bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of a Novel Major Histocompatibility Complex Class II–restricted Tumor Antigen Resulting from a Chromosomal Rearrangement Recognized by CD4+ T Cells
Authors: Wang Rong-Fu , Wang Xiang , Rosenberg Steven A.
Published: July 2002

CD4+ T cells play an important role in antitumor immune responses and autoimmune and infectious diseases. Although many major histocompatibility complex (MHC) class I–restricted tumor antigens have been identified in the last few years, little is known about MHC class II– restricted human tumor antigens recognized by CD4+ T cells. Here, we describe the identification of a novel melanoma antigen recognized by an human histocompatibility leukocyte antigen (HLA)-DR1–restricted CD4+ tumor-infiltrating lymphocyte (TIL)1363 using a genetic cloning approach. DNA sequencing analysis indicated that this was a fusion gene generated by a low density lipid receptor (LDLR) gene in the 5′ end fused to a GDP-l-fucose:β-d-galactoside 2-α-l-fucosyltransferase (FUT) in an antisense orientation in the 3′ end. The fusion gene encoded the first five ligand binding repeats of LDLR in the NH2 terminus followed by a new polypeptide translated in frame with LDLR from the FUT gene in an antisense direction. Southern blot analysis showed that chromosomal DNA rearrangements occurred in the 1363mel cell line. Northern blot analysis detected two fusion RNA transcripts present only in the autologous 1363mel, but not in other cell lines or normal tissues tested. Two minimal peptides were identified from the COOH terminus of the fusion protein. This represents the first demonstration that a fusion protein resulting from a chromosomal rearrangement in tumor cells serves as an immune target recognized by CD4+ T cells.
Lymphotoxin α/β and Tumor Necrosis Factor Are Required for Stromal Cell Expression of Homing Chemokines in B and T Cell Areas of the Spleen
Authors: Ngo Vu N. , Korner Heinrich , Gunn Michael D. , Schmidt Kerstin N. , Sean Riminton D. , Cooper Max D. , Browning Jeffrey L. , Sedgwick Jonathon D. , Cyster Jason G.
Published: July 2002

Mice deficient in the cytokines tumor necrosis factor (TNF) or lymphotoxin (LT) α/β lack polarized B cell follicles in the spleen. Deficiency in CXC chemokine receptor 5 (CXCR5), a receptor for B lymphocyte chemoattractant (BLC), also causes loss of splenic follicles. Here we report that BLC expression by follicular stromal cells is defective in TNF-, TNF receptor 1 (TNFR1)-, LTα- and LTβ-deficient mice. Treatment of adult mice with antagonists of LTα1β2 also leads to decreased BLC expression. These findings indicate that LTα1β2 and TNF have a role upstream of BLC/CXCR5 in the process of follicle formation. In addition to disrupted follicles, LT-deficient animals have disorganized T zones. Expression of the T cell attractant, secondary lymphoid tissue chemokine (SLC), by T zone stromal cells is found to be markedly depressed in LTα-, and LTβ-deficient mice. Expression of the SLC-related chemokine, Epstein Barr virus–induced molecule 1 ligand chemokine (ELC), is also reduced. Exploring the basis for the reduced SLC expression led to identification of further disruptions in T zone stromal cells. Together these findings indicate that LTα1β2 and TNF are required for the development and function of B and T zone stromal cells that make chemokines necessary for lymphocyte compartmentalization in the spleen.
Cleavage by Granzyme B Is Strongly Predictive of Autoantigen Status
Authors: Casciola-Rosen Livia , Andrade Felipe , Ulanet Danielle , Wong Wes Bang , Rosen Antony
Published: July 2002

Systemic autoimmune diseases are a genetically complex, heterogeneous group of disorders in which the immune system targets a diverse but highly specific group of intracellular autoantigens. The molecules targeted are not unified by common structure, function, or distribution in control cells but become clustered and concentrated in surface blebs when cells undergo apoptosis. We show here that the majority of autoantigens targeted across the spectrum of human systemic autoimmune diseases are efficiently cleaved by granzyme B in vitro and during cytotoxic lymphocyte granule–induced death, generating unique fragments not observed during any other form of apoptosis. These molecules are not cleaved by caspase-8, although this protease has a very similar specificity to granzyme B. The granzyme B cleavage sites in autoantigens contain amino acids in the P2 and P3 positions that are preferred by granzyme B but are not tolerated by caspase-8. In contrast to autoantigens, nonautoantigens are either not cleaved by granzyme B or are cleaved to generate fragments identical to those formed in other forms of apoptosis. The striking ability of granzyme B to generate unique fragments is therefore an exclusive property of autoantigens and unifies the majority of molecules targeted in this spectrum of diseases. These results focus attention on the role of the cytotoxic lymphocyte granule–induced death pathway in the initiation and propagation of systemic autoimmunity.
Irradiation-Mediated Rescue of T Cell–Specific V(D)j Recombination and Thymocyte Differentiation in Severe Combined Immunodeficient Mice by Bone Marrow Cells
Authors: Wang Chiyu , Bogue Molly A. , Levitt Jonathan M. , Roth David B.
Published: July 2002

