Massimo Mazzini

Viral particles of the endogenous retrovirus ZAM from Drosophila melanogaster use a pre-existing endosome/exosome pathway for transfer to the oocyte

BackgroundRetroviruses have evolved various mechanisms to optimize their transfer to new target cells via late endosomes. Here, we analyzed the transfer of ZAM, a retroelement from Drosophila melanogaster, from ovarian follicle cells to the oocyte at stage 9–10 of oogenesis, when an active yolk transfer is occurring between these two cell types.ResultsCombining genetic and microscopic approaches, we show that a functional secretory apparatus is required to tether ZAM to endosomal vesicles and to direct its transport to the apical side of follicle cells. There, ZAM egress requires an intact follicular epithelium communicating with the oocyte. When gap junctions are inhibited or yolk receptors mutated, ZAM particles fail to sort out the follicle cells.ConclusionOverall, our results indicate that retrotransposons do not exclusively perform intracellular replication cycles but may usurp exosomal/endosomal traffic to be routed from one cell to another.

Qualitative and quantitative analysis of serum immunoglobulins of four Antarctic fish species

Abstract Immunoglobulins from the Antarctic fish species Trematomus bernacchii, Trematomus hansoni, Trematomus newnesi, and Chionodraco hamatus were analysed in whole serum and after purification by affinity chromatography on protein A-sepharose. Using SDS-PAGE, the apparent masses of the heavy and light chains were, respectively, 83.5u2009kDa and 27.5u2009kDa for T. bernacchii, 83.5 kDa and 27 kDa for T. hansoni, 81 kDa and 27.5 kDa for T. newnesi, and 74.5 kDa and 30 kDa for C. hamatus. It was not possible to purify immunoglobulins from T. newnesi due to their low concentration in serum. Heterogeneity in mass of both heavy and light chains was observed in all species. By using a polyclonal antibody raised against sea bass immunoglobulins, cross-reactivity was observed with heavy and light chains of all species. With this antibody, an indirect enzyme-linked immunosorbent assay (ELISA) was developed and results showed the relative immunoglobulin concentration in sera of the Antarctic fish species considered, using as standard sea bass immunoglobulins.

Biological activity of cytokines : An evolutionary perspective

It appears evident that teleost fish are at present the vertebrate group in which, excluding mammals, most information on the immune system is available. However, despite the great impetus on the discovery of genes homologous to mammalian immunomodulatory molecules, the knowledge on biological activities exerted by cytokines is meager. This review reports the present knowledge on the biological activities of cytokine-like and cytokines in invertebrates and cold-blooded vertebrates.

Cytology of lymphomyeloid head kidney of Antarctic fishes Trematomus bernacchii (Nototheniidae) and Chionodraco hamatus (Channicthyidae).

Species that live in extreme conditions have specially adapted physiology and tissue/organ organisation. The adaptation of lymphoid organs to low temperatures in polar species could be an original field of study, indicating how the immune system works under extreme conditions. In fishes, the head kidney is a key organ for immunity and here the cytology of this organ is studied in two common Antarctic species: Trematomus bernacchii and Chionodraco hamatus. Ultrastructural analysis revealed heterogeneity of epithelial cells, with reticular cells, subcapsular- and perivascular-limiting cells. Differences in the size and morphology of epithelial cells were observed between the polar species and warm water species of fish. Intermingled with epithelial cell leucocytes, such as lymphocytes, thrombocytes and macrophages, had comparable morphology in both species, contrary to sharp differences observed in the morphology of erythrocytes and granulocytes. The functional adaptation of the head kidney to the low temperatures of polar water is discussed.

Yolk granules are differentially acidified during embryo development in the stick insect Carausius morosus

Abstract. Newly laid eggs of stick insects comprise a unique fluid ooplasm that is gradually partitioned into a number of yolk granules by invasion of secondary vitellophages. This study aimed at establishing how yolk granules become acidified in the course of embryonic development. Data show that acidified yolk granules are rather scarce and randomly distributed in vitellophages of early embryos, while they tend to increase gradually in number as development proceeds to completion. Yolk granule acidification is progressively more inhibited in the presence of increasing concentrations of chloroquine, monensin and bafilomycin. A pro-protease was identified cytochemically and by immunoblotting in yolk extracts of progressively more advanced embryos. A specific monoclonal antibody raised against this pro-protease helped to demonstrate that it is gradually processed to yield a lower molecular weight polypeptide as development proceeds to completion. This latter polypeptide was identified as a protease using electrophoresis in polyacrylamide gels containing yolk extracts. Simultaneous administration of a fluorescent substrate for cysteine protease and an acidotropic probe produced superimposable labelling patterns, suggesting that only acidified yolk granules possess a proteolytic activity. On the other hand, yolk granules probed simultaneously for acidification and latent pro-protease yielded labelling patterns partially superimposed. Pro-protease labelling is gradually lost as yolk granules are progressively more acidified during development. Distinct labelling patterns were also obtained in vitellophages processed for the simultaneous detection of pro-protease and protease, suggesting that the two activities are expressed by different yolk granule populations, and that one is gradually converted into the other as time goes by.

