bluuebunny/crossref_metadata_2025 huggingface dataset:
Heavy Metal-Induced Changes in the Glutathione Levels and Glutathione Reductase/Glutathione S-Transferase Activities in the Liver of Male Mice
Authors: Sheweita Salah A.
Published: July 2002

Glutathione S-transferases catalyze the metabolism of reactive substances of exogenous or endogenous origin and are involved in inactivation processes of xenobiotics and their metabolites. The present study aims at investigating the influence of heavy metals on the hepatic level of reduced glutathione (GSH), glutathione S-transferase, and glutathione reductase activities in the liver of male mice after single-dose (1 and 24 h) and repeated-dose treatments (three consecutive days). The hepatic level of GSH was depleted after single-dose treatments with cadmium chloride, mercuric chloride, cobalt chloride, cesium chloride, lead acetate, and silver nitrate, and percentage of GSH depletion was greater still after the repeated-dose treatments. Nickel chloride, on the other hand, did not cause any change in the level of GS H after any period of treatment. Glutathione reductase activity was increased 24 hours after treatment with cadmium chloride, mercuric chloride, lead acetate, and silver nitrate, whereas cobalt chloride decreased such activity after repeated doses. With the exception of cadmium chloride, glutathione S-transferase activity was significantly decreased 24 hours after a single-dose treatment with all of the tested heavy metals. Such alterations in the activities of phase II drug-metabolizing enzymes as a result of heavy metal treatment may change the hepatic capacity for the detoxification of many toxic compounds from endogenous or exogenous sources.
Effects of Melatonin on Oxidative Stress in Streptozotocin-Induced Diabetic Rats
Authors: Maritim Alice C. , Moore Brian H. , Sanders Ruth A. , Watkins John B.
Published: July 2002

Oxidative stress plays an important role in diabetes and other oxygen-related diseases. Melatonin, a pineal hormone thought to be a scavenger of oxygen radicals and a potentially advantageous therapeutic agent in diseases having oxidative stress, was administered (10 mg/kg ip, in gum tragacanth to prolong its absorption, once a day for 4 successive days) to normal and 30-day streptozotocin-induced diabetic Sprague-Dawley rats, after which markers of oxidative stress were assessed in the liver, kidney, intestine, and spleen. Alanine and aspartate aminotransferase activities in serum, which were increased after diabetes, were not increased further by melatonin administration, indicating that there was no melatonin-related liver toxicity. Most melatonin-induced effects were seen in the liver, and very few in extrahepatic tissues. In livers of diabetic rats, reduced concentration of nitrite and increased lipid peroxidation were both restored to normal levels following treatment with melatonin. Hepatic glutathione peroxidase activity was not changed in diabetics, but was decreased after melatonin administration in both normal and diabetic animals. Total glutathione concentrations were significantly decreased in livers of all diabetics and were not normalized by melatonin treatment. Hepatic superoxide dis-mutase activity was elevated following melatonin dosing in normal rats, but dropped below normal levels in diabetic rats and was not restored by melatonin treatment. Glutathione 5-transferase activity was higher than normal in melatonin-dosed normal rat livers. These results suggest that after 4 days of administration, melatonin may enable various enzymes of the hepatic antioxidative defense system to better detoxify harmful oxygen radicals without producing overt toxicity in a disease such as diabetes.
Matrix Metalloproteinases MMP2 and MMP9 Are Produced in Early Stages of Kidney Morphogenesis but Only MMP9 Is Required for Renal Organogenesis In Vitro
Authors: Lelongt Brigitte , Trugnan Germain , Murphy Gillian , Ronco Pierre M.
Published: July 2002

We analyzed matrix metalloproteinase (MMP) production by 11-d embryonic mouse kidneys and the effects of these enzymes on subsequent renal organogenesis. In vivo, immunolocalization of metalloproteinases by laser scanning confocal microscopy and zymograms of kidney lysates showed that the mesenchyme of embryonic kidneys synthesized both MMP9 and MMP2 enzymes. In vitro, embryonic kidneys also secreted both enzymes when cultured in a medium devoid of hormone, growth factor, and serum for 24 h during which T-shaped branching of the ureter bud appeared. We then evaluated the role of MMP2 and MMP9 in kidney morphogenesis by adding anti-MMP2 or anti-MMP9 IgGs to the culture medium of 11-d kidneys for 24 or 72 h. Although it inhibited activity of the mouse enzyme, anti-MMP2 IgGs had no effect on kidney morphogenesis. In contrast, anti-MMP9 IgGs with enzyme-blocking activity impaired renal morphogenesis, in a concentration-dependent manner, by inhibiting T-shaped branching and further divisions of the ureter bud. This effect was irreversible, still observed after inductive events and reproduced by exogenous tissue inhibitor of metalloproteinase 1 (TIMP1), the natural inhibitor of MMP9. These data provide the first demonstration of MMP9 and MMP2 production in vivo by 11-d embryonic kidneys and further show that MMP9 is required in vitro for branching morphogenesis of the ureter bud.
α3β1 Integrin Is Required for Normal Development of the Epidermal Basement Membrane
Authors: DiPersio C. Michael , Hodivala-Dilke Kairbaan M. , Jaenisch Rudolf , Kreidberg Jordan A. , Hynes Richard O.
Published: July 2002

