bluuebunny/crossref_metadata_2025 huggingface dataset:
Interferon γ (IFN-γ) Is Necessary for the Genesis of Acetylcholine Receptor–induced Clinical Experimental Autoimmune Myasthenia gravis in Mice
Authors: Balasa Balaji , Deng Caishu , Lee Jae , Bradley Linda M. , Dalton Dyana K. , Christadoss Premkumar , Sarvetnick Nora
Published: July 2002

Experimental autoimmune myasthenia gravis (EAMG) is an animal model of human myasthenia gravis (MG). In mice, EAMG is induced by immunization with Torpedo californica acetylcholine receptor (AChR) in complete Freund's adjuvant (CFA). However, the role of cytokines in the pathogenesis of EAMG is not clear. Because EAMG is an antibody-mediated disease, it is of the prevailing notion that Th2 but not Th1 cytokines play a role in the pathogenesis of this disease. To test the hypothesis that the Th1 cytokine, interferon (IFN)-γ, plays a role in the development of EAMG, we immunized IFN-γ knockout (IFN-gko) (−/−) mice and wild-type (WT) (+/+) mice of H-2b haplotype with AChR in CFA. We observed that AChR-primed lymph node cells from IFN-gko mice proliferated normally to AChR and to its dominant pathogenic α146–162 sequence when compared with these cells from the WT mice. However, the IFN-gko mice had no signs of muscle weakness and remained resistant to clinical EAMG at a time when the WT mice exhibited severe muscle weakness and some died. The resistance of IFN-gko mice was associated with greatly reduced levels of circulating anti-AChR antibody levels compared with those in the WT mice. Comparatively, immune sera from IFN-gko mice showed a dramatic reduction in mouse AChR-specific IgG1 and IgG2a antibodies. However, keyhole limpet hemocyanin (KLH)–priming of IFN-gko mice readily elicited both T cell and antibody responses, suggesting that IFN-γ regulates the humoral immune response distinctly to self (AChR) versus foreign (KLH) antigens. We conclude that IFN-γ is required for the generation of a pathogenic anti-AChR humoral immune response and for conferring susceptibility of mice to clinical EAMG.
Epstein-Barr  Virus Binding to CD21 Activates the Initial Viral Promoter via NF-κB Induction
Authors: Sugano Naoyuki , Chen Weiping , Roberts M. Luisa , Cooper Neil R.
Published: July 2002

Epstein-Barr virus (EBV), an oncogenic human herpesvirus, binds to and infects normal human B lymphocytes via CD21, the CR2 complement receptor. Studies of the mechanisms that enable EBV to infect nonactivated, noncycling B cells provide compelling evidence for a sequence of events in which EBV binding to CD21 on purified resting human B cells rapidly activates the NF-κB transcription factor, which, in turn, binds to and mediates transcriptional activation of Wp, the initial viral latent gene promoter. Thus, EBV binding to its cellular receptor on resting B cells triggers an NF-κB–dependent intracellular signaling pathway which is required for infection.
Wound Healing Is Accelerated by Agonists of Adenosine A2 (Gαs-linked) Receptors
Authors: Montesinos M. Carmen , Gadangi Pratap , Longaker Michael , Sung Joanne , Levine Jamie , Nilsen Diana , Reibman Joan , Li Min , Jiang Chuan-Kui , Hirschhorn Rochelle , Recht Phoebe A. , Ostad Edward , Levin Richard I. , Cronstein Bruce N.
Published: July 2002

The complete healing of wounds is the final step in a highly regulated response to injury. Although many of the molecular mediators and cellular events of healing are known, their manipulation for the enhancement and acceleration of wound closure has not proven practical as yet. We and others have established that adenosine is a potent regulator of the inflammatory response, which is a component of wound healing. We now report that ligation of the Gαs-linked adenosine receptors on the cells of an artificial wound dramatically alters the kinetics of wound closure. Excisional wound closure in normal, healthy mice was significantly accelerated by topical application of the specific A2A receptor agonist CGS-21680 (50% closure by day 2 in A2 receptor antagonists. In rats rendered diabetic (streptozotocin-induced diabetes mellitus) wound healing was impaired as compared to nondiabetic rats; CGS-21680 significantly increased the rate of wound healing in both nondiabetic and diabetic rats. Indeed, the rate of wound healing in the CGS-21680–treated diabetic rats was greater than or equal to that observed in untreated normal rats. These results appear to constitute the first evidence that a small molecule, such as an adenosine receptor agonist, accelerates wound healing in both normal animals and in animals with impaired wound healing.
T Cell Receptor (TCR) Engagement Leads to Activation-induced Splicing of  Tumor Necrosis Factor (TNF) Nuclear Pre-mRNA
Authors: Yang Yang , Chang Ju-Fay , Parnes Jane R. , Garrison Fathman C.
Published: July 2002

