of pre-TCR expression accelerates lymphomagenesis in E2A-deficient mice
Authors: Engel Isaac , Murre Cornelis
Published: September 2002

The helix–loop–helix proteins E47 and E12, which are encoded by the E2A gene, regulate several stages of T cell development. In addition, mice deficient for E2A are highly susceptible to thymic lymphoma. Here we report that the development of lymphoma in E2A-deficient mice did not require pre- and recombinase-activating gene expression. Rather, we found that, whereas illegitimate DNA rearrangement did not play a major role in the development of these lymphomas, defects that prevented pre-T cell antigen receptor expression tended to accelerate lymphomagenesis in E2A-deficient mice. These data and previous observations also provide insight into the role of Notch in lymphoma development. Specifically, we propose that Notch activation indirectly modulates E2A activity through induction of pre-Tα expression, ultimately leading to the development of lymphoma.
Wiskott–Aldrich syndrome protein is required for NK cell cytotoxicity and colocalizes with actin to NK cell-activating immunologic synapses
Authors: Orange Jordan S. , Ramesh Narayanaswamy , Remold-O'Donnell Eileen , Sasahara Yoji , Koopman Louise , Byrne Michael , Bonilla Francisco A. , Rosen Fred S. , Geha Raif S. , Strominger Jack L.
Published: September 2002

The Wiskott–Aldrich syndrome (WAS) is a primary immunodeficiency disorder caused by a mutation in WAS protein (WASp) that results in defective actin polymerization. Although the function of many hematopoietic cells requires WASp, the specific expression and function of this molecule in natural killer (NK) cells is unknown. Here, we report that WAS patients have increased percentages of peripheral blood NK cells and that fresh enriched NK cells from two patients with a WASp mutation have defective cytolytic function. In normal NK cells, WASp was expressed and localized to the activating immunologic synapse (IS) with filamentous actin (F-actin). Perforin also localized to the NK cell-activating IS but at a lesser frequency than F-actin and WASp. The accumulation of F-actin and WASp at the activating IS was decreased significantly in NK cells that had been treated with the inhibitor of actin polymerization, cytochalasin D. NK cells from WAS patients lacked expression of WASp and accumulated F-actin at the activating IS infrequently. Thus, WASp has an important function in NK cells. In patients with WASp mutations, the resulting NK cell defects are likely to contribute to their disease.
Mammalian mediator subunit mMED8 is an Elongin BC-interacting protein that can assemble with Cul2 and Rbx1 to reconstitute a ubiquitin ligase
Authors: Brower Christopher S. , Sato Shigeo , Tomomori-Sato Chieri , Kamura Takumi , Pause Arnim , Stearman Robert , Klausner Richard D. , Malik Sohail , Lane William S. , Sorokina Irina , Roeder Robert G. , Conaway Joan Weliky , Conaway Ronald C.
Published: September 2002

The heterodimeric Elongin BC complex has been shown to interact
in vitro
and in cells with a conserved BC-box motif found in an increasing number of proteins including RNA polymerase II elongation factor Elongin A, suppressor of cytokine signaling (SOCS)-box proteins, and the von Hippel–Lindau tumor suppressor protein. Recently, the Elongin BC complex was found to function as an adaptor that links these BC-box proteins to a module composed of Cullin family members Cul2 or Cul5 and RING-H2 finger protein Rbx1 to reconstitute a family of E3 ubiquitin ligases that activate ubiquitylation by the E2 ubiquitin-conjugating enzyme Ubc5. As part of our effort to understand the functions of Elongin BC-based ubiquitin ligases, we exploited a modified yeast two-hybrid screen to identify a mammalian BC-box protein similar in sequence to
Saccharomyces cerevisiae
Mediator subunit Med8p. In this report we demonstrate (
i
) that mammalian MED8 is a subunit of the mammalian Mediator complex and (
ii
) that MED8 can assemble with Elongins B and C, Cul2, and Rbx1 to reconstitute a ubiquitin ligase. Taken together, our findings are consistent with the model that MED8 could function to recruit ubiquitin ligase activity directly to the RNA polymerase II transcriptional machinery.
Multiple genetic loci modify susceptibility to plasmacytoma-related morbidity in Eμ-v-abltransgenic mice
Authors: Symons R. C. Andrew , Daly Mark J. , Fridlyand Jane , Speed Terence P. , Cook Wendy D. , Gerondakis Steven , Harris Alan W. , Foote Simon J.
Published: September 2002

There is a great difference in susceptibility to v-abltransgene-induced plasmacytoma between the BALB/cAn and the relatively resistant C57BL/6J mouse strains. We have used the Mapmaker/SURVIVOR algorithm to analyze genome-wide scans on over 800 transgenic F2hybrid mice, and have mapped at least six loci on chromosomes 2, 4, 11, 17, and 18 that modify tumor-related morbidity. As in human multiple myeloma, males were found to be more prone to plasmacytomagenesis. Different loci influence tumor susceptibility in male and female mice. Survival in females may be largely controlled by a pair of interacting loci on chromosomes 2 and 17.
Cell and chloroplast division requires ARTEMIS
Authors: Fulgosi Hrvoje , Gerdes Lars , Westphal Sabine , Glockmann Christel , Soll Jürgen
Published: September 2002