Integrins α3β1 and α6β4 are abundant receptors on keratinocytes for laminin-5, a major component of the basement membrane between the epidermis and the dermis in skin. These integrins are recruited to distinct adhesion structures within keratinocytes; α6β4 is present in hemidesmosomes, while α3β1 is recruited into focal contacts in cultured cells. To determine whether differences in localization reflect distinct functions of these integrins in the epidermis, we studied skin development in α3β1-deficient mice. Examination of extracellular matrix by immunofluorescence microscopy and electron microscopy revealed regions of disorganized basement membrane in α3β1-deficient skin. Disorganized matrix was first detected by day 15.5 of embryonic development and became progressively more extensive as development proceeded. In neonatal skin, matrix disorganization was frequently accompanied by blistering at the dermal-epidermal junction. Laminin-5 and other matrix proteins remained associated with both the dermal and epidermal sides of blisters, suggesting rupture of the basement membrane itself, rather than detachment of the epidermis from the basement membrane as occurs in some blistering disorders such as epidermolysis bullosa. Consistent with this notion, primary keratinocytes from α3β1-deficient skin adhered to laminin-5 through α6 integrins. However, α3β1-deficient keratinocytes spread poorly compared with wild-type cells on laminin-5, demonstrating a postattachment requirement for α3β1 and indicating distinct roles for α3β1 and α6β4. Our findings support a novel role for α3β1 in establishment and/or maintenance of basement membrane integrity, while α6β4 is required for stable adhesion of the epidermis to the basement membrane through hemidesmosomes.
Movement of Bax from the Cytosol to Mitochondria during Apoptosis
Authors: Wolter Keith G. , Hsu Yi-Te , Smith Carolyn L. , Nechushtan Amotz , Xi Xu-Guang , Youle Richard J.
Published: July 2002

Bax, a member of the Bcl-2 protein family, accelerates apoptosis by an unknown mechanism. Bax has been recently reported to be an integral membrane protein associated with organelles or bound to organelles by Bcl-2 or a soluble protein found in the cytosol. To explore Bcl-2 family member localization in living cells, the green fluorescent protein (GFP) was fused to the NH2 termini of Bax, Bcl-2, and Bcl-XL. Confocal microscopy performed on living Cos-7 kidney epithelial cells and L929 fibroblasts revealed that GFP–Bcl-2 and GFP–Bcl-XL had a punctate distribution and colocalized with a mitochondrial marker, whereas GFP–Bax was found diffusely throughout the cytosol. Photobleaching analysis confirmed that GFP–Bax is a soluble protein, in contrast to organelle-bound GFP–Bcl-2. The diffuse localization of GFP–Bax did not change with coexpression of high levels of Bcl-2 or Bcl-XL. However, upon induction of apoptosis, GFP–Bax moved intracellularly to a punctate distribution that partially colocalized with mitochondria. Once initiated, this Bax movement was complete within 30 min, before cellular shrinkage or nuclear condensation. Removal of a COOH-terminal hydrophobic domain from GFP–Bax inhibited redistribution during apoptosis and inhibited the death-promoting activity of both Bax and GFP– Bax. These results demonstrate that in cells undergoing apoptosis, an early, dramatic change occurs in the intracellular localization of Bax, and this redistribution of soluble Bax to organelles appears important for Bax to promote cell death.
Activation of Membrane-associated Procaspase-3 Is Regulated by Bcl-2
Authors: Krebs Joseph F. , Armstrong Robert C. , Srinivasan Anu , Aja Teresa , Wong Angela M. , Aboy Aileen , Sayers Rob , Pham Bryan , Vu Tam , Hoang Kim , Karanewsky Donald S. , Leist Christian , Schmitz Albert , Wu Joe C. , Tomaselli Kevin J. , Fritz Lawrence C.
Published: July 2002

The mechanism by which membrane-bound Bcl-2 inhibits the activation of cytoplasmic procaspases is unknown. Here we characterize an intracellular, membrane-associated form of procaspase-3 whose activation is controlled by Bcl-2. Heavy membranes isolated from control cells contained a spontaneously activatable caspase-3 zymogen. In contrast, in Bcl-2 overexpressing cells, although the caspase-3 zymogen was still associated with heavy membranes, its spontaneous activation was blocked. However, Bcl-2 expression had little effect on the levels of cytoplasmic caspase activity in unstimulated cells. Furthermore, the membrane-associated caspase-3 differed from cytosolic caspase-3 in its responsiveness to activation by exogenous cytochrome c. Our results demonstrate that intracellular membranes can generate active caspase-3 by a Bcl-2–inhibitable mechanism, and that control of caspase activation in membranes is distinct from that observed in the cytoplasm. These data suggest that Bcl-2 may control cytoplasmic events in part by blocking the activation of membrane-associated procaspases.
The Ribosomal RNA Processing Machinery Is Recruited to the Nucleolar Domain before RNA Polymerase I during Xenopus laevis Development
Authors: Verheggen Céline , Almouzni Geneviève , Hernandez-Verdun Danièle
Published: July 2002

Transcription and splicing of messenger RNAs are temporally and spatially coordinated through the recruitment by RNA polymerase II of processing factors. We questioned whether RNA polymerase I plays a role in the recruitment of the ribosomal RNA (rRNA) processing machinery. During Xenopus laevis embryogenesis, recruitment of the rRNA processing machinery to the nucleolar domain occurs in two steps: two types of precursor structures called prenucleolar bodies (PNBs) form independently throughout the nucleoplasm; and components of PNBs I (fibrillarin, nucleolin, and the U3 and U8 small nucleolar RNAs) fuse to the nucleolar domain before components of PNBs II (B23/NO38). This fusion process is independent of RNA polymerase I activity, as shown by actinomycin D treatment of embryos and by the lack of detectable RNA polymerase I at ribosomal gene loci during fusion. Instead, this process is concomitant with the targeting of maternally derived pre-rRNAs to the nucleolar domain. Absence of fusion was correlated with absence of these pre-rRNAs in nuclei where RNA polymerase II and III are inhibited. Therefore, during X. laevis embryogenesis, the recruitment of the rRNA processing machinery to the nucleolar domain could be dependent on the presence of pre-rRNAs, but is independent of either zygotic RNA polymerase I transcription or the presence of RNA polymerase I itself.
RNA-mediated interaction of Cajal bodies and U2 snRNA genes
Authors: Frey Mark R. , Matera A. Gregory
Published: July 2002