Inducible gene expression is primarily regulated at the level of transcription. Additional steps of “processing” pre-mRNA, involved in the regulation of induced gene expression, have not been previously reported. Here we report a novel mechanism of “activation-induced splicing” of preexisting tumor necrosis factor (TNF) message (pre-mRNA) in naive T lymphocytes after engagement of the T cell receptor (TCR), which still occurs after inhibition of transcription. Expression of TNF has been previously demonstrated to be regulated at both the transcriptional and translational levels. However, neither the large pool of TNF mRNA observed in activated T cells nor TNF protein production, which peaks very shortly after activation, can be solely attributed to increased transcription. Evidence is presented that activation-induced splicing of TNF pre-mRNA plays a significant role in the rapid production of TNF seen in activated T cells. Activation triggers processing of TNF pre-mRNA that has accumulated in naive T cells (before activation-induced transcription), and the mature TNF mRNA is translocated to the cytoplasm for rapid translation and protein production. This novel form of activation-induced splicing of TNF may allow T cells to mount an immediate response to activation stimuli under physiological conditions.
Selective Expression and Functions of Interleukin 18 Receptor on T Helper (Th) Type 1 but not Th2 Cells
Authors: Xu Damo , Chan Woon Ling , Leung Bernard P. , Hunter David , Schulz Kerstin , Carter Robert W. , McInnes Iain B. , Robinson John H. , Liew Foo Y.
Published: July 2002

Interleukin (IL)-18 induces interferon (IFN)-γ synthesis and synergizes with IL-12 in T helper type 1 (Th1) but not Th2 cell development. We report here that IL-18 receptor (IL-18R) is selectively expressed on murine Th1 but not Th2 cells. IL-18R mRNA was expressed constitutively and consistently in long-term cultured clones, as well as on newly polarized Th1 but not Th2 cells. IL-18 sustained the expression of IL-12Rβ2 mRNA, indicating that IL-18R transmits signals that maintain Th1 development through the IL-12R complex. In turn, IL-12 upregulated IL-18R mRNA. Antibody against an IL-18R–derived peptide bound Th1 but not Th2 clones. It also labeled polarized Th1 but not Th2 cells derived from naive ovalbumin–T cell antigen receptor-αβ transgenic mice (D011.10). Anti–IL-18R antibody inhibited IL-18– induced IFN-γ production by Th1 clones in vitro. In vivo, anti–IL-18R antibody reduced local inflammation and lipopolysaccharide-induced mortality in mice. This was accompanied by shifting the balance from Th1 to Th2 responses, manifest as decreased IFN-γ and proinflammatory cytokine production and increased IL-4 and IL-5 synthesis. Therefore, these data provide a direct mechanism for the selective effect of IL-18 on Th1 but not Th2 cells. They also show that the synergistic effect of IL-12 and IL-18 on Th1 development may be due to the reciprocal upregulation of their receptors. Furthermore, IL-18R is a cell surface marker distinguishing Th1 from Th2 cells and may be a therapeutic target.
Transgenic Interleukin 10 Prevents Induction of Experimental Autoimmune Encephalomyelitis
Authors: Cua Daniel J. , Groux Herve , Hinton David R. , Stohlman Stephen A. , Coffman Robert L.
Published: July 2002

The effectiveness of interleukin 10 (IL-10) in the treatment of autoimmune-mediated central nervous system inflammation is controversial. Studies of the model system, experimental autoimmune encephalomyelitis (EAE), using various routes, regimens, and delivery methods of IL-10 suggest that these variables may affect its immunoregulatory function. To study the influence of these factors on IL-10 regulation of EAE pathogenesis, we have analyzed transgenic mice expressing human IL-10 (hIL-10) transgene under the control of a class II major histocompatibility complex (MHC) promoter. The hIL-10 transgenic mice are highly resistant to EAE induced by active immunization, and this resistance appears to be mediated by suppression of autoreactive T cell function. Myelin-reactive T helper 1 cells are induced but nonpathogenic in the IL-10 transgenic mice. Antibody depletion confirmed that EAE resistance is dependent on the presence of the transgenic IL-10. Mice expressing the hIL-10 transgene but not the endogenous murine IL-10 gene demonstrated that transgenic IL-10 from MHC class II–expressing cells is sufficient to block induction of EAE. This study demonstrates that IL-10 can prevent EAE completely if present at appropriate levels and times during disease induction.
Resistance of CD7-deficient Mice to Lipopolysaccharide-induced Shock Syndromes
Authors: Sempowski Gregory D. , Lee David M. , Scearce Richard M. , Patel Dhavalkumar D. , Haynes Barton F.
Published: July 2002

CD7 is an immunoglobulin superfamily molecule involved in T and natural killer (NK) cell activation and cytokine production. CD7-deficient animals develop normally but have antigen-specific defects in interferon (IFN)-γ production and CD8+ CTL generation. To determine the in vivo role of CD7 in systems dependent on IFN-γ, the response of CD7-deficient mice to lipopolysaccharide (LPS)-induced shock syndromes was studied. In the high-dose LPS-induced shock model, 67% of CD7-deficient mice survived LPS injection, whereas 19% of control C57BL/6 mice survived LPS challenge (P < 0.001). CD7-deficient or C57BL/6 control mice were next injected with low-dose LPS (1 μg plus 8 mg D-galactosamine [D-gal] per mouse) and monitored for survival. All CD7-deficient mice were alive 72 h after injection of LPS compared with 20% of C57BL/6 control mice (P < 0.001). After injection of LPS and D-gal, CD7-deficient mice had decreased serum IFN-γ and tumor necrosis factor (TNF)-α levels compared with control C57BL/6 mice (P < 0.001). Steady-state mRNA levels for IFN-γ and TNF-α in liver tissue were also significantly decreased in CD7-deficient mice compared with controls (P < 0.05). In contrast, CD7-deficient animals had normal liver interleukin (IL)-12, IL-18, and interleukin 1 converting enzyme (ICE) mRNA levels, and CD7-deficient splenocytes had normal IFN-γ responses when stimulated with IL-12 and IL-18 in vitro. NK1.1+/ CD3+ T cells are known to be key effector cells in the pathogenesis of toxic shock. Phenotypic analysis of liver mononuclear cells revealed that CD7-deficient mice had fewer numbers of liver NK1.1+/CD3+ T cells (1.5 ± 0.3 × 105) versus C57BL/6 control mice (3.7 ± 0.8 × 105; P < 0.05), whereas numbers of liver NK1.1+/CD3− NK cells were not different from controls. Thus, targeted disruption of CD7 leads to a selective deficiency of liver NK1.1+/ CD3+ T cells, and is associated with resistance to LPS shock. These data suggest that CD7 is a key molecule in the inflammatory response leading to LPS-induced shock.
Stem Cell Emergence and Hemopoietic Activity Are Incompatible in Mouse Intraembryonic Sites
Authors: Godin Isabelle , Garcia-Porrero Juan Antonio , Dieterlen-Lièvre Françoise , Cumano Ana
Published: July 2002

