title: Expression of tuberculosis antigen ESAT-6 in Nicotiana tabacum using a potato virus X-based vector.
abstract: A good candidate antigen to create a therapeutic vaccine against TB is the ESAT-6 protein. Antigens produced in plants have already been successfully used as experimental vaccines, and small single-stranded RNA plant viruses have emerged as promising tools to rapidly express large amounts of foreign proteins in susceptible host plants. Here, we present the expression of ESAT-6 protein in Nicotiana tabacum using a vector based on potato virus X (PVX). The complete ESAT-6 open reading frame is expressed as a fusion protein with the 2A peptide of Foot and Mouth Disease Virus and the amino terminal of the PVX coat protein (CP) (PVXESAT-6). This strategy allows the production of free CP and ESAT-6 as well as fused ESAT-2A-CP to obtain recombinant chimaeric virions expressing ESAT-6 at the surface to be used as particulate antigen in vaccination. ESAT-6 expression was tested in agroinfiltrated tobacco leaves and products of the expected molecular masses corresponding to cleaved CP and ESAT-2A-CP fusion protein were observed, with ESAT-6 yields ranging from 0.5% to 1% of total soluble protein. Our study describes for the first time the expression of the ESAT-6 protein in tobacco plants using a PVX-derived vector. This strategy should serve as a convenient, rapid, low-cost expression system and can also be used for the assessment of ESAT-6 production and function prior to stable plant transformation.
authors.affiliation: Instituto de Investigaciones en Ingeniería Genética y Biología Molecular, CONICET and FCEN-UBA Vuelta de Obligado 2490, Buenos Aires, Argentina. azelada@dna.uba.ar
authors.forename: Alicia M
authors.initials: AM
authors.lastname: Zelada
authors.forename: Gabriela
authors.initials: G
authors.lastname: Calamante
authors.forename: María
authors.initials: M
authors.lastname: de la Paz Santangelo
authors.forename: Fabiana
authors.initials: F
authors.lastname: Bigi
authors.forename: Florencia
authors.initials: F
authors.lastname: Verna
authors.forename: Alejandro
authors.initials: A
authors.lastname: Mentaberry
authors.forename: Angel
authors.initials: A
authors.lastname: Cataldi
mesh_terms: D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D005822:Genetic Vectors; D009169:Mycobacterium tuberculosis / Q000276:immunology*; D018515:Plant Leaves / Q000378:metabolism; D017863:Potexvirus / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D014026:Nicotiana / Q000378:metabolism*
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D011993:Recombinant Fusion Proteins