title: Efficient production of chimeric human papillomavirus 16 L1 protein bearing the M2e influenza epitope in Nicotiana benthamiana plants.
abstract: BACKGROUND
  Human papillomavirus 16 (HPV-16) L1 protein has the capacity to self-assemble into capsomers or virus-like particles (VLPs) that are highly immunogenic, allowing their use in vaccine production. Successful expression of HPV-16 L1 protein has been reported in plants, and plant-produced VLPs have been shown to be immunogenic after administration to animals.
  
  
  RESULTS
  We investigated the potential of HPV-16 L1 to act as a carrier of two foreign epitopes from Influenza A virus: (i) M2e2-24, ectodomain of the M2 protein (M2e), that is highly conserved among all influenza A isolates, or (ii) M2e2-9, a shorter version of M2e containing the N-terminal highly conserved epitope, that is common for both M1 and M2 influenza proteins. A synthetic HPV-16 L1 gene optimized with human codon usage was used as a backbone gene to design four chimeric sequences containing either the M2e2-24 or the M2e2-9 epitope in two predicted surface-exposed L1 positions. All chimeric constructs were transiently expressed in plants using the Cowpea mosaic virus-derived expression vector, pEAQ-HT. Chimeras were recognized by a panel of linear and conformation-specific anti HPV-16 L1 MAbs, and two of them also reacted with the anti-influenza MAb. Electron microscopy showed that chimeric proteins made in plants spontaneously assembled in higher order structures, such as VLPs of T = 1 or T = 7 symmetry, or capsomers.
  
  
  CONCLUSIONS
  In this study, we report for the first time the transient expression and the self-assembly of a chimeric HPV-16 L1 bearing the M2e influenza epitope in plants, representing also the first record of a successful expression of chimeric HPV-16 L1 carrying an epitope of a heterologous virus in plants. This study further confirms the usefulness of human papillomavirus particles as carriers of exogenous epitopes and their potential relevance for the production in plants of monovalent or multivalent vaccines.
authors.affiliation: Istituto di Virologia Vegetale, CNR, Strada delle Cacce 73, 10135 Torino, Italy.
authors.forename: Slavica
authors.initials: S
authors.lastname: Matić
authors.forename: Riccardo
authors.initials: R
authors.lastname: Rinaldi
authors.forename: Vera
authors.initials: V
authors.lastname: Masenga
authors.forename: Emanuela
authors.initials: E
authors.lastname: Noris
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D036022:Capsid Proteins / Q000096:biosynthesis* / Q000378:metabolism; D003001:Cloning, Molecular; D017798:Comovirus; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000235:genetics*; D005822:Genetic Vectors; D052162:Human papillomavirus 16 / Q000378:metabolism*; D006801:Humans; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009856:Oncogene Proteins, Viral / Q000096:biosynthesis* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D016415:Sequence Alignment; D014026:Nicotiana; D014763:Viral Matrix Proteins / Q000378:metabolism*; D000086082:Viroporin Proteins
chemical_list: D000911:Antibodies, Monoclonal; D036022:Capsid Proteins; D000939:Epitopes; D009856:Oncogene Proteins, Viral; D011993:Recombinant Fusion Proteins; D014763:Viral Matrix Proteins; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins; C063317:L1 protein, Human papillomavirus type 16
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