Cajal bodies (CBs) are nuclear structures involved in RNA metabolism that accumulate high concentrations of small nuclear ribonucleoproteins (snRNPs). Notably, CBs preferentially associate with specific genomic loci in interphase human cells, including several snRNA and histone gene clusters. To uncover functional elements involved in the interaction of genes and CBs, we analyzed the expression and subcellular localization of stably transfected artificial arrays of U2 snRNA genes. Although promoter substitution arrays colocalized with CBs, constructs containing intragenic deletions did not. Additional experiments identified factors within CBs that are important for association with the native U2 genes. Inhibition of nuclear export or targeted degradation of U2 snRNPs caused a marked decrease in the levels of U2 snRNA in CBs and strongly disrupted the interaction with U2 genes. Together, the results illustrate a specific requirement for both the snRNA transcripts as well as the presence of snRNPs (or snRNP proteins) within CBs. Our data thus provide significant insight into the mechanism of CB interaction with snRNA loci, strengthening the putative role for this nuclear suborganelle in snRNP biogenesis.
Force transduction by Triton cytoskeletons
Authors: Sawada Yasuhiro , Sheetz Michael P.
Published: July 2002

Force-initiated signal transduction can occur either via membrane-based ionic mechanisms or through changes in cytoskeletal–matrix linkages. We report here the stretch-dependent binding of cytoplasmic proteins to Triton X-100 cytoskeletons of L-929 cells grown on collagen-coated silicone. Triton X-100–insoluble cytoskeletons were stretched by 10% and incubated with biotinylated cytoplasmic proteins. Analysis with two-dimensional gel electrophoresis showed stretch-dependent binding of more than 10 cytoplasmic protein spots. Bound cytoplasmic proteins were purified by a photocleavable biotin tag and stretch-dependent binding of paxillin, focal adhesion kinase, and p130Cas was found, whereas the binding of vinculin was unchanged and actin binding decreased with stretch. Paxillin binding upon stretch was morphologically and biochemically similar in vitro and in vivo, that is, enhanced in the periphery and inhibited by the tyrosine phosphatase inhibitor, phenylarsine oxide. Thus, we suggest that transduction of matrix forces occurs through force-dependent conformation changes in the integrated cytoskeleton.
Phospholipids undergo hop diffusion in compartmentalized cell membrane
Authors: Fujiwara Takahiro , Ritchie Ken , Murakoshi Hideji , Jacobson Ken , Kusumi Akihiro
Published: July 2002

The diffusion rate of lipids in the cell membrane is reduced by a factor of 5–100 from that in artificial bilayers. This slowing mechanism has puzzled cell biologists for the last 25 yr. Here we address this issue by studying the movement of unsaturated phospholipids in rat kidney fibroblasts at the single molecule level at the temporal resolution of 25 μs. The cell membrane was found to be compartmentalized: phospholipids are confined within 230-nm-diameter (ϕ) compartments for 11 ms on average before hopping to adjacent compartments. These 230-nm compartments exist within greater 750-nm-ϕ compartments where these phospholipids are confined for 0.33 s on average. The diffusion rate within 230-nm compartments is 5.4 μm2/s, which is nearly as fast as that in large unilamellar vesicles, indicating that the diffusion in the cell membrane is reduced not because diffusion per se is slow, but because the cell membrane is compartmentalized with regard to lateral diffusion of phospholipids. Such compartmentalization depends on the actin-based membrane skeleton, but not on the extracellular matrix, extracellular domains of membrane proteins, or cholesterol-enriched rafts. We propose that various transmembrane proteins anchored to the actin-based membrane skeleton meshwork act as rows of pickets that temporarily confine phospholipids.
Keratinocyte junctions and the epidermal barrier
Authors: Bazzoni Gianfranco , Dejana Elisabetta
Published: July 2002

Although intercellular junctions are known to be the major regulators of permeability of simple epithelia, they had not been thought to be important in regulating the permeability of stratified mammalian epithelia. Furuse et al. (2002)(this issue) demonstrate that functional tight junctions may indeed be a necessary part of the permeability barrier of the skin.
A Novel Inhibitory Receptor (ILT3) Expressed on Monocytes, Macrophages, and Dendritic Cells Involved in Antigen Processing
Authors: Cella Marina , Döhring Christian , Samaridis Jacqueline , Dessing Mark , Brockhaus Manfred , Lanzavecchia Antonio , Colonna Marco
Published: July 2002

Immunoglobulin-like transcript (ILT) 3 is a novel cell surface molecule of the immunoglobulin superfamily, which is selectively expressed by myeloid antigen presenting cells (APCs) such as monocytes, macrophages, and dendritic cells. The cytoplasmic region of ILT3 contains putative immunoreceptor tyrosine-based inhibitory motifs that suggest an inhibitory function of ILT3. Indeed, co-ligation of ILT3 to stimulatory receptors expressed by APCs results in a dramatic blunting of the increased [Ca2+]i and tyrosine phosphorylation triggered by these receptors. Signal extinction involves SH2-containing protein tyrosine phosphatase 1, which is recruited by ILT3 upon cross-linking. ILT3 can also function in antigen capture and presentation. It is efficiently internalized upon cross-linking, and delivers its ligand to an intracellular compartment where it is processed and presented to T cells. Thus, ILT3 is a novel inhibitory receptor that can negatively regulate activation of APCs and can be used by APCs for antigen uptake.
Distinct Roles for Signals Relayed through the Common Cytokine Receptor γ Chain and Interleukin 7 Receptor α Chain in Natural T Cell Development
Authors: Boesteanu Alina , Silva A. Dharshan De , Nakajima Hiroshi , Leonard Warren J. , Peschon Jacques J. , Joyce Sebastian
Published: July 2002