In SCID (severe combined immunodeficient) mice, proper assembly of immunoglobulin and T cell receptor (TCR) genes is blocked by defective V(D)J recombination so that B and T lymphocyte differentiation is arrested at an early precursor stage. Treating the mice with gamma irradiation rescues V(D)J rearrangement at multiple TCR loci, promotes limited thymocyte differentiation, and induces thymic lymphomas. These effects are not observed in the B cell lineage. Current models postulate that irradiation affects intrathymic T cell precursors. Surprisingly, we found that transfer of irradiated SCID bone marrow cells to unirradiated host animals rescues both TCR rearrangements and thymocyte differentiation. These data indicate that irradiation affects precursor cells at an earlier stage of differentiation than was previously thought and suggest new models for the mechanism of irradiation rescue.
Helicobacter pylori Caga Protein Can Be Tyrosine Phosphorylated in Gastric Epithelial Cells
Authors: Asahi Momoyo , Azuma Takeshi , Ito Shigeji , Ito Yoshiyuki , Suto Hiroyuki , Nagai Yukifumi , Tsubokawa Misao , Tohyama Yumi , Maeda Shin , Omata Masao , Suzuki Toshihiko , Sasakawa Chihiro
Published: July 2002

Attachment of Helicobacter pylori to gastric epithelial cells induces various cellular responses, including the tyrosine phosphorylation of an unknown 145-kD protein and interleukin 8 production. Here we show that this 145-kD protein is the cagA product of H. pylori, an immunodominant, cytotoxin-associated antigen. Epithelial cells infected with various H. pylori clinical isolates resulted in generation of tyrosine-phosphorylated proteins ranging from 130 to 145 kD in size that were also induced in vitro by mixing host cell lysate with bacterial lysate. When epithelial cells were infected with [35S]methionine-labeled H. pylori, a radioactive 145-kD protein was detected in the immunoprecipitates with antiphosphotyrosine antibody or anti-CagA (cytotoxin-associated gene A) antibody. Consistently, the 145-kD protein recognized by the anti-CagA and antiphosphotyrosine antibodies was induced in epithelial cells after infection of wild-type H. pylori but not the cagA::Km mutant. Furthermore, the amino acid sequence of the phosphorylated 145-kD protein induced by H. pylori infection was identical to the H. pylori CagA sequence. These results reveal that the tyrosine-phosphorylated 145-kD protein is H. pylori CagA protein, which may be delivered from attached bacteria into the host cytoplasm. The identification of the tyrosine-phosphorylated protein will thus provide further insights into understanding the precise roles of CagA protein in H. pylori pathogenesis.
Extracellular K+ and Opening of Voltage-Gated Potassium Channels Activate T Cell Integrin Function
Authors: Levite Mia , Cahalon Liora , Peretz Asher , Hershkoviz Rami , Sobko Alex , Ariel Amiram , Desai Rooma , Attali Bernard , Lider Ofer
Published: July 2002

Elevated extracellular K+ ([K+]o), in the absence of “classical” immunological stimulatory signals, was found to itself be a sufficient stimulus to activate T cell β1 integrin moieties, and to induce integrin-mediated adhesion and migration. Gating of T cell voltage-gated K+ channels (Kv1.3) appears to be the crucial “decision-making” step, through which various physiological factors, including elevated [K+]o levels, affect the T cell β1 integrin function: opening of the channel leads to function, whereas its blockage prevents it. In support of this notion, we found that the proadhesive effects of the chemokine macrophage-inflammatory protein 1β, the neuropeptide calcitonin gene–related peptide (CGRP), as well as elevated [K+]o levels, are blocked by specific Kv1.3 channel blockers, and that the unique physiological ability of substance P to inhibit T cell adhesion correlates with Kv1.3 inhibition. Interestingly, the Kv1.3 channels and the β1 integrins coimmunoprecipitate, suggesting that their physical association underlies their functional cooperation on the T cell surface. This study shows that T cells can be activated and driven to integrin function by a pathway that does not involve any of its specific receptors (i.e., by elevated [K+]o). In addition, our results suggest that undesired T cell integrin function in a series of pathological conditions can be arrested by molecules that block the Kv1.3 channels.
A Macrophage Invasion Mechanism for Mycobacteria Implicating the Extracellular Domain of Cd43
Authors: Fratazzi Candida , Manjunath N. , Arbeit Robert D. , Carini Claudio , Gerken Thomas A. , Ardman Blair , Remold-O'Donnell Eileen , Remold Heinz G.
Published: July 2002

We studied the role of CD43 (leukosialin/sialophorin), the negatively charged sialoglycoprotein of leukocytes, in the binding of mycobacteria to host cells. CD43-transfected HeLa cells bound Mycobacterium avium, but not Salmonella typhimurium or Shigella flexneri. Quantitative bacteriology showed that macrophages (Mφ) from wild-type mice (CD43+/+) bound M. avium, Mycobacterium bovis (bacillus Calmette-Guérin), and Mycobacterium tuberculosis (strain H37Rv), whereas Mφ from CD43 knockout mice (CD43−/−) did not. Fluorescence microscopy demonstrated that the associated M. avium had been ingested by the CD43+/+ Mφ. The inability of CD43−/− Mφ to bind M. avium could be restored by addition of galactoglycoprotein (Galgp), the extracellular mucin portion of CD43. The effect of Galgp is not due to opsonization of the bacteria, but required its interaction with the Mφ; other mucins had no effect. CD43 expression by the Mφ was also required for optimal induction by M. avium of tumor necrosis factor (TNF)-α production, which likewise could be reconstituted by Galgp. In contrast, interleukin (IL)-10 production by M. avium–infected Mφ was CD43 independent, demonstrating discordant regulation of TNF-α and IL-10. These findings describe a novel role of CD43 in promoting stable interaction of mycobacteria with receptors on the Mφ enabling the cells to respond specifically with TNF-α production.
In Vivo Overexpression of IL-13 Receptor α2 Chain Inhibits Tumorigenicity of Human Breast and Pancreatic Tumors in Immunodeficient Mice
Authors: Kawakami Koji , Kawakami Mariko , Snoy Philip J. , Husain Syed R. , Puri Raj K.
Published: July 2002

