title: Translational fusion of chloroplast-expressed human papillomavirus type 16 L1 capsid protein enhances antigen accumulation in transplastomic tobacco.
abstract: Human Papillomavirus (HPV) is the causal agent of cervical cancer, one of the most common causes of death for women. The major capsid L1 protein self-assembles in Virus Like Particles (VLPs), which are highly immunogenic and suitable for vaccine production. In this study, a plastid transformation approach was assessed in order to produce a plant-based HPV-16 L1 vaccine. Transplastomic plants were obtained after transformation with vectors carrying a chimeric gene encoding the L1 protein either as the native viral (L1(v) gene) or a synthetic sequence optimized for expression in plant plastids (L1(pt) gene) under control of plastid expression signals. The L1 mRNA was detected in plastids and the L1 antigen accumulated up to 1.5% total leaf proteins only when vectors included the 5'-UTR and a short N-terminal coding segment (Downstream Box) of a plastid gene. The half-life of the engineered L1 protein, determined by pulse-chase experiments, is at least 8 h. Formation of immunogenic VLPs in chloroplasts was confirmed by capture ELISA assay using antibodies recognizing conformational epitopes and by electron microscopy.
authors.affiliation: CNR-IGV, Institute of Plant Genetics-Research Division Portici, via Università 133, 80055 Portici, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Lenzi
authors.forename: Nunzia
authors.initials: N
authors.lastname: Scotti
authors.forename: Fiammetta
authors.initials: F
authors.lastname: Alagna
authors.forename: Maria L
authors.initials: ML
authors.lastname: Tornesello
authors.forename: Andrea
authors.initials: A
authors.lastname: Pompa
authors.forename: Alessandro
authors.initials: A
authors.lastname: Vitale
authors.forename: Angelo
authors.initials: A
authors.lastname: De Stradis
authors.forename: Luigi
authors.initials: L
authors.lastname: Monti
authors.forename: Stefania
authors.initials: S
authors.lastname: Grillo
authors.forename: Franco M
authors.initials: FM
authors.lastname: Buonaguro
authors.forename: Pal
authors.initials: P
authors.lastname: Maliga
authors.forename: Teodoro
authors.initials: T
authors.lastname: Cardi
mesh_terms: D001669:Biochemical Phenomena; D002213:Capsid / Q000276:immunology / Q000378:metabolism; D036022:Capsid Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology* / Q000648:ultrastructure; D002736:Chloroplasts / Q000378:metabolism* / Q000648:ultrastructure; D004797:Enzyme-Linked Immunosorbent Assay; D005240:Feasibility Studies; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D009856:Oncogene Proteins, Viral / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology* / Q000648:ultrastructure; D030821:Plants, Genetically Modified; D010957:Plasmids; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology; D014026:Nicotiana / Q000235:genetics / Q000276:immunology* / Q000378:metabolism / Q000648:ultrastructure; D019076:Transgenes
chemical_list: D036022:Capsid Proteins; D009856:Oncogene Proteins, Viral; D012333:RNA, Messenger; D011994:Recombinant Proteins; C063317:L1 protein, Human papillomavirus type 16