Ultrastructure and proteins of the egg chorion of the antarctic fish Chionodraco hamatus (Teleostei, Notothenioidei)

Abstract. The chorion morphology and protein content of unfertilised eggs of the antarctic icefish Chionodraco hamatus were characterised. Scanning and transmission electron microscopy showed that the chorion of this antarctic species is organised differently from that of non-polar species, with several concentric layers varying in thickness. Purified chorions were dissolved in 8-M urea buffer, and polypeptides were revealed by one- and two-dimensional gel electrophoresis. Glycoproteins were detected using affinity blotting with concanavalin-A. The principal glycoproteins had a molecular weight of 49 and 93xa0kDa. Purification of the main polypeptides between 40 and 92xa0kDa was achieved by preparative electrophoresis followed by electroelution of excised bands. The main biochemical composition of C. hamatus chorion was similar to that of other teleost species investigated.

Histological observations on lymphomyeloid organs of the Antarctic fish Trematomus bernacchii (Teleostei: Nototheniidae)

Abstract Lymphomyeloid organs of the Antarctic fish Trematomus bernacchii were studied with the aim of analysing some morphological aspects related to adaptation to low environmental temperature. The thymus of T. bernacchii was flattened, incompletely lobated and scarcely regionalised. It was filled by lymphoid elements intermingled with stromal elements. The head kidney appeared highly vascularised and mainly lymphopoietic. The spleen appeared mainly erythropoietic, with scarcely developed areas of white pulp.

An Ultrastructural Investigation on Vitellophage Invasion of the Yolk Mass during and after Germ Band Formation in Embryos of the Stick Insect Carausius morosus Br.

Developing embryos of the stick insect Carausius morosus were examined ultrastructurally with a view to studying vitellophage invasion of the yolk mass during and after germ band formation. Newly laid eggs in C.morosus have a unique yolk fluid compartment surrounded by a narrow fringe of cytoplasm comprising several small yolk granules. Vitellophages originate mainly from a thin layer of stem cells, the so‐called yolk cell membrane, interposed between the germ band and the yolk mass. Throughout development, a thin basal lamina separates the yolk cell membrane from the overlying embryo.

Oocyte growth, follicle cell differentiation and vitellin processing in the stick insect, Carausius morosus Br. (Phasmatodea)

Abstract Ovarian growth in stick insects (Phasmatodea) was examined ultrastructurally and cytochemically with a view to studying: (1) the kinetics of oocyte growth and the staging characteristics of ovarian follicles undergoing vitellogenesis; (2) the endocytic capability of the growing oocyte, including the post-endocytic fate of the vitellins sequestered by the oocyte during vitellogenesis; (3) the differentiation of the follicular epithelium in relation to the appearance of intercellular spaces and the extracellular release of a follicle cell product. These structural observations were interpreted in relation to the nature and kinetics of the vitellin processing in follicles undergoing vitellogenesis.

Characterization of a Monoclonal Antibody Against a 180 kDa Hemocyte Polypeptide Involved in Cellular Defence Reactions of the Stick Insect Bacillus rossius

Defence properties of hemoctyes were investigated using the anti-hemocyte monoclonal antibody BrH1 obtained by immunizing mice with 2% paraformaldehyde-fixed hemoctyes of the stick insect Bacillus rossius. In Western blot analysis, the antibody recognized a 180 kDa antigen in hemocyte cell lysates, whereas fat body lysates and cell-free hemolymph were negative. In immunofluorescence analysis of cultured or freshly collected hemoctyes, BrH1 stained intracellular antigen(s) in detergent-treated cells. Transverse cryosections of adult stick insects probed by immunofluorescence with BrH1 showed in situ the scattered distribution of hemoctyes inside the haemocoel. The antigen(s) recognized by BrH1 appears to be involved in cell defence hemocyte-mediated mechanisms, as evidenced by the fact that cryosections of insects challenged in vivo with yeast cells, bacteria, or polystyrene latex particles and probed with BrH1 showed an accumulation of antigen surrounding the injected stimuli. Copyright 1997 Elsevier Science Ltd. All rights reserved