Interleukin 13 receptor α2 (IL-13Rα2) chain is highly expressed on some tumor cell lines and primary cell cultures. This receptor chain plays an important role in ligand binding and internalization. To determine the functional significance of overexpression of this chain, we stably transfected IL-13Rα2 chain in human breast (MDA-MB-231) and pancreatic (PANC-1) cancer cell lines that naturally do not express this chain. There was no difference in growth between vector only transfected and IL-13Rα2 chain transfected cells in vitro. However, surprisingly, in immunodeficient mice, tumorigenicity was profoundly inhibited in IL-13Rα2 chain overexpressing tumors. Because breast tumors that grew later showed loss of IL-13Rα2 gene expression, lack of tumorigenicity correlated positively with IL-13Rα2 chain expression. Inflammatory cells including neutrophils and macrophages were identified in IL-13Rα2 overexpressing regressing tumors and neutrophils were found to produce IL-13. IL-13 showed a modest antitumor activity to IL-13Rα2 chain overexpressing tumors in vitro and in vivo. Furthermore, IL-13Rα2 chain overexpressing tumors constitutively produced IL-8 that has been shown to have antitumor effect. These results establish a novel function of a cytokine receptor chain and further suggest that the presence of this chain on tumor cells by itself may play a key role in tumorigenicity.
Cysteinyl Leukotrienes and Uridine Diphosphate Induce Cytokine Generation by Human Mast Cells Through an Interleukin 4–regulated Pathway that Is Inhibited by Leukotriene Receptor Antagonists
Authors: Mellor Elizabeth A. , Austen K. Frank , Boyce Joshua A.
Published: July 2002

We previously reported that interleukin (IL)-4 upregulates the expression of leukotriene C4 synthase (LTC4S) by human cord blood–derived mast cells (hMCs), augments their high-affinity Fc receptor for IgE (FcϵRI)-dependent generation of eicosanoids and cytokines, and induces a calcium flux in response to cysteinyl leukotrienes (cys-LTs) and uridine diphosphate (UDP) that is blocked by cys-LT receptor antagonists. We speculated that this IL-4–dependent, receptor-mediated response to the cys-LTs and UDP might induce cytokine generation by hMCs without concomitant exocytosis. Unlike hMCs maintained in cytoprotective stem cell factor (SCF) alone, hMCs primed for 5 d with IL-4 responded to UDP (1 μM), LTC4 (100 nM), and LTD4 (100 nM) by producing IL-5, tumor necrosis factor (TNF)-α, and especially large quantities of macrophage inflammatory protein (MIP)-1β de novo at 6 h, preceded by the induced expression of the corresponding mRNAs. Cys-LT– and UDP-mediated cytokine production by the primed hMCs occurred without histamine release or PGD2 generation and was inhibited by the CysLT1 receptor antagonist MK571. Additionally, pretreatment of hMCs with MK571 or with the cys-LT biosynthetic inhibitor MK886 decreased IL-5 and TNF-α production in response to IgE receptor cross-linkage, implying a positive feedback by endogenously produced cys-LTs. Cys-LTs and UDP thus orchestrate a novel, IL-4–regulated, non-IgE–dependent hMC activation for cytokine gene induction that could be initiated by microbes, cellular injury, or neurogenic or inflammatory signals; and this pathobiologic event would not be recognized in tissue studies where hMC activation is classically defined by exocytosis.
Mechanisms of Barbiturate Inhibition of Acetylcholine Receptor Channels
Authors: Dilger James P. , Boguslavsky Rebecca , Barann Martin , Katz Tamir , Vidal Ana Maria
Published: July 2002

We used patch clamp techniques to study the inhibitory effects of pentobarbital and barbital on nicotinic acetylcholine receptor channels from BC3H-1 cells. Single channel recording from outside-out patches reveals that both drugs cause acetylcholine-activated channel events to occur in bursts. The mean duration of gaps within bursts is 2 ms for 0.1 mM pentobarbital and 0.05 ms for 1 mM barbital. In addition, 1 mM barbital reduces the apparent single channel current by 15%. Both barbiturates decrease the duration of openings within a burst but have only a small effect on the burst duration. Macroscopic currents were activated by rapid perfusion of 300 μM acetylcholine to outside-out patches. The concentration dependence of peak current inhibition was fit with a Hill function; for pentobarbital, Ki = 32 μM, n = 1.09; for barbital, Ki = 1900 μM, n = 1.24. Inhibition is voltage independent. The kinetics of inhibition by pentobarbital are at least 30 times faster than inhibition by barbital (3 ms vs. <0.1 ms at the Ki). Pentobarbital binds ≥10-fold more tightly to open channels than to closed channels; we could not determine whether the binding of barbital is state dependent. Experiments performed with both barbiturates reveal that they do not compete for a single binding site on the acetylcholine receptor channel protein, but the binding of one barbiturate destabilizes the binding of the other. These results support a kinetic model in which barbiturates bind to both open and closed states of the AChR and block the flow of ions through the channel. An additional, lower-affinity binding site for pentobarbital may explain the effects seen at >100 μM pentobarbital.
Cation-selective Mutations in the M2 Domain of the Inhibitory Glycine Receptor Channel Reveal Determinants of Ion-Charge Selectivity
Authors: Keramidas Angelo , Moorhouse Andrew J. , Pierce Kerrie D. , Schofield Peter R. , Barry Peter H.
Published: July 2002

