Unitus DSpace

Studio dell’espressione di geni coinvolti nello sviluppo di un sistema immunitario funzionale nella spigola (Dicentrarchus labrax L.)

2011-03-08T01:30:18Z

Title: Studio dell’espressione di geni coinvolti nello sviluppo di un sistema immunitario funzionale nella spigola (Dicentrarchus labrax L.) Authors: Guerra, Laura Description: Dottorato di ricerca in Evoluzione biologica e biochimica

A CD4 homologue in sea bass (Dicentrarchus labrax): Molecular characterization and structural analysis

2011-06-27T20:38:24Z

Title: A CD4 homologue in sea bass (Dicentrarchus labrax): Molecular characterization and structural analysis Authors: Buonocore, Francesco; Randelli, Elisa; Casani, Daniela; Guerra, Laura; Picchietti, Simona; Costantini, Susan; Facchiano, Angelo M.; Zou, Jun; Secombes, Chris J.; Scapigliati, Giuseppe Abstract: CD4 is a transmembrane glycoprotein fundamental for cell-mediated immunity. Its action as a T cell coreceptor increases the avidity of association between a T cell and an antigen-presenting cell by interacting with portions of the complex between MHC class II and TR molecules. In this paper we report the cDNA cloning, expression and structural analysis of a CD4 homologue from sea bass (Dicentrarchus labrax). The sea bass CD4 cDNA consists of 2071 bp that translates in one reading frame to give the entire molecule containing 480 amino acids. The analysis of the sequence shows the presence of four putative Ig-like domains and that some fundamental structural features, like a disulphide bond in domain D2 and the CXC signalling motif in the cytoplasmic tail, are conserved from sea bass to mammals. Real-time PCR analysis showed that very high levels of CD4 mRNA transcripts are present in thymus, followed by gut and gills. In vitro stimulation of head kidney leukocytes with LPS and PHA-L gave an increase of CD4 mRNA levels after 4 h and a decrease after 24 h. Homology modelling has been applied to create a 3D model of sea bass CD4 and to investigate its interaction with sea bass MHC-II. The analysis of the 3D complex between sea bass CD4 and sea bass MHC-II suggests that the absence of a disulfide bond in the CD4 D1 domain could make this molecule more flexible, inducing a different conformation and affecting the binding and the way of interaction between CD4 and MHC-II. Our results will add new insights into the sea bass T cell immune responses and will help in the identification of T cell subsets in teleost fishes to better understand the evolution of cell-mediated immunity from fish to mammals. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/

CD4 homologue in sea bass (Dicentrarchus labrax): molecular characterization and structural analysis

2011-06-28T09:57:19Z

Title: CD4 homologue in sea bass (Dicentrarchus labrax): molecular characterization and structural analysis Authors: Buonocore, Francesco; Randelli, Elisa; Casani, Daniela; Guerra, Laura; Picchietti, Simona; Costantini, Susan; Facchiano, Angelo M.; Zou, Jun; Secombes, Chris J.; Scapigliati, Giuseppe Abstract: CD4 is a transmembrane glycoprotein fundamental for cell-mediated immunity. Its action as a T cell coreceptor increases the avidity of association between a T cell and an antigen-presenting cell by interacting with portions of the complex between MHC class II and TR molecules. In this paper we report the cDNA cloning, expression and structural analysis of a CD4 homologue from sea bass (Dicentrarchus labrax). The sea bass CD4 cDNA consists of 2071 bp that translates in one reading frame to give the entire molecule containing 480 amino acids. The analysis of the sequence shows the presence of four putative Ig-like domains and that some fundamental structural features, like a disulphide bond in domain D2 and the CXC signalling motif in the cytoplasmic tail, are conserved from sea bass to mammals. Real-time PCR analysis showed that very high levels of CD4 mRNA transcripts are present in thymus, followed by gut and gills. In vitro stimulation of head kidney leukocytes with LPS and PHA-L gave an increase of CD4 mRNA levels after 4 h and a decrease after 24 h. Homology modelling has been applied to create a 3D model of sea bass CD4 and to investigate its interaction with sea bass MHC-II. The analysis of the 3D complex between sea bass CD4 and sea bass MHC-II suggests that the absence of a disulfide bond in the CD4 D1 domain could make this molecule more flexible, inducing a different conformation and affecting the binding and the way of interaction between CD4 and MHC-II. Our results will add new insights into the sea bass T cell immune responses and will help in the identification of T cell subsets in teleost fishes to better understand the evolution of cell-mediated immunity from fish to mammals. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/

Do all stoneflies nymphs have respiratory proteins? Further data on the presence of hemocyanin in the larval stages of plecoptera species

2011-06-13T10:44:47Z

Title: Do all stoneflies nymphs have respiratory proteins? Further data on the presence of hemocyanin in the larval stages of plecoptera species Authors: Amore, Valentina; Belardinelli, Maria Cristina; Guerra, Laura; Buonocore, Francesco; Fausto, Anna Maria; Fochetti, Romolo Description: L'articolo è disponibile sul sito dell'editore http://onlinelibrary.wiley.com