The commitment, differentiation, and expansion of mainstream α/β T cells during ontogeny depend on the highly controlled interplay of signals relayed by cytokines through their receptors on progenitor cells. The role of cytokines in the development of natural killer (NK)1+ natural T cells is less clearly understood. In an approach to define the role of cytokines in the commitment, differentiation, and expansion of NK1+ T cells, their development was studied in common cytokine receptor γ chain (γc) and interleukin (IL)-7 receptor α (IL-7Rα)–deficient mice. These mutations block mainstream α/β T cell ontogeny at an early prethymocyte stage. Natural T cells do not develop in γc-deficient mice; they are absent in the thymus and peripheral lymphoid organs such as the liver and the spleen. In contrast, NK1+ T cells develop in IL-7Rα–deficient mice in the thymus, and they are present in the liver and in the spleen. However, the absolute number of NK1+ T cells in the thymus of IL-7Rα–deficient mice is reduced to ∼10%, compared to natural T cell number in the wild-type thymus. Additional data revealed that NK1+ T cell ontogeny is not impaired in IL-2– or IL-4–deficient mice, suggesting that neither IL-2, IL-4, nor IL-7 are required for their development. From these data, we conclude that commitment and/or differentiation to the NK1+ natural T cell lineage requires signal transduction through the γc, and once committed, their expansion requires signals relayed through the IL-7Rα.
Association of Glucocorticoid Insensitivity with Increased Expression of Glucocorticoid Receptor β
Authors: Leung Donald Y.M. , Hamid Qutayba , Vottero Alessandra , Szefler Stanley J. , Surs Wendy , Minshall Eleanor , Chrousos George P. , Klemm Dwight J.
Published: July 2002

In many chronic inflammatory disorders, glucocorticoid (GC) insensitivity is a challenging clinical problem associated with life-threatening disease progression. The molecular basis of GC insensitivity, however, is unknown. Alternative splicing of the GC receptor (R) pre–messenger RNA generates a second GCR, termed GCR-β, which does not bind GCs but antagonizes the transactivating activity of the classic GCR, termed GCR-α. In the current study, we demonstrate that GC-insensitive asthma is associated with a significantly higher number of GCR-β–immunoreactive cells in peripheral blood than GC-sensitive asthmatics or normal controls. Furthermore, we show that patients with GC-insensitive asthma have cytokine-induced abnormalities in the DNA binding capability of the GCR. These abnormalities can be reproduced by transfection of cell lines with the GCR-β gene resulting in significant reduction of their GCR-α DNA binding capacity. We conclude that increased expression of GCR-β is cytokine inducible and may account for GC insensitivity in this common inflammatory condition.
Identification of an Astrocyte Cell Population from Human Brain that Expresses Perforin, a Cytotoxic Protein Implicated in Immune Defense
Authors: Gasque Philippe , Jones Jane , Singhrao Sim K. , Morgan B.
Published: July 2002

The brain is an immunoprivileged organ isolated from the peripheral immune system. However, it has been shown that resident cells, notably astrocytes and microglia, can express numerous innate immune molecules, providing the capacity to generate a local antipathogen system. Perforin is a cytolytic protein present in the granules of cytotoxic T lymphocytes and natural killer cells. Expression in cells other than those of the hemopoetic lineage has not been described. We report here that fetal astrocytes in culture (passages 2 to 15), astrocytoma, and adult astrocytes expressed perforin. Reverse transcriptase polymerase chain reaction followed by Southern blot was carried out using multiple specific primers and all cDNAs were cloned and sequenced. Human fetal astrocyte perforin cDNA sequence was ∼100% identical to the reported perforin cDNA cloned from T cells. Western blot analysis using monoclonal and polyclonal antiperforin peptide antibodies revealed a protein of 65 kD in both human fetal astrocyte and rat natural killer cell lysates (n = 4). Immunostaining followed by FACS® and confocal and electron microscopy analysis revealed that perforin was expressed by 40–50% of glial fibrillary acidic protein positive cells present in the fetal brain culture (n = 11). Perforin was not localized to granules in astrocytes but was present throughout the cytoplasm, probably in association with the endoplasmic reticulum. Perforin was not detected in normal adult brain tissue but was present in and around areas of inflammation (white and grey matter) in multiple sclerosis and neurodegenerative brains. Perforin-positive cells were identified as reactive astrocytes. These findings demonstrate that perforin expression is not unique to lymphoid cells and suggest that perforin produced by a subpopulation of astrocytes plays a role in inflammation in the brain.
The Transcription Factor Early Growth Response 1 (Egr-1) Advances Differentiation of Pre-B and Immature B Cells
Authors: Dinkel Adelheid , Warnatz Klaus , Ledermann Birgit , Rolink Antonius , Zipfel Peter F. , Bürki Kurt , Eibel Hermann
Published: July 2002