Ligand-gated ion channel receptors mediate neuronal inhibition or excitation depending on their ion charge selectivity. An investigation into the determinants of ion charge selectivity of the anion-selective α1 homomeric glycine receptor (α1 glycine receptor [GlyR]) was undertaken using point mutations to residues lining the extra- and intracellular ends of the ion channel. Five mutant GlyRs were studied. A single substitution at the intracellular mouth of the channel (A-1′E GlyR) was sufficient to convert the channels to select cations over anions with PCl/PNa = 0.34. This result delimits the selectivity filter and provides evidence that electrostatic interactions between permeating ions and pore residues are a critical factor in ion charge selectivity. The P-2′Δ mutant GlyR retained its anion selectivity (PCl/PNa = 3.81), but it was much reduced compared with the wild-type (WT) GlyR (PCl/PNa = 27.9). When the A-1′E and the P-2′Δ mutations were combined (selectivity double mutant [SDM] GlyR), the relative cation permeability was enhanced (PCl/PNa = 0.13). The SDM GlyR was also Ca2+ permeable (PCa/PNa = 0.29). Neutralizing the extracellular mouth of the SDM GlyR ion channel (SDM+R19′A GlyR) produced a more Ca2+-permeable channel (PCa/PNa = 0.73), without drastically altering monovalent charge selectivity (PCl/PNa = 0.23). The SDM+R19′E GlyR, which introduces a negatively charged ring at the extracellular mouth of the channel, further enhanced Ca2+ permeability (PCa/PNa = 0.92), with little effect on monovalent selectivity (PCl/PNa = 0.19). Estimates of the minimum pore diameter of the A-1′E, SDM, SDM+R19′A, and SDM+R19′E GlyRs revealed that these pores are larger than the α1 GlyR, with the SDM-based GlyRs being comparable in diameter to the cation-selective nicotinic acetylcholine receptors. This result provides evidence that the diameter of the ion channel is also an important factor in ion charge selectivity.
Growth of graded noetherian rings
Authors: Stephenson Darin , Zhang James
Published: July 2002

We show that every graded locally finite right noetherian algebra has sub-exponential growth. As a consequence, every noetherian algebra with exponential growth has no finite dimensional filtration which leads to a right (or left) noetherian associated graded algebra. We also prove that every connected graded right noetherian algebra with finite global dimension has finite GK-dimension. Using this, we can classify all connected graded noetherian algebras of global dimension two.
Responses of Neurons in Macaque Area V4 During Memory-guided Visual Search
Authors: Chelazzi Leonardo , Miller Earl K. , Duncan John , Desimone Robert
Published: July 2002

In a typical scene with many different objects, attentional mechanisms are needed to select relevant objects for visual processing and control over behavior. To test the role of area V4 in the selection of objects based on non-spatial features, we recorded from V4 neurons in the monkey, using a visual search paradigm. A cue stimulus was presented at the center of gaze, followed by a blank delay period. After the delay, a two-stimulus array was presented extrafoveally, and the monkey was rewarded for detecting the target stimulus matching the cue. The array was composed of one ‘good’ stimulus (effective in driving the cell when presented alone) and one ‘poor’ stimulus (ineffective in driving the cell when presented alone). When the choice array was presented in the receptive field (RF) of the neuron, many cells showed suppressive interactions between the stimuli as well as strong attention effects. Within 150–200 ms of array onset, responses to the array were determined by the target stimulus. If the target was the good stimulus, the response to the array became equal to the response to the good stimulus presented alone. If the target was the poor stimulus, the response approached the response to that stimulus presented alone. Thus the influence of the nontarget stimulus was filtered out. These effects were reduced or eliminated when the poor stimulus was located outside the RF and, therefore, no longer competing for the cell's response. Overall, the results support a ‘biased competition’ model of attention, according to which objects in the visual field compete for representation in the cortex, and this competition is biased in favor of the behaviorally relevant object.
Genome‐wide insertional mutagenesis in human cells by the Drosophila mobile element Minos
Authors: Klinakis Apostolos G , Zagoraiou Laskaro , Vassilatis Demetrios K , Savakis Charalambos
Published: July 2002

The development of efficient non‐viral methodologies for genome‐wide insertional mutagenesis and gene tagging in mammalian cells is highly desirable for functional genomic analysis. Here we describe transposon mediated mutagenesis (TRAMM), using naked DNA vectors based on the Drosophila hydei transposable element Minos. By simple transfections of plasmid Minos vectors in HeLa cells, we have achieved high frequency generation of cell lines, each containing one or more stable chromosomal integrations. The Minos‐derived vectors insert in different locations in the mammalian genome. Genome‐wide mutagenesis in HeLa cells was demonstrated by using a Minos transposon containing a lacZ–neo gene‐trap fusion to generate a HeLa cell library of at least 105 transposon insertions in active genes. Multiple gene traps for six out of 12 active genes were detected in this library. Possible applications of Minos‐based TRAMM in functional genomics are discussed.
The archaeal TFIIEα homologue facilitates transcription initiation by enhancing TATA‐box recognition
Authors: Bell Stephen D , Brinkman Arie B , van der Oost John , Jackson Stephen P
Published: July 2002