In the mouse embryo, the generation of candidate progenitors for long-lasting hemopoiesis has been reported in the paraaortic splanchnopleura (P-Sp)/aorta-gonad-mesonephros (AGM) region. Here, we address the following question: can the P-Sp/AGM environment support hemopoietic differentiation as well as generate stem cells, and, conversely, are other sites where hemopoietic differentiation occurs capable of generating stem cells? Although P-Sp/AGM generates de novo hemopoietic stem cells between 9.5 and 12.5 days post coitus (dpc), we show here that it does not support hemopoietic differentiation. Among mesoderm-derived sites, spleen and omentum were shown to be colonized by exogenous cells in the same fashion as the fetal liver. Cells colonizing the spleen were multipotent and pursued their evolution to committed progenitors in this organ. In contrast, the omentum, which was colonized by lymphoid-committed progenitors that did not expand, cannot be considered as a hemopoietic organ. From these data, stem cell generation appears incompatible with hemopoietic activity. At the peak of hemopoietic progenitor production in the P-Sp/AGM, between 10.5 and 11.5 dpc, multipotent cells were found at the exceptional frequency of 1 out of 12 total cells and 1 out of 4 AA4.1+ cells. Thus, progenitors within this region constitute a pool of undifferentiated hemopoietic cells readily accessible for characterization.
Selective Expansion of Cross-Reactive Cd8+ Memory T Cells by Viral Variants
Authors: Haanen John B.A.G. , Wolkers Monika C. , Kruisbeek Ada M. , Schumacher Ton N.M.
Published: July 2002

The role of memory T cells during the immune response against random antigenic variants has not been resolved. Here, we show by simultaneous staining with two tetrameric major histocompatibility complex (MHC)–peptide molecules, that the polyclonal CD8+ T cell response against a series of natural variants of the influenza A nucleoprotein epitope is completely dominated by infrequent cross-reactive T cells that expand from an original memory population. Based on both biochemical and functional criteria, these cross-reactive cytotoxic T cells productively recognize both the parental and the mutant epitope in vitro and in vivo. These results provide direct evidence that the repertoire of antigen-specific T cells used during an infection critically depends on prior antigen encounters, and indicate that polyclonal memory T cell populations can provide protection against a range of antigenic variants.
Human Nerve Growth Factor Protects Common Marmosets against Autoimmune Encephalomyelitis by Switching the Balance of T Helper Cell Type 1 and 2 Cytokines within the Central Nervous System
Authors: Villoslada Pablo , Hauser Stephen L. , Bartke Ilse , Unger Jurgen , Heald Nathan , Rosenberg Daniel , Cheung Steven W. , Mobley William C. , Fisher Stefan , Genain Claude P.
Published: July 2002

Multiple sclerosis is a demyelinating disorder of the central nervous system (CNS), in which an immune attack directed against myelin constituents causes myelin destruction and death of oligodendrocytes, the myelin-producing cells. Here, the efficacy of nerve growth factor (NGF), a growth factor for neurons and oligodendrocytes, in promoting myelin repair was evaluated using the demyelinating model of experimental allergic encephalomyelitis (EAE) in the common marmoset. Surprisingly, we found that NGF delayed the onset of clinical EAE and, pathologically, prevented the full development of EAE lesions. We demonstrate by immunocytochemistry that NGF exerts its antiinflammatory effect by downregulating the production of interferon γ by T cells infiltrating the CNS, and upregulating the production of interleukin 10 by glial cells in both inflammatory lesions of EAE and normal-appearing CNS white matter. Thus, NGF, currently under investigation in human clinical trials as a neuronal trophic factor, may be an attractive candidate for therapy of autoimmune demyelinating disorders.
Expression Cloning of an Immunodominant Family of Mycobacterium tuberculosis Antigens Using Human Cd4+ T Cells
Authors: Alderson Mark R. , Bement Teresa , Day Craig H. , Zhu Liqing , Molesh David , Skeiky Yasir A. W. , Coler Rhea , Lewinsohn David M. , Reed Steven G. , Dillon Davin C.
Published: July 2002

