Person: Gil Dones, Félix
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First Name
Félix
Last Name
Gil Dones
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Biológicas
Department
Genética, Fisiología y Microbiología
Area
Genética
Identifiers
4 results
Search Results
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Item Oral immunogenicity of the plant derived spike protein from swine-transmissible gastroenteritis coronavirus(2000) Gómez, Nestor; Wigdorovitz, Andres; Castañón, Sonia; Gil Dones, Félix; Ordá, Ricardo; Borca, Manuel; Escribano, JoséTransgenic plants represent an inexpensive alternative to classical fermentation systems for production of recombinant subunit vaccines. Transgenic potato plants were created that express the N-terminal domain of the glycoprotein S (N-gS) from Transmissible gastroenteritis coronavirus (TGEV), containing the major antigenic sites of the protein. Extracts from potato tubers expressing N-gS were inoculated intraperitoneally to mice, and the vaccinated mice developed serum IgG specific for TGEV. Furthermore, when potato tubers expressing N-gS were fed directly to mice, they developed serum antibodies specific for gS protein, demonstrating the oral immunogenicity of the plant derived spike protein from TGEV.Item A novel methodology to develop a foot and mouth disease virus (FMDV) peptide-based vaccine in transgenic plants(2002) Dus Santos, María ; Wigdorovitz, Andrés; Trono, Karina; Rı́os, Raúl ; Franzone, Pascual ; Gil Dones, Félix; Moreno, Javier; Carrillo, Consuelo; Escribano, José ; Borca, ManuelThe expression of antigens in transgenic plants has been increasingly used as an alternative to the classical methodologies for antigen expression in the development of experimental vaccines. However, an important limitation in most cases is the low concentration of the recombinant antigens in the plant tissues, which reduces the possibilities of practical applications. Because the site of insertion of the transferred DNA into the cellular chromosomal DNA is at random, different levels of foreign protein expression in independent transformants is expected. Strategies to allow the evaluation of a high number of the transgenic individuals, usually an expensive and very time consuming process, would permit the selection of those plants presenting the highest levels of recombinant protein expression. Here, we present the development of an experimental immunogen based in the expression of a highly immunogenic epitope from foot and mouth disease virus (FMDV) fused to the glucuronidase (gus A) reporter gene, which allows selection of the transgenic plants by the ß-glucuronidase (ßGUS) enzymatic activity. We produced transgenic plants of alfalfa expressing the immunogenic site between amino acid residues 135–160 of structural protein VP1 (VP135–160), fused to the ßGUS protein. Plants expressing the highest levels of the immunogenic epitope VP135–160, analyzed by Western blot, were efficiently selected based on their levels of ßGUS enzymatic activity. The FMDV epitope expressed in plants was highly immunogenic in mice which developed, after immunization, a strong anti-FMDV antibody response against a synthetic peptide representing the region VP135–160, to native virus VP1, and to purified FMDV particles. Additionally, these mice were completely protected against experimental challenge with the virulent virus. To our knowledge, this constitutes the first report of a peptide-based vaccine produced in transgenic plants that induces a protective immune response when used in experimental hosts. Also, these results demonstrated the possibility of using a novel and simple methodology for obtaining transgenic plants expressing high levels of foreign immunogenic epitopes, which could be directly applied in the development of plant-based vaccines.Item High‐yield expression of a viral peptide vaccine in transgenic plants(FEBS Letters, 2001) Gil Dones, Félix; Brun, Alejandro; Wigdorovitz, Andrés; Catalá, Rafael; Martı́nez-Torrecuadrada, Jorge ; Casal, Ignacio; Salinas, Julio; Borca, Manuel ; Escribano, JoséA high-yield production of a peptide vaccine in transgenic plants is described here. A 21-mer peptide, which confers protection to dogs against challenge with virulent canine parvovirus, has been expressed in transgenic plants as an amino-terminal translational fusion with the GUS gene. Transformants were selected on the basis of their GUS activities, showing expression levels of the recombinant protein up to 3% of the total leaf soluble protein, a production yield comparable to that obtained with the same epitope expressed by chimeric plant viruses. The immunogenicity of the plant-derived peptide was demonstrated in mice immunized either intraperitoneally or orally with transgenic plant extracts, providing the suitability of the GUS fusions approach for low-cost production of peptide vaccines.Item Multimerization of peptide antigens for production of stable immunogens in transgenic plants(Journal of Biotechnology, 2007) Gil Dones, Félix; Reytor, Edel; Pérez-Filgueira, Daniel Mariano; Escribano, JoséPrevious literature addressing the production of recombinant proteins in heterologous systems has consistently shown that proteins capable of forming complex structures tend to accumulate within host cells at relatively higher levels than monomeric forms. In this report, we translationally fused a 21-aminoacids long highly immunogenic peptide (2L21), derived from canine parvovirus (CPV) VP2 protein to a 41-aminoacid long tetramerization domain (TD) from the transcriptional factor p53. The chimerical DNA construction 2L21-TD was cloned in a binary plant transformation vector and used to transform Arabidopsis thaliana plants. Fifteen of the 25 transgenic lines obtained in the experiment showed detectable 2L21-TD RNA accumulation and from these we chose 4 to study 2L21-TD protein accumulation. Non-denaturing immunoblotting assays revealed that 2L21-TD chimeras effectively formed tetrameric complexes with yields reaching up to 12 μg/mg of soluble protein. Mice immunized by oral or intraperitoneal routes with crude protein extracts containing 2L21-TD protein were able to detect both 2L21-synthetic peptide and CPV VP2 proteins, with titers similar to those elicited by a previously reported fusion between 2L21 and the β-glucuronidase protein. These results demonstrate that multimerization directed by the small TD domain contributed to the stabilization and consequently to the accumulation of the 2L21 peptide in transgenic plants, without altering its native antigenicity and immunogenicity.