Transcription from many archaeal promoters can be reconstituted in vitro using recombinant TATA‐box binding protein (TBP) and transcription factor B (TFB)—homologues of eukaryal TBP and TFIIB—together with purified RNA polymerase (RNAP). However, all archaeal genomes sequenced to date reveal the presence of TFE, a homologue of the α‐subunit of the eukaryal general transcription factor, TFIIE. We show that, while TFE is not absolutely required for transcription in the reconstituted in vitro system, it nonetheless plays a stimulatory role on some promoters and under certain conditions. Mutagenesis of the TATA box or reduction of TBP concentration in transcription reactions sensitizes a promoter to TFE addition. Conversely, saturating reactions with TBP de‐sensitizes promoters to TFE. These results suggest that TFE facilitates or stabilizes interactions between TBP and the TATA box.
Prominent mitochondrial DNA recombination intermediates in human heart muscle
Authors: Kajander Olli A , Karhunen Pekka J , Holt Ian J , Jacobs Howard T
Published: July 2002

Recombination intermediates containing four‐way (Holliday) junctions are generated during DNA repair and replication in many systems, including yeast mitochondrial DNA (mtDNA). In contrast, convincing evidence for recombination in mammalian mtDNA is lacking. We have used two‐dimensional agarose‐gel electrophoresis to analyse non‐linear forms of mtDNA in human heart muscle. Replication intermediates from both the coupled and strand‐asynchronous mtDNA replication pathways were detected. An additional class of non‐linear molecules, with the electrophoretic properties of four‐way junctions, was also prominent. These molecules were insensitive to topoisomerase I or RNase H, but were diminished by branch migration or RuvC treatment. Junctional molecules were detected in all regions of the mitochondrial genome, were found in myocardial DNA from young and old adults, but were present at lower levels in skeletal muscle and placenta. We suggest that they could represent intermediates of mtDNA repair, given their prevalence in the oxyradical‐rich environment of heart muscle mitochondria.
Differential expression of thymosin β‐10 by early passage and senescent vascular endothelium is modulated by VPF/VEGF: evidence for senescent endothelial cells in vivo at sites of atherosclerosis
Authors: VASILE ELIZA , TOMITA YASUHIKO , BROWN LAWRENCE F. , KOCHER OLIVIER , DVORAK HAROLD F.
Published: July 2002

VPF/VEGF acts selectively on the vascular endothelium to enhance permeability, induce cell migration and division, and delay replicative senescence. To understand the changes in gene expression during endothelial senescence, we investigated genes that were differentially expressed in early vs. late passage (senescent) human dermal endothelial cells (HDMEC) using cDNA array hybridization. Early passage HDMEC cultured with or without VPF/VEGF overexpressed 9 and underexpressed 6 genes in comparison with their senescent counterparts. Thymosin β‐10 expression was modulated by VPF/VEGF and was strikingly down‐regulated in senescent EC. The β‐thymosins are actin G‐sequestering peptides that regulate actin dynamics and are overexpressed in neoplastic transformation. We have also identified senescent EC in the human aorta at sites overlying atherosclerotic plaques. These EC expressed senescence‐associated neutral β‐galactosidase and, in contrast to adventitial microvessel endothelium, exhibited weak staining for thymosin β‐10. ISH performed on human malignant tumors revealed strong thymosin β‐10 expression in tumor blood vessels. This is the first report that Tβ‐10 expression is significantly reduced in senescent EC, that VPF/VEGF modulates thymosin β‐10 expression, and that EC can become senescent in vivo. The reduced expression of thymosin β‐10 may contribute to the senescent phenotype by reducing EC plasticity and thus impairing their response to migratory stimuli.—Vasile, E., Tomita, Y., Brown, L. F., Kocher, O., Dvorak, H. F. Differential expression of thymosin Β‐10 by early passage and senescent vascular endothelium is modulated by VPF/VEGF: evidence for senescent endothelial cells in vivo at sites of atherosclerosis. FASEB J. 15, 458‐466 (2001)
Phylogenomic analysis of 16S rRNA:(guanine‐N2) methyltransferases suggests new family members and reveals highly conserved motifs and a domain structure similar to other nucleic acid amino‐methyltransferases
Authors: BUJNICKI JANUSZ M.
Published: July 2002

ABSTRACT The sequences of known Escherichia coli 16S rRNA:m2G1207 methyltransferase (MTase) RsmC and hypothetical 16S rRNA:m2G966 MTase encoded by the ygjo open reading frame were used to carry out a database search of other putative m2G‐generating enzymes in finished and unfinished genomic sequences. Sequence comparison and phy‐logenetic analysis of 21 close homologs of RsmC and YgjO revealed the presence of the third paralogous lineage in E. coli and other γ‐Proteobacteria, which might correspond to the subfamily of MTases spe¬cific for G1516 in 16S rRNA. In addition, the com¬parative sequence analysis supported by sequence/ structure threading suggests that rRNA:m2G MTases are very closely related to RNA and DNA:m6A MTases and that these two enzyme families share common architecture of the active site and presum‐ably a similar mechanism of methyl group transfer onto the exocyclic amino group of their target bases.—Bujnicki, J. M. Phylogenomic analysis of 16S rRNA:(guanine‐N2) methyltransferases suggests new family members and reveals highly conserved motifs and a domain structure similar to other nucleic acid amino‐methyltransferases. FASEB J. 14, 2365‐2368 (2000)
Coordinated expression in chronically unconscious persons
Authors:   The American Association for Research into Nervous and Mental Diseases, Plum F. , Schiff N. , Ribary U. , Llinás R.
Published: July 2002