Development of a subunit vaccine for Mycobacterium tuberculosis (Mtb) is likely to be dependent on the identification of T cell antigens that induce strong proliferation and interferon γ production from healthy purified protein derivative (PPD)+ donors. We have developed a sensitive and rapid technique for screening an Mtb genomic library expressed in Escherichia coli using Mtb-specific CD4+ T cells. Using this technique, we identified a family of highly related Mtb antigens. The gene of one family member encodes a 9.9-kD antigen, termed Mtb9.9A. Recombinant Mtb9.9A protein, expressed and purified from E. coli, elicited strong T cell proliferation and IFN-γ production by peripheral blood mononuclear cells from PPD+ but not PPD− individuals. Southern blot analysis and examination of the Mtb genome sequence revealed a family of highly related genes. A T cell line from a PPD+ donor that failed to react with recombinant Mtb9.9A recognized one of the other family members, Mtb9.9C. Synthetic peptides were used to map the T cell epitope recognized by this line, and revealed a single amino acid substitution in this region when compared with Mtb9.9A. The direct identification of antigens using T cells from immune donors will undoubtedly be critical for the development of vaccines to several intracellular pathogens.
Low-level Hypermutation in T Cell–independent Germinal Centers Compared with High Mutation Rates Associated with T Cell–dependent Germinal Centers
Authors: Toellner Kai-Michael , Jenkinson William E. , Taylor Dale R. , Khan Mahmood , Sze Daniel M.-Y. , Sansom David M. , Vinuesa Carola G. , MacLennan Ian C.M.
Published: July 2002

Exceptionally germinal center formation can be induced without T cell help by polysaccharide-based antigens, but these germinal centers involute by massive B cell apoptosis at the time centrocyte selection starts. This study investigates whether B cells in germinal centers induced by the T cell–independent antigen (4-hydroxy-3-nitrophenyl)acetyl (NP) conjugated to Ficoll undergo hypermutation in their immunoglobulin V region genes. Positive controls are provided by comparing germinal centers at the same stage of development in carrier-primed mice immunized with a T cell–dependent antigen: NP protein conjugate. False positive results from background germinal centers and false negatives from non-B cells in germinal centers were avoided by transferring B cells with a transgenic B cell receptor into congenic controls not carrying the transgene. By 4 d after immunization, hypermutation was well advanced in the T cell–dependent germinal centers. By contrast, the mutation rate for T cell–independent germinal centers was low, but significantly higher than in NP-specific B cells from nonimmunized transgenic mice. Interestingly, a similar rate of mutation was seen in extrafollicular plasma cells at this stage. It is concluded that efficient activation of hypermutation depends on interaction with T cells, but some hypermutation may be induced without such signals, even outside germinal centers.
Pivotal Role of KARAP/DAP12 Adaptor Molecule in the Natural Killer Cell–mediated Resistance to Murine Cytomegalovirus Infection
Authors: Sjölin Hanna , Tomasello Elena , Mousavi-Jazi Mehrdad , Bartolazzi Armando , Kärre Klas , Vivier Eric , Cerboni Cristina
Published: July 2002

Natural killer (NK) cells are major contributors to early defense against infections. Their effector functions are controlled by a balance between activating and inhibiting signals. To date, however, the involvement of NK cell activating receptors and signaling pathways in the defense against pathogens has not been extensively investigated. In mice, several NK cell activating receptors are coexpressed with and function through the immunoreceptor tyrosine-based activation motif (ITAM)-bearing molecule KARAP/DAP12. Here, we have analyzed the role of KARAP/DAP12 in the early antiviral response to murine cytomegalovirus (MCMV). In KARAP/DAP12 mutant mice bearing a nonfunctional ITAM, we found a considerable increase in viral titers in the spleen (30–40-fold) and in the liver (2–5-fold). These effects were attributed to NK cells. The formation of hepatic inflammatory foci appeared similar in wild-type and mutant mice, but the latter more frequently developed severe hepatitis with large areas of focal necrosis. Moreover, the percentage of hepatic NK cells producing interferon γ was reduced by 56 ± 22% in the absence of a functional KARAP/DAP12. This is the first study that shows a crucial role for a particular activating signaling pathway, in this case the one induced through KARAP/DAP12, in the NK cell–mediated resistance to an infection. Our results are discussed in relation to recent reports demonstrating that innate resistance to MCMV requires the presence of NK cells expressing the KARAP/DAP12-associated receptor Ly49H.
Single Streptomyces lividans K+ Channels
Authors: Heginbotham Lise , LeMasurier Meredith , Kolmakova-Partensky Ludmilla , Miller Christopher
Published: July 2002

Basic electrophysiological properties of the KcsA K+ channel were examined in planar lipid bilayer membranes. The channel displays open-state rectification and weakly voltage-dependent gating. Tetraethylammonium blocking affinity depends on the side of the bilayer to which the blocker is added. Addition of Na+ to the trans chamber causes block of open-channel current, while addition to the cis side has no effect. Most striking is the activation of KcsA by protons; channel activity is observed only when the trans bilayer chamber is at low pH. To ascertain which side of the channel faces which chamber, residues with structurally known locations were mapped to defined sides of the bilayer. Mutation of Y82, an external residue, results in changes in tetraethylammonium affinity exclusively from the cis side. Channels with cysteine residues substituted at externally exposed Y82 or internally exposed Q119 are functionally modified by methanethiosulfonate reagents from the cis or trans chambers, respectively. Block by charybdotoxin, known to bind to the channel's external mouth, is observed only when the toxin is added to the cis side of channels mutated to be toxin sensitive. These results demonstrate unambiguously that the protonation sites linked to gating are on the intracellular portion of the KcsA protein.
Immobilizing the Moving Parts of Voltage-Gated Ion Channels
Authors: Horn Richard , Ding Shinghua , Gruber Hermann J.
Published: July 2002