In mature B lymphocytes, the zinc finger transcription factor early growth response 1 (Egr-1) is one of the many immediate-early genes induced upon B cell antigen receptor engagement. However, its role during earlier stages of lymphopoiesis has remained unclear. By examining bone marrow B cell subsets, we found Egr-1 transcripts in pro/pre-B and immature B lymphocytes, and Egr-1 protein in pro/pre-B–I cells cultivated on stroma cells in the presence of interleukin (IL)-7. In recombinase-activating gene (RAG)-2–deficient mice overexpressing an Egr-1 transgene in the B lymphocyte lineage, pro/pre-B–I cells could differentiate past a developmental block at the B220low BP-1− stage to the stage of B220low BP-1+ pre-B–I cells, but not further to the B220low BP-1+ CD25+ stage of pre-B–II cells. Therefore, during early B lymphopoiesis progression from the B220low BP-1− IL-2R− pro/pre-B–I stage to the B220low BP-1+ IL-2R+ pre-B–II stage seems to occur in at least two distinct steps, and the first step to the stage of B220low BP-1+ pre-B–I cells can be promoted by the overexpression of Egr-1 alone. Wild-type mice expressing an Egr-1 transgene had increased proportions of mature immunoglobulin (Ig)M+ B220high and decreased proportions of immature IgM+ B220low bone marrow B cells. Since transgenic and control precursor B cells show comparable proliferation patterns, overexpression of Egr-1 seems also to promote entry into the mature B cell stage. Analysis of changes in the expression pattern of potential Egr-1 target genes revealed that Egr-1 enhances the expression of the aminopeptidase BP-1/6C3 in pre-B and immature B cells and upregulates expression of the orphan nuclear receptor nur77 in IgM+ B cells.
Modulation of  Thymic Selection by Expression of an Immediate-early Gene, Early Growth Response 1 (Egr-1)
Authors: Miyazaki Toru , Lemonnier François A.
Published: July 2002

The potential involvement of early growth response (Egr)-1, a zinc-finger transcription factor belonging to the immediate-early genes, in positive/negative selection of thymocytes has been implicated by its expression in the population of CD4+CD8+ double positive (DP) cells undergoing selection. To further investigate this possibility, transgenic mice overexpressing Egr-1 in thymocytes were bred with a transgenic mouse line expressing a T cell receptor (TCR) recognizing the H-Y male antigen in the context of H-2b class I major histocompatibility complex (MHC) molecules. In Egr-1/TCR H-Y double-transgenic mice, efficient positive selection of H-Y CD8+ T cells occurred, even in mice on either a nonselecting H-2d background or a β2-microglobulin (β2m)-deficient background in which the expression of class I MHC heavy chains is extremely low; no positive selection was observed on a Kb−/−Db−/−β2m−/− background where class I MHC expression is entirely absent. Similarly, when the Egr-1 transgene was introduced into a class II MHC–restricted TCR transgenic mouse line, Egr-1/TCR double-transgenic mice revealed increased numbers of CD4+ T cells selected by class II MHC, as well as significant numbers of CD8+ T cells selected by class I MHC (for which the transgenic TCR might have weak affinity). Thus, Egr-1 overexpression allows positive selection of thymocytes via TCR–MHC interactions of unusually low avidity, possibly by lowering the threshold of avidity required for positive selection. Supporting this possibility, increased numbers of alloreactive T cells were positively selected in Egr-1 transgenic mice, resulting in a strikingly enhanced response against allo-MHC. These results suggest that expression of Egr-1 and/or its target gene(s) may directly influence the thresholds required for thymocyte selection.
Mature Follicular Dendritic Cell Networks Depend on Expression of Lymphotoxin β Receptor by Radioresistant Stromal Cells and of Lymphotoxin β and Tumor Necrosis Factor by B Cells
Authors: Endres Robert , Alimzhanov Marat B. , Plitz Thomas , Fütterer Agnes , Kosco-Vilbois Marie H. , Nedospasov Sergei A. , Rajewsky Klaus , Pfeffer Klaus
Published: July 2002

The formation of germinal centers (GCs) represents a crucial step in the humoral immune response. Recent studies using gene-targeted mice have revealed that the cytokines tumor necrosis factor (TNF), lymphotoxin (LT) α, and LTβ, as well as their receptors TNF receptor p55 (TNFRp55) and LTβR play essential roles in the development of GCs. To establish in which cell types expression of LTβR, LTβ, and TNF is required for GC formation, LTβR−/−, LTβ−/−, TNF−/−, B cell–deficient (BCR−/−), and wild-type mice were used to generate reciprocal or mixed bone marrow (BM) chimeric mice. GCs, herein defined as peanut agglutinin–binding (PNA+) clusters of centroblasts/centrocytes in association with follicular dendritic cell (FDC) networks, were not detectable in LTβR−/− hosts after transfer of wild-type BM. In contrast, the GC reaction was restored in LTβ−/− hosts reconstituted with either wild-type or LTβR−/− BM. In BCR−/− recipients reconstituted with compound LTβ−/−/BCR−/− or TNF−/−/BCR−/− BM grafts, PNA+ cell clusters formed in splenic follicles, but associated FDC networks were strongly reduced or absent. Thus, development of splenic FDC networks depends on expression of LTβ and TNF by B lymphocytes and LTβR by radioresistant stromal cells.
Activation-Induced Inhibition of Interleukin 6–Mediated T Cell Survival and Signal Transducer and Activator of Transcription 1 Signaling
Authors: Teague T. Kent , Schaefer Brian C. , Hildeman David , Bender Jeremy , Mitchell Tom , Kappler John W. , Marrack Philippa
Published: July 2002