Lymphocyte differentiation in sea bass thymus: CD4 and CD8-α gene expression studies

2011-06-28T14:12:59Z

Title: Lymphocyte differentiation in sea bass thymus: CD4 and CD8-α gene expression studies Authors: Picchietti, Simona; Guerra, Laura; Buonocore, Francesco; Randelli, Elisa; Fausto, Anna Maria; Abelli, Luigi Abstract: Different developmental stages (from eggs to 1-year-old juveniles) of the teleost fish Dicentrarchus labrax (L.) were assayed for CD4 gene expression. RT-PCR revealed the appearance of CD4 transcripts in postlarvae from 51 days post-hatching (dph). This finding overlaps the first detection of CD8-a mRNA. Real-time PCR with specific primers quantified CD4, CD8-a and TCR-b transcripts in larvae and postlarvae (25, 51, 75 and 92 dph) and 1-year-old thymus. At 92 dph, TcR-b and CD8-a transcripts were significantly higher (P < 0.001) than in previous stages, as CD4 transcripts compared with 51 dph (P < 0.01). High levels of TCR-b and CD8-a transcripts were found in the thymus, while CD4 transcripts were lower (P < 0.05 vs. TCR-b). In situ hybridization identified CD4 mRNAs at 51 dph, localized in thymocytes of the outer and lateral zones of the thymic glands. From 75 dph on the signal was mainly detected in the outer region, drawing a cortex–medulla demarcation. Developmental expression of CD4 and CD8-a almost coincided. In each adult thymic lobe CD4þ and CD8-aþ thymocytes filled the cortex. The expression patterns of CD4 and CD8-a largely overlap, except in the medulla, where CD4þ thymocytes were isolated, while CD8-aþ ones mainly arranged in cords. These results provide new information about the thymic compartmentalization and lymphocyte differentiation pathways in a teleost, almost demonstrating that double negative thymocytes fill the cortex giving rise to further selection in the medulla. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/

Compartmentalisation of T cells expressing CD8α and TCRβ in developing thymus of sea bass Dicentrarchus labrax (L.)

2011-06-28T10:25:57Z

Title: Compartmentalisation of T cells expressing CD8α and TCRβ in developing thymus of sea bass Dicentrarchus labrax (L.) Authors: Picchietti, Simona; Guerra, Laura; Selleri, Luigi; Buonocore, Francesco; Abelli, Luigi; Scapigliati, Giuseppe; Mazzini, Massimo; Fausto, Anna Maria Abstract: Eggs, larvae, post-larvae and sexually immature juveniles of the teleost Dicentrarchus labrax (L.) were assayed for the expression of genes encoding the T cell receptor b and CD8a. RT-PCR of RNA extracted from larvae revealed TCRb transcripts from day 25 posthatching (ph) and CD8a transcripts from 26 days later. At day 51 ph, CD8a and TCRb mRNAs were localised by in situ hybridisation in thymocytes of the outer and lateral zones of the thymic paired glands. From day 75 ph onwards the signal was mainly detected in the outer region, drawing a cortex–medulla demarcation. In 1-year-old fish, CD8a+ and TCRb+ thymocytes almost filled the cortex and extended in large cords in the medulla. A CD8a TCRb+ subcapsular lymphoid zone was evident near the septa coming from the inner connective capsule that delimited the thymus. The localisation of CD8a and TCRb transcripts demonstrated a compartmentalisation of the juvenile thymus due to distinct localisation of thymocytes at different developmental stages. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/

Intestinal T cells of Dicentrarchus labrax (L.): gene expression and functional studies

2011-08-04T23:05:11Z

Title: Intestinal T cells of Dicentrarchus labrax (L.): gene expression and functional studies Authors: Picchietti, Simona; Guerra, Laura; Bertoni, Fabrizio; Randelli, Elisa; Belardinelli, Maria Cristina; Buonocore, Francesco; Rombout, Jan; Scapigliati, Giuseppe; Abelli, Luigi Abstract: Cellular and molecular data have evidenced a gut-associated lymphoid tissue in a variety of teleost species, abundantly containing T cells, whose origin, selection and functions are still unclear. This study reports CD4, CD8-α, MHCI-α, MHCII-β, rag-1 and TCR-β gene transcription along the intestine (anterior, middle and posterior segments) and in the thymus of one year-old Dicentrarchus labrax (L.). Real-time PCR findings depicted a main role of the thymus in T cell development, but also rag-1 and CD8-α transcripts are detected in the intestine, having significant expression in the posterior segment. In the whole intestine TCR-β and CD8-α exceeded CD4 transcripts. RNA ISH confirmed these data and detailed that mucosal CD8-α+ cells were especially numerous in the epithelium and in aggregates in the lamina propria. Regional differences in T cell-specific gene expressions are first described in the intestine of a bony fish. High non-specific cytotoxic activity against xenogeneic and allogeneic cells was found in lymphocytes purified from the intestinal mucosa, providing further insight into their local defence roles. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/