Voltage-gated ion channels have at least two classes of moving parts, voltage sensors that respond to changes in the transmembrane potential and gates that create or deny permeant ions access to the conduction pathway. To explore the coupling between voltage sensors and gates, we have systematically immobilized each using a bifunctional photoactivatable cross-linker, benzophenone-4-carboxamidocysteine methanethiosulfonate, that can be tethered to cysteines introduced into the channel protein by mutagenesis. To validate the method, we first tested it on the inactivation gate of the sodium channel. The benzophenone-labeled inactivation gate of the sodium channel can be trapped selectively either in an open or closed state by ultraviolet irradiation at either a hyperpolarized or depolarized voltage, respectively. To verify that ultraviolet light can immobilize S4 segments, we examined its relative effects on ionic and gating currents in Shaker potassium channels, labeled at residue 359 at the extracellular end of the S4 segment. As predicted by the tetrameric stoichiometry of these potassium channels, ultraviolet irradiation reduces ionic current by approximately the fourth power of the gating current reduction, suggesting little cooperativity between the movements of individual S4 segments. Photocross-linking occurs preferably at hyperpolarized voltages after labeling residue 359, suggesting that depolarization moves the benzophenone adduct out of a restricted environment. Immobilization of the S4 segment of the second domain of sodium channels prevents channels from opening. By contrast, photocross-linking the S4 segment of the fourth domain of the sodium channel has effects on both activation and inactivation. Our results indicate that specific voltage sensors of the sodium channel play unique roles in gating, and suggest that movement of one voltage sensor, the S4 segment of domain 4, is at least a two-step process, each step coupled to a different gate.
New practices for electronic publishing 1: Will the scientific paper keep its form?
Authors: Kircz Joost G.
Published: July 2002

Discussion about the value of electronic documents is often hampered by starting from what is usual in the paper world and attempting to impose that on an electronic environment. In order to grasp the impact of the current electronic revolution, and formulate a policy for the future, we examine the aims and content of scientific communication. We then critically discuss the recommendations of an International Working Group [see Learned Publishing 2000:13(4) Oct. 251–8], and show the tension between these very reasonable recommendations and the reality of electronic publishing. We conclude that the scientific article will change considerably but that, in its new more composite form as an ensemble of various textual and non‐textual components, it will retain many of the current cultural and scientific requirements with regard to editorial, quality and integrity.
Hypercyclic operators that commute with the Bergman backward shift
Authors: Bourdon Paul , Shapiro Joel
Published: July 2002

The backward shift

B
B

on the Bergman space of the unit disc is known to be hypercyclic (meaning: it has a dense orbit). Here we ask: “Which operators that commute with

B
B

inherit its hypercyclicity?” We show that the problem reduces to the study of operators of the form

ϕ
(
B
)

\phi (B)

where

ϕ
\phi

is a holomorphic self-map of the unit disc that multiplies the Dirichlet space into itself, and that the question of hypercyclicity for such an operator depends on how freely

ϕ
(
z
)

\phi (z)

is allowed to approach the unit circle as

|

z

|

→
1
−

|z|\to 1-

.
Meshless Galerkin methods using radial basis functions
Authors: Wendland Holger
Published: July 2002

We combine the theory of radial basis functions with the field of Galerkin methods to solve partial differential equations. After a general description of the method we show convergence and derive error estimates for smooth problems in arbitrary dimensions.
Proof of the double bubble conjecture
Authors: Hutchings Michael , Morgan Frank , Ritoré Manuel , Ros Antonio
Published: July 2002

We prove that the standard double bubble provides the least-area way to enclose and separate two regions of prescribed volume in

R

3

{\mathbb R}^3

.
Proof of Conway’s lost cosmological theorem
Authors: Ekhad Shalosh , Zeilberger Doron
Published: July 2002

John Horton Conway’s Cosmological Theorem about sequences like 1, 11, 21, 1211, 111221, 312211,…, for which no extant proof existed, is given a new proof, this time hopefully for good.
Statistical mechanical simulation of polymeric DNA melting with MELTSIM.
Authors: Blake R D , Bizzaro J W , Blake J D , Day G R , Delcourt S G , Knowles J , Marx K A , SantaLucia J
Published: July 2002

MOTIVATION: MELTSIM is a windows-based statistical mechanical program for simulating melting curves of DNAs of known sequence and genomic dimensions under different conditions of ionic strength with great accuracy. The program is useful for mapping variations of base compositions of sequences, conducting studies of denaturation, establishing appropriate conditions for hybridization and renaturation, determinations of sequence complexity, and sequence divergence. RESULTS: Good agreement is achieved between experimental and calculated melting curves of plasmid, bacterial, yeast and human DNAs. Denaturation maps that accompany the calculated curves indicate non-coding regions have a significantly lower (G+C) composition than coding regions in all species examined. Curves of partially sequenced human DNA suggest the current database may be heavily biased with coding regions, and excluding large (A+T)-rich elements. AVAILABILITY: MELTSIM 1.0 is available at: //www.uml.edu/Dept/Chem/UMLBIC/Apps/MEL TSIM/MELTSIM-1.0-Win/meltsim. zip. Melting curve plots in this paper were made with GNUPLOT 3.5, available at: http://www.cs.dartmouth.edu/gnuplot_inf o.html Contact : blake@maine.maine.edu;
A historical perspective on gene/protein functional assignment
Authors: Hodgman T. C.
Published: July 2002

Sequence determination and analysis began on proteins in the 1950s, with RNA starting about a decade later and DNA a similar period later still. Hence many of the concepts for function prediction were first developed by looking at amino acid sequences. Over time these methods have become much more sophisticated, allowing better discrimination of only weak similarities. The most recent developments concern an examination of contextual information, such as operon structure, metabolic reconstruction or co-expression profiles.