Chloroplasts are endosymbiotic organelles of cyanobacterial origin. It seems reasonable to assume that cell division and organelle division still share general principles, as shown for the FtsZ proteins. However, further components involved in this process are largely unknown. Here we describe ARTEMIS, a nuclear-encoded protein of chloroplast inner envelope membranes that is required for organelle division. ARTEMIS consists of three distinct modules: an N-terminal receptor-like region, a centrally positioned glycine-rich stretch containing a nucleoside triphosphate-binding site, and a C-terminal YidC/Oxa1p/Alb3 protein translocase-like domain. Analysis of
Arabidopsis En-1
transposon mutants as well as ARTEMIS antisense plants revealed chloroplasts arrested in the late stages of division. Chloroplasts showed clearly separated and distinct multiple thylakoid systems, whereas the final organelle fission remained unaccomplished. Inactivation of a cyanobacterial gene with sequence similarity to the YidC/Oxa1p/Alb3-like domain of ARTEMIS resulted in aberrant cell division, which could be rescued by the
Arabidopsis
protein. ARTEMIS represents a so-far-unrecognized link between prokaryotic cell fission and chloroplast division.
CREB activity in the nucleus accumbens shell controls gating of behavioral responses to emotional stimuli
Authors: Barrot Michel , Olivier Jocelien D. A. , Perrotti Linda I. , DiLeone Ralph J. , Berton Olivier , Eisch Amelia J. , Impey Soren , Storm Daniel R. , Neve Rachael L. , Yin Jerry C. , Zachariou Venetia , Nestler Eric J.
Published: September 2002

The transcription factor cAMP response element (CRE)-binding protein (CREB) has been shown to regulate neural plasticity. Drugs of abuse activate CREB in the nucleus accumbens, an important part of the brain's reward pathways, and local manipulations of CREB activity have been shown to affect cocaine reward, suggesting an active role of CREB in adaptive processes that follow exposure to drugs of abuse. Using CRE-LacZ reporter mice, we show that not only rewarding stimuli such as morphine, but also aversive stimuli such as stress, activate CRE-mediated transcription in the nucleus accumbens shell. Using viral-mediated gene transfer to locally alter the activity of CREB, we show that this manipulation affects morphine reward, as well as the preference for sucrose, a more natural reward. We then show that local changes in CREB activity induce a more general syndrome, by altering reactions to anxiogenic, aversive, and nociceptive stimuli as well. Increased CREB activity in the nucleus accumbens shell decreases an animal's responses to each of these stimuli, whereas decreased CREB activity induces an opposite phenotype. These results show that environmental stimuli regulate CRE-mediated transcription within the nucleus accumbens shell, and that changes in CREB activity within this brain area subsequently alter gating between emotional stimuli and their behavioral responses. This control appears to be independent of the intrinsic appetitive or aversive value of the stimulus. The potential relevance of these data to addiction and mood disorders is discussed.
Angiopoietin-2 displays VEGF-dependent modulation of capillary structure and endothelial cell survival
in vivo
Authors: Lobov Ivan B. , Brooks Peter C. , Lang Richard A.
Published: September 2002

Modulation of Tie2 receptor activity by its angiopoietin ligands is crucial for angiogenesis, blood vessel maturation, and vascular endothelium integrity. It has been proposed that angiopoietins 1 (Ang1) and 2 (Ang2) are pro- and anti-angiogenic owing to their respective agonist and antagonist signaling action through the Tie2 receptor. The function of Ang2 has remained controversial, however, with recent reports suggesting that in some circumstances, it may be pro-angiogenic. We have examined this issue using the transient ocular microvessel network called the pupillary membrane as a unique
in vivo
model for studying the effects of vascular regulators. We show that
in vivo
, in the presence of endogenous vascular endothelial growth factor (VEGF)-A, Ang2 promotes a rapid increase in capillary diameter, remodeling of the basal lamina, proliferation and migration of endothelial cells, and stimulates sprouting of new blood vessels. By contrast, Ang2 promotes endothelial cell death and vessel regression if the activity of endogenous VEGF is inhibited. These observations support a model for regulation of vascularity where VEGF can convert the consequence of Ang2 stimulation from anti- to pro-angiogenic.
Sucrose-mediated transcriptional regulation of sucrose symporter activity in the phloem
Authors: Vaughn Matthew W. , Harrington Gregory N. , Bush Daniel R.
Published: September 2002

A proton–sucrose symporter mediates the key step in carbon export from leaves of most plants. Sucrose transport activity and steady-state mRNA levels of
BvSUT1
, a sugar beet leaf sucrose symporter, are negatively regulated specifically by sucrose. Results reported here show that
BvSUT1
mRNA was localized to companion cells of the leaf's vascular system, which supports its role in the systemic distribution of photoassimilate. Immunoblot analysis showed that decreased transport activity was caused by a reduction in the abundance of symporter protein. RNA gel blot analysis of the leaf symporter revealed that message levels also declined, and nuclear run-on experiments demonstrated that this was the result of decreased transcription. Further analysis showed that symporter protein and message are both degraded rapidly. Taken together, these data show that phloem loading is regulated by means of sucrose-mediated changes in transcription of a phloem-specific sucrose symporter gene in a regulatory system that may play a pivotal role in balancing photosynthetic activity with resource utilization.