The cytokines interleukin (IL)-2, IL-4, IL-6, IL-7, and IL-15 have all previously been shown to inhibit resting T cell death in vitro. We have found a difference in the response of T cells to IL-6, depending on the activation status of the cells. IL-6 inhibited the death of naive T cells, but had no effect on the death of either superantigen-activated T cells, or T cells bearing memory markers. This was true even when the resting and activated T cells were isolated from the same animal; thus, the determining factor for IL-6 insensitivity was the activation status or activation history of the cell, and not the milieu in the animal from which the cells were isolated. Activated T cells expressed lower levels of IL-6 receptors on their surfaces, yet there were sufficient levels of receptors for signaling, as we observed similar levels of signal transducer and activator of transcription (Stat)3 phosphorylation in resting and activated T cells treated with IL-6. However, there was profound inhibition of IL-6–induced Stat1 phosphorylation in activated T cells compared with resting T cells. These data suggest that there is activation-induced inhibition of IL-6 receptor signaling in T cells. This inhibition appears to be specific for some but not all of the IL-6–mediated signaling cascades in these cells.
Failure to Suppress the Expansion of the Activated Cd4 T Cell Population in Interferon γ–Deficient Mice Leads to Exacerbation of Experimental Autoimmune Encephalomyelitis
Authors: Chu Cong-Qiu , Wittmer Susan , Dalton Dyana K.
Published: July 2002

Mice deficient in interferon (IFN)-γ or IFN-γ receptor develop progressive and fatal experimental autoimmune encephalomyelitis (EAE). We demonstrate that CD4 T cells lacking IFN-γ production were required to passively transfer EAE, indicating that they were disease-mediating cells in IFN-γ knockout (KO) mice. IFN-γ KO mice accumulated 10–16-fold more activated CD4 T cells (CD4+CD44hi) than wild-type mice in the central nervous system during EAE. CD4+CD44hi T cells in the spleen and central nervous system of IFN-γ KO mice during EAE showed markedly increased in vivo proliferation and significantly decreased ex vivo apoptosis compared with those of wild-type mice. IFN-γ KO CD4+CD44hi T cells proliferated extensively to antigen restimulation in vitro and accumulated larger numbers of live CD4+ CD44hi T cells. IFN-γ completely suppressed proliferation and significantly induced apoptosis of CD4+CD44hi T cells responding to antigen and hence inhibited accumulation of live, activated CD4 T cells. We thus present novel in vivo and in vitro evidence that IFN-γ may limit the extent of EAE by suppressing expansion of activated CD4 T cells.
T Cell Development and T Cell Responses in Mice with Mutations Affecting Tyrosines 292 or 315 of the Zap-70 Protein Tyrosine Kinase
Authors: Magnan Antoine , Di Bartolo Vincenzo , Mura Anne-Marie , Boyer Claude , Richelme Mireille , Lin Yea-Lih , Roure Agnès , Gillet Anne , Arrieumerlou Cécile , Acuto Oreste , Malissen Bernard , Malissen Marie
Published: July 2002

After stimulation of the T cell receptor (TCR), the tyrosine residues 292 and 315 in interdomain B of the protein tyrosine kinase ZAP-70 become phosphorylated and plausibly function as docking sites for Cbl and Vav1, respectively. The two latter proteins have been suggested to serve as substrates for ZAP-70 and to fine-tune its function. To address the role of these residues in T cell development and in the function of primary T cells, we have generated mice that express ZAP-70 molecules with Tyr to Phe substitution at position 292 (Y292F) or 315 (Y315F). When analyzed in a sensitized TCR transgenic background, the ZAP-70 Y315F mutation reduced the rate of positive selection and delayed the occurrence of negative selection. Furthermore, this mutation unexpectedly affected the constitutive levels of the CD3-ζ p21 phosphoisoform. Conversely, the ZAP-70 Y292F mutation upregulated proximal events in TCR signaling and allowed more T cells to produce interleukin 2 and interferon γ in response to a given dose of antigen. The observation that ZAP-70 Y292F T cells have a slower rate of ligand-induced TCR downmodulation suggests that Y292 is likely involved in regulating the duration activated TCR reside at the cell surface. Furthermore, we showed that Y292 and Y315 are dispensable for the TCR-induced tyrosine phosphorylation of Cbl and Vav1, respectively. Therefore, other molecules present in the TCR signaling cassette act as additional adaptors for Cbl and Vav1. The present in vivo analyses extend previous data based on transformed T cell lines and suggest that residue Y292 plays a role in attenuation of TCR signaling, whereas residue Y315 enhances ZAP-70 function.
A note on the weighted norm inequality for the one-sided maximal operator
Authors: Qinsheng Lai
Published: July 2002

Let

M
g
+

M_g^+

be the one-sided maximal function. In this note we obtain some necessary and sufficient conditions in order that the weighted weak type inequality holds for

M
g
+

M_g^+

. Meanwhile, some necessary or sufficient conditions for the weighted inequality for

M
g
+

M_g^+

are given.
Single-point condensation and least-energy solutions
Authors: Ren Xiaofeng , Wei Juncheng
Published: July 2002

We prove a conjecture raised in our earlier paper which says that the least-energy solutions to a two-dimensional semilinear problem exhibit single-point condensation phenomena as the nonlinear exponent gets large. Our method is based on a sharp form of a well-known borderline case of the Sobolev embedding theory. With the help of this embedding, we can use the Moser iteration scheme to carefully estimate the upper bound of the solutions. We can also determine the location of the condensation points.
Steiner systems 𝑆(5,6,𝑣) with 𝑣=72 and 84
Authors: Grannell M. , Griggs T. , Mathon R.
Published: July 2002