Contact: ch18380@ggr.co.uk
JenPep: a database of quantitative functional peptide data for
immunology
Authors: Blythe Martin J. , Doytchinova Irini A. , Flower Darren R.
Published: July 2002

Motivation: The compilation of quantitative binding data underlies attempts to derive tools for the accurate prediction of epitopes in cellular immunology and is part of our concerted goal to develop practical computational vaccinology.

Results: JenPep is a family of relational databases supporting the growing community of immunoinformaticians. It contains quantitative data on peptide binding to Major Histocompatibility Complexes (MHCs) and to Transmembrane Peptide Transporter (TAP), as well as an annotated list of T-cell epitopes.

Availability: The database is available via the Internet. An HTML interface allowing searching of the database can be found at the following address: http://www.jenner.ac.uk/JenPep.

Contact: darren.flower@jenner.ac.uk

* To whom correspondence should be addressed.
Dating the common ancestor of SIVcpz and HIV‐1 group M and the origin of HIV‐1 subtypes by using a new method to uncover clock‐like molecular evolution
Authors: Salemi Marco , Strimmer Korbinian , Hall William W. , Duffy Margaret , Delaporte Eric , Mboup Souleymane , Peeters Martine , Vandamme Anne‐Mieke
Published: July 2002

Attempts to estimate the time of origin of human immunodeficiency virus (HIV)‐1 by using phylogenetic analysis are seriously flawed because of the unequal evolutionary rates among different viral lineages. Here, we report a new method of molecular clock analysis, called Site Stripping for Clock Detection (SSCD), which allows selection of nucleotide sites evolving at an equal rate in different lineages. The method was validated on a dataset of patients all infected with hepatitis C virus in 1977 by the same donor, and it was able to date exactly the ‘known’ origin of the infection. Using the same method, we calculated that the origin of HIV‐1 group M radiation was in the 1930s. In addition, we show that the coalescence time of the simian ancestor of HIV‐1 group M and its closest related cpz strains occurred around the end of the XVII century, a date that could be considered the upper limit to the time of simian‐to‐human transmission of HIV‐1 group M. The results show also that SSCD is an easy‐to‐use method of general applicability in molecular evolution to calibrate clock‐like phylogenetic trees.
Trophic structure stability and extinction dynamics of beetles (Coleoptera) in tropical forest fragments
Authors: Didham R. K. , Lawton J. H. , Hammond P. M. , Eggleton P.
Published: July 2002

A first analysis of the stability of trophic structure following tropical forest fragmentation was performed in an experimentally fragmented tropical forest landscape in Central Amazonia. A taxonomically and trophically diverse assemblage of 993 species of beetles was sampled from 920 m
2
of leaf litter at 46 sites varying in distance from forest edge and fragment area. Beetle density increased significantly towards the forest edge and showed non-linear changes with fragment area, due to the influx of numerous disturbed-area species into 10 ha and 1 ha fragments. There was a marked change in species composition with both decreasing distance from forest edge and decreasing fragment area, but surprisingly this change in composition was not accompanied by a change in species richness. Rarefied species richness did not vary significantly across any of the sites, indicating that local extinctions of deep forest species were balanced by equivalent colonization rates of disturbed-area species. The change in species composition with fragmentation was non-random across trophic groups. Proportions of predator species and xylophage species changed significantly with distance from forest edge, but no area-dependent changes in proportions of species in trophic groups were observed. Trophic structure was also analysed with respect to proportions of abundance in six trophic groups. Proportions of abundance of all trophic groups except xylomycetophages changed markedly with respect to both distance from forest edge and fragment area. Local extinction probabilities calculated for individual beetle species supported theoretical predictions of the differential susceptibility of higher trophic levels to extinction, and of changes in trophic structure following forest fragmentation. To reduce random effects due to sampling error, only abundant species (
n
≥ 46) were analysed for extinction probabilities, as defined by absence from samples. Of these common species, 27% had significantly higher probabilities of local extinction following fragmentation. The majority of these species were predators; 42% of all abundant predator species were significantly more likely to be absent from samples in forest fragments than in undisturbed forest. These figures are regarded as minimum estimates for the entire beetle assemblage because rarer species will inevitably have higher extinction probabilities. Absolute loss of biodiversity will affect ecosystem process rates, but the differential loss of species from trophic groups will have an even greater destabilizing effect on food web structure and ecosystem function.
Developmental biology and the redirection or replacement of cells
Authors: Gurdon John
Published: July 2002

