title: Optimization of codon usage is required for effective genetic immunization against Art v 1, the major allergen of mugwort pollen.
abstract: BACKGROUND
  As the major allergen of mugwort pollen, Art v 1 is an important target for specific immunotherapy. However, both recombinant protein as well as a gene vaccine for Art v 1 failed to be immunogenic in mice. In order to improve immunogenicity we focused on genetic immunization because interspecific differences of codon usage have been shown as an obstacle for effective induction of immune responses with gene vaccines encoding infectious pathogens.
  
  
  OBJECTIVE
  In order to find out, whether codon usage might also be used to improve genetic immunization with allergen genes, the response against a gene vaccine expressing the wild-type gene of Art v 1 (pCMV-wtArt) was compared with a synthetic codon-optimized vector with human codon usage (pCMV-humArt).
  
  
  METHODS
  Balb/c mice were injected intradermally with pCMV-wtArt or pCMV-humArt. In vitro expression levels of both constructs were compared in transfection experiments. Total immunoglobulin G (IgG), IgG1, IgG2a and IgE antibodies were analyzed by enzyme-linked immunosorbent assay and the anaphylactic activity of the sera was determined by allergen-specific degranulation of rat basophil leukemia-2H3 cells.
  
  
  RESULTS
  No immune response was detectable with the gene vaccine expressing the wildtype Art v 1, but immunization with pCMV-humArt revealed a strong and allergen-specific induction of antibody responses. The antibodies recognized both the recombinant as well as the purified natural (glycosylated) Art v 1 molecule. The response type was Th1-biased, as indicated by high levels of IgG2a antibodies. Expression analysis with B16 mouse melanoma cells transfected with pCMV-humArt or pCMV-wtArt revealed an impaired expression of the wild-type vector but normal translation after recoding.
  
  
  CONCLUSIONS
  The results demonstrate that optimization of codon usage offers a simple way to improve immunogenicity and therefore should be routinely considered in the development of gene vaccines for the treatment of allergy.
mesh_terms: D000485:Allergens / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D000595:Amino Acid Sequence; D000818:Animals; D052179:Antigens, Plant; D001483:Base Sequence; D045744:Cell Line, Tumor; D003062:Codon; D005260:Female; D005822:Genetic Vectors; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D051381:Rats; D016415:Sequence Alignment; D013045:Species Specificity; D019444:Vaccines, DNA / Q000276:immunology*
chemical_list: D000485:Allergens; D052179:Antigens, Plant; C102724:Art v 1 protein, mugwort; D003062:Codon; D007074:Immunoglobulin G; D010940:Plant Proteins; D019444:Vaccines, DNA