It is proved that there are precisely 4204 pairwise non-isomorphic Steiner systems

S
(
5
,
6
,
72
)

S(5,6,72)

invariant under the group

P
S
L

2

(
71
)

\mathrm {PSL}_2(71)

and which can be constructed using only short orbits. It is further proved that there are precisely 38717 pairwise non-isomorphic Steiner systems

S
(
5
,
6
,
84
)

S(5,6,84)

invariant under the group

P
S
L

2

(
83
)

\mathrm {PSL}_2(83)

and which can be constructed using only short orbits.
Intersection theory on \overline{ℳ}_{1,4} and elliptic Gromov-Witten invariants
Authors: Getzler E.
Published: July 2002

We find a new relation among codimension

2
2

algebraic cycles in the moduli space

M

¯

1
,
4

\bar {\mathcal {M}}_{1,4}

, and use this to calculate the elliptic Gromov-Witten invariants of projective spaces

C
P

2

\mathbb {CP}^2

and

C
P

3

\mathbb {CP}^3

.
Virtual moduli cycles and Gromov-Witten invariants of algebraic varieties
Authors: Li Jun , Tian Gang
Published: July 2002

In this paper, we define the virtual moduli cycle of moduli spaces with perfect tangent-obstruction theory. The two interesting moduli spaces of this type are moduli spaces of vector bundles over surfaces and moduli spaces of stable morphisms from curves to projective varieties. As an application, we define the Gromov-Witten invariants of smooth projective varieties and prove all its basic properties.
Une compactification des champs classifiant les chtoucas de Drinfeld
Authors: Lafforgue Laurent
Published: July 2002

One knows that the notion of Harder–Narasimhan’s canonical polygon allows one to write the stacks classifying Drinfeld’s shtukas of rank at least

2
2

as inductive limits of open substacks of finite type. When there is no level structure, we present here a smooth modular compactification of each such open substack, generalizing Drinfeld’s construction for rank 

2
2

. Résumé. On sait qu’en rang au moins

2
2

, la notion de polygone canonique de Harder–Narasimhan permet d’écrire les champs classifiant les chtoucas de Drinfeld comme des réunions filtrantes d’ouverts de type fini. Quand il n’y a pas de structure de niveau, on présente ici une compactification modulaire lisse de chacun de ces ouverts, généralisant celles de Drinfeld en rang 

2
2

.
MUTbase: maintenance and analysis of distributed mutation databases.
Authors: Riikonen P , Vihinen M
Published: July 2002

MOTIVATION: To develop tools for the submission of mutations to databases and maintenance of locus-specific mutation databases. Advanced, integrated computer systems are needed to store and organize the increasing mutation information. RESULTS: The MUTbase program suite provides an easy, interactive and quality-controlled submission of information to mutation databases. For further study of the databases on the World Wide Web, a number of tools are provided. The program package also writes and updates a large number of Web pages, e.g. about the distribution and statistics of disease-causing mutations, and changes in restriction patterns.
Semiotes: a semantics for sharing
Authors: Kazic Toni
Published: July 2002

Motivation: Reliable, automated communication of biological information requires methods to declare the information’s semantics. In this paper I describe an approach to semantic declaration intended to permit independent, distributed databases, algorithms, and servers to exchange and process requests for information and computations without requiring coordination or agreement among them on universe of discourse, data model, schema, or implementation.

Results: This approach uses Glossa , a formal language defining the semantics of biological ideas, information, and algorithms, to executably define the semantics of complex ideas and computations by constructs of semiotes , terms which axiomatically define very simple notions. A database or algorithm wishing to exchange information or computations maintains a set of mappings between its particular notions and semiotes, and a parser to translate between its indigenous ideas and implementation and the semiotes. Requests from other databases or algorithms are issued as semiotic messages, locally interpreted and processed, and the results returned as semiotes to the requesting entity. Thus, semiotes serve as a shared, abstract layer of definitions which can be computably combined by each database or algorithm according to its own needs and ideas. By combining the explicit declaration of semantics with the computation of the semantics of complex ideas, Glossa and its semiotes permit independent computational entities to lightly federate their capabilities as desired while maintaining their unique perspectives on both scientific and technical questions.

Availability: http://www.ibc.wustl.edu/agora/semiotes/ and files therein.

Contact: toni@athe.wustl.edu
An activation‐independent role of transcription factors in insulator function
Authors: Fourel Geneviève , Boscheron Cécile , Revardel Émanuelle , Lebrun Éléonore , Hu Yan‐Fen , Simmen Katia Carmine , Müller Karin , Li Rong , Mermod Nicolas , Gilson Éric
Published: July 2002

Chromatin insulators are defined as transcriptionally neutral elements that prevent negative or positive influence from extending across chromatin to a promoter. Here we show that yeast subtelomeric anti‐silencing regions behave as boundaries to telomere‐driven silencing and also allow discontinuous propagation of silent chromatin. These two facets of insulator activity, boundary and silencing discontinuity, can be recapitulated by tethering various transcription activation domains to tandem sites on DNA. Importantly, we show that these insulator activities do not involve direct transcriptional activation of the reporter promoter. These findings predict that certain promoters behave as insulators and partition genomes in functionally independent domains.
NurA, a novel 5′–3′ nuclease gene linked to rad50 and mre11 homologs of thermophilic Archaea
Authors: Constantinesco Florence , Forterre Patrick , Elie Christiane
Published: July 2002