The aim of developmental biology is to understand how an egg converts itself into a complete organism through the processes of cell differentiation, morphogenesis and size regulation. The principles that have emerged over recent decades include the constancy of the genome in nearly all cells of an individual, the existence of stem cells in many organs and the overwhelming importance of signalling between cells for the determination of their fate. These and other characteristics of development are discussed here in relation to the prospect of achieving cell and tissue correction or replacement with the help of nuclear transplantation and signalling factors. Nuclear transplantation offers a one–step procedure for generating multipotent embryo cells from the cells of an adult tissue such as skin. It should be possible to proliferate the resulting cells as can be done for mouse embryonic stem cells. Embryo cells can be made to differentiate in many directions by exposing them to various agents or to different concentrations of a single factor such as the transforming growth factor β class signalling molecule activin. The possibility of a cancerous condition being acquired during these experimental manipulations can be guarded against by transfecting cells with a conditional suicide gene. Thus it may be possible to generate replacement cells or tissues from an adult human for transplantation back to the original donor, without the disadvantage of any genetic incompatibility.
Reverse genetic studies of homologous DNA recombination using the chicken B–lymphocyte line, DT40
Authors: Sonoda Eiichiro , Morrison Ciaran , Yamashita Yukiko M. , Takata Minoru , Takeda Shunichi
Published: July 2002

DT40 is an avian leucosis virus–transformed chicken B–lymphocyte line which exhibits high ratios of targeted to random integration of transfected DNA constructs. This efficient targeted integration may be related to the ongoing diversification of the variable segment of the immunoglobulin gene through homologous DNA recombination–controlled gene conversion. DT40s are a convenient model system for making gene–targeted mutants. Another advantage is the relative tractability of these cells, which makes it possible to disrupt multiple genes in a single cell and to generate conditionally gene–targeted mutants including temperature–sensitive mutants. There are strong phenotypic similarities between murine and DT40 mutants of various genes involved in DNA recombination. These similarities confirm that the DT40 cell line is a reasonable model for the analysis of vertebrate DNA recombination, despite obvious concerns associated with the use of a transformed cell line, which may have certain cell–line–specific characteristics. Here we describe our studies of homologous DNA recombination in vertebrate somatic cells using reverse genetics in DT40 cells.
Enzymatic Regional Methylation Assay: A Novel Method to Quantify Regional CpG Methylation Density
Authors: Galm Oliver , Rountree Michael R. , Bachman Kurtis E. , Jair Kam-Wing , Baylin Stephen B. , Herman James G.
Published: July 2002

We have developed a novel quantitative method for rapidly assessing the CpG methylation density of a DNA region in mammalian cells. After bisulfite modification of genomic DNA, the region of interest is PCR amplified with primers containing two dam sites (GATC). The purified PCR products are then incubated with 14C-labeled S-adenosyl-L-methionine (SAM) and dam methyltransferase as an internal control to standardize DNA quantity. This is followed by an incubation with 3H-labeled SAM and SssI methyltransferase for methylation quantification. By use of standard mixtures of cell line DNA with a defined methylation status in every assay, the ratio (3H/14C signal) of each sample can be converted into percentage values to assess the methylation density of the amplified sequence. This methylation-sensitive technique, termed ERMA (Enzymatic Regional Methylation Assay) provides several advantages over existing methods used for methylation analysis as it determines an exact measurement of the methylation density of the region studied. We demonstrate a use of this technique in determining the methylation density of the promoter region of the tumor suppressor genep15INK4B and changes that occur after treatment with demethylating agents.
Linear motion correction in three dimensions applied to dynamic gadolinium enhanced breast imaging
Authors: Krishnan Sumati , Chenevert Thomas L. , Helvie Mark A. , Londy Frank L.
Published: July 2002

Quantitative analysis of dynamic gadolinium‐DTPA (diethylenetriamine pentaacetic acid) enhanced magnetic resonance imaging (MRI) is emerging as a highly sensitive tool for detecting malignant breast tissue. Three‐dimensional rapid imaging techniques, such as keyhole MRI, yield high temporal sampling rates to accurately track contrast enhancement and washout in lesions over the course of multiple volume acquisitions. Patient motion during the dynamic acquisitions is a limiting factor that degrades the image quality, particularly of subsequent subtraction images used to identify and quantitatively evaluate regions suggestive of malignancy. Keyhole imaging is particularly sensitive to motion since datasets acquired over an extended period are combined in k‐space. In this study, motion is modeled as set of translations in each of the three orthogonal dimensions. The specific objective of the study is to develop and implement an algorithm to correct the consequent phase shifts in k‐space data prior to offline keyhole reconstruction three‐dimensional (3D) volume breast MR acquisitions.
Determination of the urethral dose in prostate brachytherapy when the urethra cannot be visualized in the postimplant CT scan
Authors: Waterman F. M. , Dicker A. P.
Published: July 2002

This study investigates the feasibility of locating the urethra at the geometric center of peripherally loaded prostate implant when a urinary catheter is not utilized for the postimplant CT scan. Twenty postimplant CT scans utilizing a urinary catheter were randomly selected. The urethra was localized in each study and, in addition, a surrogate urethra was localized at the geometric center of the prostate. Dose‐volume histograms of the urethra and surrogate urethra were compiled and compared. The values obtained for the urethra and were in good agreement and demonstrate that the urethral dose can be determined reliably by locating a surrogate urethra at the geometric center of the prostate in a peripherally loaded implant when the urethra cannot be visualized.
The shape of the Eiffel Tower
Authors: Gallant Joseph
Published: July 2002

