The antiapoptotic herpes simplex virus glycoprotein J localizes to multiple cellular organelles and induces reactive oxygen species formation

The Us5 gene of herpes simplex virus (HSV) encodes glycoprotein J (gJ). The only previously reported function of gJ was its ability to inhibit apoptosis. However, the mechanism by which gJ prevents apoptosis is not understood, and it is not known whether gJ mediates additional cellular effects. In this study, we evaluated the expression, localization, and cellular effects of Us5/gJ. Us5 was first expressed 4 h after infection. gJ was detectable at 6 h and was expressed in glycosylated and unglycosylated forms. Us5 was regulated as a late gene, with partial dependency on DNA replication for expression. Us5 expression was delayed in the absence of ICP22; furthermore, expression of Us5 in trans protected cells from apoptosis induced by an HSV mutant with deletion of ICP27, suggesting that the antiapoptotic effects of ICP22 and ICP27 are mediated in part through effects on gJ expression. Within HSV-infected or Us5-transfected cells, gJ was distributed widely, especially to the endoplasmic reticulum, trans-Golgi network, and early endosomes. gJ interacted with F_oF₁ ATP synthase subunit 6 by a yeast two-hybrid screen and had strong antiapoptotic effects, which were mediated by protein rather than mRNA. Antiapoptotic activity required the extracellular and transmembrane domains of gJ, but not the intracellular domain. Consistent with inhibition of F_oF₁ ATP synthase function, Us5 was required for HSV-induced reactive oxygen species (ROS) formation, and gJ was sufficient to induce ROS in Us5-transfected cells. Thus, HSV gJ is a multifunctional protein, modulating other cellular processes in addition to inhibition of apoptosis.     

Polysialic acid regulates the clustering, migration, and neuronal differentiation of progenitor cells in the adult hippocampus

Newborn cells of the adult dentate gyrus in the hippocampus are characterized by their abundant expression of polysialic acid (PSA), a carbohydrate attached to the neural cell adhesion molecule (NCAM). PSA+ newborn cells of the dentate gyrus form clusters with proliferating neural progenitor cells, migrate away from these clusters, and terminally differentiate. To identify the roles of PSA in the development of adult progenitors of the dentate gyrus, we injected endoneuraminidase N (endoN) into the hippocampus of adult rats to specifically cleave PSA from NCAM. Two days later, we administered the mitotic marker, 5-bromo-2'-deoxyuridine (BrdU). Three days after BrdU injection, BrdU+ cells were found inside and outside the clusters of newborn cells. In endoN-treated animals, the total number of BrdU+ cells was not changed but significantly more BrdU+ cells were present within clusters, suggesting that PSA normally facilitates the migration of progenitors away from the clusters. Seven days post-BrdU injection, endoN-treated animals had significantly more BrdU+ cells which were also positive for the mature neuronal nuclear marker NeuN compared with controls, indicating that the loss of PSA from progenitor cells increases neuronal differentiation. This report is the first demonstration that PSA is involved in controlling the spatio-temporal neuronal maturation of adult hippocampal progenitors in the normal brain. In vitro, the removal of PSA from adult-derived neural progenitors significantly enhanced neuronal differentiation, strengthening our in vivo findings and indicating that PSA removal on isolated progenitor cells, apart from a complex in vivo environment, induces neuronal maturation.     

Generation and characterization of anti-MUC4 monoclonal antibodies reactive with normal and cancer cells in humans.