The clinically described ‘persistent vegetative state’ (PVS), consists of wakefulness unaccompanied by any evidence of the subject's awareness of self or environment. Past studies from our own and other laboratories have used positron emission tomography (PET) to study brain metabolism in approximately 20 such patients during wakeful periods. All those efforts identified global cerebral glucose metabolism at or below levels encountered during deep barbiturate anaesthesia. Nevertheless, the clinical literature includes rare reports of relatively isolated cognitive functions expressed by PVS patients late in their course. The observation raises the question of whether such activity reflects awareness or unconscious automatic behaviour. We employed magnetometry (MEG), PET scanning, MR imaging and 24–hour EEG recordings to evaluate three patients clinically vegetative between six months and 20 years after onset.

Neither meticulous clinical examinations nor 24–hour EEG and video monitoring provided any hint of cognitive interaction in any subject. Nevertheless, patient 1 uttered single words once every 48 hours or more; patient 2 frequently expressed coordinated, non–purposeful, non–dystonic movements in arms and/or legs; and, patient 3 expressed strong emotional negativity without motor responses to noxious stimuli with occasional quieting in response to prosodic stimuli.

All patients had whole–brain averaged global metabolism levels below 50% of normal. Patient 1, however, demonstrated preserved islands of increased metabolism in the posterior frontal and posterior temporal lobes, as well as MEG activations of Heschl's gyrus all located in the left hemisphere. In patient 2, selected increased metabolism was confined to the frontal poles and related subcortical structures. MRI in patient 3 demonstrated severe, bilateral post–traumatic cerebral atrophy. PET metabolism was diffusely reduced to 40% of normal but MEG evoked potentials indicated early and late sensory processing with abnormal later evoked components.

The correlation of fragmentary behaviour with preserved metabolic and physiologic activity in cortical and subcortical regions known to support specific modular functions is novel. The finding demonstrates the capacity of severely damaged brains to partially express surviving modular functions without evidence of integrative processes that would be necessary to produce consciousness. These data represent a baseline against which less severely damaged brains may be assessed and ultimately tested the recovery of conscious expression.

We conclude that the mere expression of isolated neuropsychologic activity by isolated modules is insufficient to generate consciousness in overwhelmingly damaged brains.
Event-related functional magnetic resonance imaging: modelling, inference and optimization
Authors: Josephs Oliver , Henson Richard N. A.
Published: July 2002

Event–related functional magnetic resonance imaging is a recent and popular technique for detecting haemodynamic responses to brief stimuli or events. However, the design of event–related experiments requires careful consideration of numerous issues of measurement, modelling and inference. Here we review these issues, with particular emphasis on the use of basis functions within a general linear modelling framework to model and make inferences about the haemodynamic response. With these models in mind, we then consider how the properties of functional magnetic resonance imaging data determine the optimal experimental design for a specific hypothesis, in terms of stimulus ordering and interstimulus interval. Finally, we illustrate various event–related models with examples from recent studies.
The Saccharomyces cerevisiae ubiquitin–proteasome system
Authors: Hochstrasser M. , Johnson P. R. , Arendt C. S. , Amerik A. Y. , Swaminathan S. , Swanson R. , Li S. , Laney J. , Pals-Rylaarsdam R. , Nowak J. , Connerly P. L.
Published: July 2002

Our studies of the yeast ubiquitin-proteasome pathway have uncovered a number of general principles that govern substrate selectivity and proteolysis in this complex system. Much of the work has focused on the destruction of a yeast transcription factor, MATα2. The α2 protein is polyubiquitinated and rapidly degraded in α–haploid cells. One pathway of proteolytic targeting, which depends on two distinct endoplasmic reticulum–localized ubiquitin–conjugating enzymes, recognizes the hydrophobic face of an amphipathic helix in α2. Interestingly, degradation of α2 is blocked ina/α–diploid cells by heterodimer formation between the α2 anda1 homeodomain proteins. The data suggest that degradation signals may overlap protein–protein interaction surfaces, allowing a straightforward steric mechanism for regulated degradation. Analysis of α2 degradation led to the identification of both 20S and 26S proteasome subunits, and several key features of proteasome assembly and active–site formation were subsequently uncovered. Finally, it has become clear that protein (poly)ubiquitination is highly dynamicin vivo, and our studies of yeast de–ubiquitinating enzymes illustrate how such enzymes can facilitate the proteolysis of diverse substrates.
The pathophysiology of multiple sclerosis⋮ the mechanisms underlying the production of symptoms and the natural history of the disease
Authors: Smith Kenneth J. , McDonald W. I.
Published: July 2002