The distinctive shape of the Eiffel Tower is based on simple physics and is designed so that the maximum torque created by the wind is balanced by the torque due to the Tower’s weight. We use this idea to generate an equation for the shape of the Tower. The solution depends only on the width of the base and the maximum wind pressure. We parametrize the wind pressure and reproduce the shape of the Tower. We also discuss some of the Tower’s interesting history and characteristics.
From Maupertuis to Schrödinger. Quantization of classical variational principles
Authors: Gray C. G. , Karl G. , Novikov V. A.
Published: July 2002

In his first paper on wave mechanics, Schrödinger presented a heuristic argument which led from the Hamilton–Jacobi equation through the quantum variational principle to his famous wave equation. In his second paper, Schrödinger withdrew this heuristic argument as “incomprehensible.” We show by using a recently generalized form of Maupertuis’ principle that Schrödinger’s original heuristic argument can be made more logical. Aside from pedagogical interest, this path is useful as a method of quantization of general mechanical systems.
Periodic orbit theory analysis of the circular disk or annular billiard: Nonclassical effects and the distribution of energy eigenvalues
Authors: Robinett R. W.
Published: July 2002

Periodic orbit (PO) theory can be used to make connections between the quantum energy eigenvalue spectrum and the closed orbits of the corresponding classical system. The two-dimensional annular billiard or circular disk system (namely, a particle in the plane confined between inner and outer infinite circular walls at fR≡Rin<Rout≡R where 0<f<1) is examined in the context of periodic orbit theory to illustrate several novel aspects of the PO analysis. Important features of this problem include (i) the appearance and disappearance of various features in the classical path length spectrum as a parameter (in this case f=Rin/Rout), is continuously varied, (ii) the presence of path length features which do not correspond to classical trajectories, but are rather due to purely wavelike phenomena (namely, diffraction around the inner annulus), and (iii) the study of the contribution of different regions in the energy eigenvalue space to different classes of classical trajectories. This last feature is a general observation about periodic orbit theory which can be usefully applied to a number of billiard systems. In addition, since the annular disk system is seldom studied in detail in either classical or quantum mechanics, we provide an appendix which reviews some of the more traditional connections between the classical and quantum probability densities for this geometry, exemplifying some of the important differences between one- and two-dimensional systems which arise from purely geometrical rather than kinematical effects.
Semantics in teaching introductory physics
Authors: Williams H. Thomas
Published: July 2002

One reason that many able students find introductory college physics difficult is the way language is used in this course. The large vocabulary we use for precise purposes in physics contains many words which have related, but potentially confusing meanings in everyday usage. A surprising number of other words we use frequently are not used consistently in the language of introductory textbooks. Even our statements of important and well-established principles like Newton’s laws of motion are inconsistent. I analyze the treatment of Newton’s laws of motion in several well-known introductory textbooks for evidence of such problems. A series of suggestions for ways of reducing these difficulties concludes the paper.
Suspension of a disk on a surface of water
Authors: Penner A. Raymond
Published: July 2002

The suspension of a disk on the surface of water by the forces of surface tension and buoyancy is considered. Both theoretical and experimental results are presented along with a discussion of the relative importance of the buoyant force.
Turboprop and rotary-wing aircraft flight parameter estimation using both narrow-band and broadband passive acoustic signal-processing methods
Authors: Ferguson Brian G. , Lo Kam W.
Published: July 2002

Flight parameter estimation methods for an airborne acoustic source can be divided into two categories, depending on whether the narrow-band lines or the broadband component of the received signal spectrum is processed to estimate the flight parameters. This paper provides a common framework for the formulation and test of two flight parameter estimation methods: one narrow band, the other broadband. The performances of the two methods are evaluated by applying them to the same acoustic data set, which is recorded by a planar array of passive acoustic sensors during multiple transits of a turboprop fixed-wing aircraft and two types of rotary-wing aircraft. The narrow-band method, which is based on a kinematic model that assumes the source travels in a straight line at constant speed and altitude, requires time-frequency analysis of the acoustic signal received by a single sensor during each aircraft transit. The broadband method is based on the same kinematic model, but requires observing the temporal variation of the differential time of arrival of the acoustic signal at each pair of sensors that comprises the planar array. Generalized cross correlation of each pair of sensor outputs using a cross-spectral phase transform prefilter provides instantaneous estimates of the differential times of arrival of the signal as the acoustic wavefront traverses the array.
Neural system identification model of human sound localization
Authors: Jin Craig , Schenkel Markus , Carlile Simon
Published: July 2002

This paper examines the role of biological constraints in the human auditory localization process. A psychophysical and neural system modeling approach was undertaken in which performance comparisons between competing models and a human subject explore the relevant biologically plausible “realism constraints.” The directional acoustical cues, upon which sound localization is based, were derived from the human subject’s head-related transfer functions (HRTFs). Sound stimuli were generated by convolving bandpass noise with the HRTFs and were presented to both the subject and the model. The input stimuli to the model were processed using the Auditory Image Model of cochlear processing. The cochlear data were then analyzed by a time-delay neural network which integrated temporal and spectral information to determine the spatial location of the sound source. The combined cochlear model and neural network provided a system model of the sound localization process. Aspects of humanlike localization performance were qualitatively achieved for broadband and bandpass stimuli when the model architecture incorporated frequency division (i.e., the progressive integration of information across the different frequency channels) and was trained using variable bandwidth and center-frequency sounds. Results indicate that both issues are relevant to human sound localization performance.