We isolated and characterized a new nuclease (NurA) exhibiting both single‐stranded endonuclease activity and 5′–3′ exonuclease activity on single‐stranded and double‐stranded DNA from the hyperthermophilic archaeon Sulfolobus acidocaldarius. Nuclease homologs are detected in all thermophilic archaea and, in most species, the nurA gene is organized in an operon‐like structure with rad50 and mre11 archaeal homologs. This nuclease might thus act in concert with Rad50 and Mre11 proteins in archaeal recombination/repair. To our knowledge, this is the first report of a 5′–3′ nuclease potentially associated with Rad50 and Mre11‐like proteins that may lead to the processing of double‐stranded breaks in 3′ single‐stranded tails.
Genetic requirement of p47 phox for superoxide production by murine microglia
Authors: Lavigne Mark C. , Malech Harry L. , Holland Steven M. , Leto Thomas L.
Published: July 2002

An NADPH oxidase is thought to function in microglial cells of the central nervous system. These conclusions are based on pharmacological and immunochemical evidence, although these approaches are indirect and raise issues of specificity. For example, diphenyleneiodonium inhibits a variety of flavoenzymes, including xanthine oxidase, NADH dehydrogenase, and NADPH oxidase. Here, we provide genetic evidence that p47 phox, an essential component of the phagocyte NADPH oxidase, is required for superoxide anion release from microglia. Microglia derived from newborn wild‐type mice, but not from newborn p47 phox‐deficient (“knockout”; ‐/‐) mice, produced superoxide after stimulation by opsonized zymosan or phorbol myristate acetate. Endogenous p47 phox was detected only in wild‐type microglia, consistent with selective superoxide production in these cells. Superoxide release was restored in p47 phox‐deficient microglia that were retrovirally transduced with human p47 phox cDNA. Similar kinetics of superoxide generation were observed, consistent with the same enzyme functioning in wild‐type and restored microglia. Immuno‐detection of p47 phox in transduced cells confirmed that restoration of superoxide release correlated with production of recombinant protein. These data provide genetic proof that p47 phox is necessary for superoxide release by microglial cells and indicate that a system related to the phagocyte oxidase is active in these cells.
Single units and conscious vision
Authors:   The American Association for Research into Nervous and Mental Diseases, Logothetis N. K.
Published: July 2002

Figures that can be seen in more than one way are invaluable tools for the study of the neural basis of visual awareness, because such stimuli permit the dissociation of the neural responses that underlie what we perceive at any given time from those forming the sensory representation of a visual pattern. To study the former type of responses, monkeys were subjected to binocular rivalry, and the response of neurons in a number of different visual areas was studied while the animals reported their alternating percepts by pulling levers. Perception–related modulations of neural activity were found to occur to different extents in different cortical visual areas. The cells that were affected by suppression were almost exclusively binocular, and their proportion was found to increase in the higher processing stages of the visual system. The strongest correlations between neural activity and perception were observed in the visual areas of the temporal lobe. A strikingly large number of neurons in the early visual areas remained active during the perceptual suppression of the stimulus, a finding suggesting that conscious visual perception might be mediated by only a subset of the cells exhibiting stimulus selective responses. These physiological findings, together with a number of recent psychophysical studies, offer a new explanation of the phenomenon of binocular rivalry. Indeed, rivalry has long been considered to be closely linked with binocular fusion and stereopsis, and the sequences of dominance and suppression have been viewed as the result of competition between the two monocular channels. The physiological data presented here are incompatible with this interpretation. Rather than reflecting interocular competition, the rivalry is most probably between the two different central neural representations generated by the dichoptically presented stimuli. The mechanisms of rivalry are probably the same as, or very similar to, those underlying multistable perception in general, and further physiological studies might reveal a much about the neural mechanisms of our perceptual organization.
Virus reconstitution and the proof of the existence of genomic RNA
Authors: Fraenkel-Conrat H. , Singer B.
Published: July 2002

This paper is a historical overview of the work done on the tobacco mosaic virus. The primary finding was that a virus is capable of reassembling itself from its component protein and RNA, and that only the RNA carries the genomic capability of the virus. This was followed by detailed studies of the chemical and biological properties of viral RNA.
Energy dissipation and radical scavenging by the plant phenylpropanoid pathway
Authors: Grace Stephen C. , Logan Barry A.
Published: July 2002

Environmental stresses such as high light, low temperatures, pathogen infection and nutrient deficiency can lead to increased production of free radicals and other oxidative species in plants. A growing body of evidence suggests that plants respond to these biotic and abiotic stress factors by increasing their capacity to scavenge reactive oxygen species. Efforts to understand this acclimatory process have focused on the components of the ‘classical’ antioxidant system, i.e. superoxide dismutase, ascorbate peroxidase, catalase, monodehydroascorbate reductase, glutathione reductase and the low molecular weight antioxidants ascorbate and glutathione. However, relatively few studies have explored the role of secondary metabolic pathways in plant response to oxidative stress. A case in point is the phenylpropanoid pathway, which is responsible for the synthesis of a diverse array of phenolic metabolites such as flavonoids, tannins, hydroxycinnamate esters and the structural polymer lignin. These compounds are often induced by stress and serve specific roles in plant protection, i.e. pathogen defence, ultraviolet screening, antiherbivory, or structural components of the cell wall. This review will highlight a novel antioxidant function for the taxonomically widespread phenylpropanoid metabolite chlorogenic acid (CGA; 5-
O
-caffeoylquinic acid) and assess its possible role in abiotic stress tolerance. The relationship between CGA biosynthesis and photosynthetic carbon metabolism will also be discussed. Based on the properties of this model phenolic metabolite, we propose that under stress conditions phenylpropanoid biosynthesis may represent an alternative pathway for photochemical energy dissipation that has the added benefit of enhancing the antioxidant capacity of the cell.