We have previously cloned the full-length cDNA (approximately 28 Kb) and established the complete genomic organization (25 exons/introns over 100 kb) of the human MUC4 mucin. This large molecule is predicted to protrude over 2 microm above the cell surface, in which MUC4alpha is an extracellular mucin-type glycoprotein subunit and MUC4beta is the transmembrane subunit. Over two thirds of the encoded protein sequence consists of 16-amino-acid tandem repeats (TR), which are flanked by unique sequences. In this study we generated and characterized monoclonal antibodies (MAbs) directed against the TR region of MUC4. Mice were immunized with a KLH-conjugated MUC4 TR peptide, STGDTTPLPVTDTSSV. Several clones were purified by three rounds of limited dilutions and stable clones presenting a sustained antibody production were selected for subsequent characterization. Antibodies were tested for their reactivity and specificity to recognize the MUC4 peptide and further screened by enzyme-linked immunosorbent assay (ELISA) and Western blotting analyses. One of the MAbs (8G7) was strongly reactive against the MUC4 peptide and with native MUC4 from human tissues or pancreatic cancer cells in Western blotting, immunohistochemistry, and confocal analysis. Anti-MUC4 MAb may represent a powerful tool for the study of MUC4 function under normal and pathological conditions and for diagnosis of solid tumors including those in the breast, pancreas, lungs, and ovaries.     

Modelling of the Coupling between Brain Electrical Activity and Metabolism

In order to make an attempt at grouping the various aspects of brain functional imaging (fMRI, MRS, EEG-MEG, ...) within a coherent frame, we implemented a model consisting of a system of differential equations, that includes: (1) sodium membrane transport, (2) Na/K ATPase, (3) neuronal energy metabolism (i.e. glycolysis, buffering effect of phosphocreatine, and mitochondrial respiration), (4) blood-brain barrier exchanges and (5) brain hemodynamics, all the processes which are involved in the activation of brain areas. We assumed that the correlation between brain activation and metabolism could be due to either changes in the concentrations of ATP and ADP following activation of Na/K ATPase that result from the changes in ion concentrations, or the involvement, in different phases of metabolism, of a second messenger such as calcium. In this article, we show how this type of model enables interpretation of MRS and fMRI published data that were obtained during prolonged stimulations.     

Molecular phylogeny of swallowtail butterflies of the tribe Papilionini (Papilionidae, Lepidoptera).

Swallowtail butterflies of the tribe Papilionini number about 225 species and are currently used as model organisms in several research areas, including genetics, chemical ecology and phylogenetics of host plant utilization and mimicry, mechanisms of speciation, and conservation. We have inferred phylogenetic relationships for a sample of 18 species of the genus Papilio (sensu lato) and five outgroup taxa by sequencing two stretches of mitochondrial DNA that correspond to segments 12886-13370 and 12083-12545 of Drosophila melanogaster mitochondrial DNA and consist of sections of the genes for the large ribosomal RNA and subunit 1 of NADH-dehydrogenase. Our data support the monophyly of Papilio and, within it, of several traditionally recognized subgroups. Species belonging to groups that utilize primarily Rutaceae as larval foodplants form two clusters, corresponding to Old World and American taxa, respectively, while two previously recognized clades-of American and South Asian-Austronesian origin-whose members were known to feed mostly on Lauraceae and Magnoliaceae, are observed to form a clade. The sister group of Papilio is found to be the South Asian genus Meandrusa, which also happens to feed on Lauraceae. The latter plant family is therefore the probable larval host of the ancestor Papilio and the shift to Rutaceae (which four-fifths of extant Papilio species use as foodplants) is more likely to have occurred only after the initial diversification of the genus.     

Circular dichroic studies on Cu(II) interactions with a protamine,scylliorhinine Z3

The interaction of Cu(II) with the protamine scylliorhinine Z₃ was studied by means of CD measurements. At a 1:1 molar ratio, three complexes are formed. (1) In the pH range 5–6.5, the results suggest the formation of a five-membered chelate ring through the coordination of two nitrogen atoms, the N-terminal and the contiguous peptide nitrogen. (2) At pH ≥ 6.4, there is involvement of the lateral NH₂ group of Arg; at pH 6.5–8, the formation of a 3N cupric complex is strongly suggested. (3) At pH ≥ 8, results indicate the formation of a 4N complex as a major species in Cu(II)-Z₃ solution. The transformation from a 2N to a 3N complex, and from a 3N to a 4N complex was followed with the help of the σ(αNH₂) → Cu(II) charge-transfer dichroic band transitions. At Cu(II):Z₃ molar ratios ≥ 2 and at pH > 8, a new dichroic band appears, indicating the involvement of the tyrosine residue side chain in metal-ion complexation.