The pathophysiology of multiple sclerosis is reviewed, with emphasis on the axonal conduction properties underlying the production of symptoms, and the course of the disease. The major cause of the negative symptoms during relapses (e.g. paralysis, blindness and numbness) is conduction block, caused largely by demyelination and inflammation, and possibly by defects in synaptic transmission and putative circulating blocking factors. Recovery from symptoms during remissions is due mainly to the restoration of axonal function, either by remyelination, the resolution of inflammation, or the restoration of conduction to axons which persist in the demyelinated state. Conduction in the latter axons shows a number of deficits, particularly with regard to the conduction of trains of impulses and these contribute to weakness and sensory problems. The mechanisms underlying the sensitivity of symptoms to changes in body temperature (Uhthoff's phenomenon) are discussed. The origin of ‘positive’ symptoms, such as tingling sensations, are described, including the generation of ectopic trains and bursts of impulses, ephaptic interactions between axons and/or neurons, the triggering of additional, spurious impulses by the transmission of normal impulses, the mechanosensitivity of axons underlying movement–induced sensations (e.g. Lhermitte's phenomenon) and pain. The clinical course of the disease is discussed, together with its relationship to the evolution of lesions as revealed by magnetic resonance imaging and spectroscopy. The earliest detectable event in the development of most new lesions is a breakdown of the blood–brain barrier in association with inflammation. Inflammation resolves after approximately one month, at which time there is an improvement in the symptoms. Demyelination occurs during the inflammatory phase of the lesion. An important mechanism determining persistent neurological deficit is axonal degeneration, although persistent conduction block arising from the failure of repair mechanisms probably also contributes.
Anatomical connectivity defines the organization of clusters of cortical areas in the macaque and the cat
Authors: Hilgetag Claus–C. , Burns Gully A. P. C. , O'Neill Marc A. , Scannell Jack W. , Young Malcolm P.
Published: July 2002

The number of different cortical structures in mammalian brains and the number of extrinsic fibres linking these regions are both large. As with any complex system, systematic analysis is required to draw reliable conclusions about the organization of the complex neural networks comprising these numerous elements. One aspect of organization that has long been suspected is that cortical networks are organized into ‘streams’ or ‘systems’. Here we report computational analyses capable of showing whether clusters of strongly interconnected areas are aspects of the global organization of cortical systems in macaque and cat. We used two different approaches to analyse compilations of corticocortical connection data from the macaque and the cat. The first approach, optimal set analysis, employed an explicit definition of a neural ‘system’ or ‘stream’, which was based on differential connectivity. We defined a two–component cost function that described the cost of the global cluster arrangement of areas in terms of the areas‘ connectivity within and between candidate clusters. Optimal cluster arrangements of cortical areas were then selected computationally from the very many possible arrangements, using an evolutionary optimization algorithm. The second approach, non–parametric cluster analysis (NPCA), grouped cortical areas on the basis of their proximity in multidimensional scaling representations. We used non–metric multidimensional scaling to represent the cortical connectivity structures metrically in two and five dimensions. NPCA then analysed these representations to determine the nature of the clusters for a wide range of different cluster shape parameters. The results from both approaches largely agreed. They showed that macaque and cat cortices are organized into densely intra–connected clusters of areas, and identified the constituent members of the clusters. These clusters reflected functionally specialized sets of cortical areas, suggesting that structure and function are closely linked at this gross, systems level.
Communication and camouflage with the same ‘bright’ colours in reef fishes
Authors: Justin Marshall N.
Published: July 2002

Reef fishes present the observer with the most diverse and stunning assemblage of animal colours anywhere on earth. The functions of some of these colours and their combinations are examined using new non-subjective spectrophotometric measurements of the colours of fishes and their habitat. Conclusions reached are as follows: (i) the spectra of colours in high spatial frequency patterns are often well designed to be very conspicuous to a colour vision system at close range but well camouflaged at a distance; (ii) blue and yellow, the most frequently used colours in reef fishes, may be good for camouflage or communication depending on the background they are viewed against; and (iii) reef fishes use a combination of colour and behaviour to regulate their conspicuousness and crypsis.
Evolution and the molecular basis of somatic hypermutation of antigen receptor genes
Authors: Diaz Marilyn , Flajnik Martin F. , Klinman Norman
Published: July 2002

Somatic hypermutation of immunoglobulin genes occurs in many vertebrates including sharks, frogs, camels, humans and mice. Similarities among species reveal a common mechanism and these include the AGC/T sequence hot spot, preponderance of base substitutions, a bias towards transitions and strand bias. There are some differences among species, however, that may unveil layers of the mechanism. These include a G:C bias in frog and shark IgM but not in nurse shark antigen receptor (NAR), a high frequency of doublets in NAR hypermutation, and the co–occurrence of somatic hypermutation with gene conversion in some species. Here we argue that some of the similarities and differences among species are best explained by error–prone DNA synthesis by the translesion synthesis DNA polymerase zeta (Pol ζ) and, as suggested by others, induction of DNA synthesis by DNA breaks in antigen receptor variable genes. Finally, targeting of the variable genes is probably obtained via transcription–related elements, and it is the targeting phase of somatic hypermutation that is the most likely to reveal molecules unique to adaptive immunity.
Theories of episodic memory
Authors: Mayes Andrew R. , Roberts Neil
Published: July 2002

Theories of episodic memory need to specify the encoding (representing), storage, and retrieval processes that underlie this form of memory and indicate the brain regions that mediate these processes and how they do so. Representation and re–representation (retrieval) of the spatiotemporally linked series of scenes, which constitute an episode, are probably mediated primarily by those parts of the posterior neocortex that process perceptual and semantic information. However, some role of the frontal neocortex and medial temporal lobes in representing aspects of context and high–level visual object information at encoding and retrieval cannot currently be excluded. Nevertheless, it is widely believed that the frontal neocortex is mainly involved in coordinating episodic encoding and retrieval and that the medial temporal lobes store aspects of episodic information. Establishing where storage is located is very difficult and disagreement remains about the role of the posterior neocortex in episodic memory storage. One view is that this region stores all aspects of episodic memoryab initiofor as long as memory lasts. This is compatible with evidence that the amygdala, basal forebrain, and midbrain modulate neocortical storage. Another view is that the posterior neocortex only gradually develops the ability to store some aspects of episodic information as a function of rehearsal over time and that this information is initially stored by the medial temporal lobes. A third view is that the posterior neocortex never stores these aspects of episodic information because the medial temporal lobes store them for as long as memory lasts in an increasingly redundant fashion. The last two views both postulate that the medial temporal lobes initially store contextual markers that serve to cohere featural information stored in the neocortex. Lesion and functional neuroimaging evidence still does not clearly distinguish between these views. Whether the feeling that an episodic memory is familiar depends on retrieving an association between a retrieved episode and this feeling, or by an attribution triggered by a priming process, is unclear. Evidence about whether the hippocampus and medial temporal lobe cortices play different roles in episodic memory is conflicting. Identifying similarities and differences between episodic memory and both semantic memory and priming will require careful componential analysis of episodic memory.
Subcellular localization of the Snf1 kinase is regulated by specific β subunits and a novel glucose signaling mechanism
Authors: Vincent Olivier , Townley Robert , Kuchin Sergei , Carlson Marian
Published: July 2002

The Snf1/AMP-activated protein kinase family has broad roles in transcriptional, metabolic, and developmental regulation in response to stress. In Saccharomyces cerevisiae, Snf1 is required for the response to glucose limitation. Snf1 kinase complexes contain the α (catalytic) subunit Snf1, one of the three related β subunits Gal83, Sip1, or Sip2, and the γ subunit Snf4. We present evidence that the β subunits regulate the subcellular localization of the Snf1 kinase. Green fluorescent protein fusions to Gal83, Sip1, and Sip2 show different patterns of localization to the nucleus, vacuole, and/or cytoplasm. We show that Gal83 directs Snf1 to the nucleus in a glucose-regulated manner. We further identify a novel signaling pathway that controls this nuclear localization in response to glucose phosphorylation. This pathway is distinct from the glucose signaling pathway that inhibits Snf1 kinase activity and responds not only to glucose but also to galactose and sucrose. Such independent regulation of the localization and the activity of the Snf1 kinase, combined with the distinct localization of kinases containing different β subunits, affords versatility in regulating physiological responses.
Fine-Resolution Physical Mapping of Genomic Diversity in Candida albicans
Authors: Chibana Hiroji , Beckerman Janna L. , Magee P.T.
Published: July 2002

It has been suggested that Candida albicans, a diploid asexual fungus, achieves genetic diversity by genomic rearrangement. This important human pathogen may provide a system in which to analyze alternate routes to genomic diversity. C. albicans has a highly variable karyotype; its chromosomes contain a middle repeated DNA sequence called the Major Repeat Sequence (MRS), composed of subrepeats HOK, RPS, and RB2. RPS is tandemly repeated while the other subrepeats occur once in each MRS. Chromosome 7, the smallest of the eight chromosomes, has been previously mapped. The complete physical map of this chromosome was used to analyze chromosome 7 diversity in six strains, including two well-characterized laboratory strains (1006 and WO-1) and four clinical ones. We found four types of events to explain the genomic diversity: 1) Chromosome length polymorphism (CLP) results from expansion and contraction of the RPS; 2) reciprocal translocation occurs at the MRS loci; 3) chromosomal deletion; and (4) trisomy of individual chromosomes. These four phenomena play an important role in generating genomic diversity in C. albicans.
A mixed‐encoding genetic algorithm with beam constraint for conformal radiotherapy treatment planning
Authors: Wu Xingen , Zhu Yunping
Published: July 2002

In this paper we propose a new hierarchical evolutionary algorithm that combines binary encoding and floating‐point encoding to automatically select the beam directions and determine the weights of the selected beams. With traditional optimization methods the beam directions are fixed a priori by the operator in recognition of the fact that computer selection of beam directions is a difficult problem. In this investigation, we used a hybrid‐encoding scheme. The binary encoding part of each chromosome was used to select the beam directions, and its corresponding floating‐point encoding part of the same chromosome was used to determine the weights of those selected beams. Before beginning the optimization process, we set a constraint on the number of the beam directions we wanted in the final solution. We present three examples to verify this method. These examples differ with each other in tumor sites, problem sizes, and optimization parameters. Three‐dimensional optimization results and statistical data showed that this method is feasible. We think this method can be easily extended to solve more complex target problems (such as nonconvex target problems).
Investigation of human brain hemodynamics by simultaneous near‐infrared spectroscopy and functional magnetic resonance imaging
Authors: Toronov Vladislav , Webb Andrew , Choi Jee Hyun , Wolf Martin , Michalos Antoios , Gratton Enrico , Hueber Dennis
Published: July 2002

The aim of this study was to compare functional cerebral hemodynamic signals obtained simultaneously by near infrared spectroscopy (NIRS) and by functional magnetic resonance imaging (fMRI). The contribution of superficial layers (skin and skull) to the NIRS signal was also assessed. Both methods were used to generate functional maps of the motor cortex area during a periodic sequence of stimulation by finger motion and rest. In all subjects we found a good collocation of the brain activity centers revealed by both methods. We also found a high temporal correlation between the BOLD signal (fMRI) and the deoxy‐hemoglobin concentration (NIRS) in the subjects who exhibited low fluctuations in superficial head tissues.