title: Production of hybrid-IgG/IgA plantibodies with neutralizing activity against Shiga toxin 1.
abstract: Shiga toxin 1 (Stx1) is a virulence factor of enterohemorrhagic Escherichia coli, such as the O157:H7 strain. In the intestines, secretory IgA (SIgA) is a major component of the immune defense against pathogens and toxins. To form SIgA, the production of dimeric IgA that retains biological activity is an important step. We previously established hybrid-IgG/IgA having variable regions of the IgG specific for the binding subunit of Stx1 (Stx1B) and the heavy chain constant region of IgA. If hybrid-IgG/IgA cDNAs can be expressed in plants, therapeutic or preventive effects may be expected in people eating those plants containing a "plantibody". Here, we established transgenic Arabidopsis thaliana expressing dimeric hybrid-IgG/IgA. The heavy and light chain genes were placed under the control of a bidirectional promoter and terminator of the chlorophyll a/b-binding protein of Arabidopsis thaliana (expression cassette). This expression cassette and the J chain gene were subcloned into a single binary vector, which was then introduced into A. thaliana by means of the Agrobacterium method. Expression and assembly of the dimeric hybrid-IgG/IgA in plants were revealed by ELISA and immunoblotting. The hybrid-IgG/IgA bound to Stx1B and inhibited Stx1B binding to Gb3, as demonstrated by ELISA. When Stx1 holotoxin was pre-treated with the resulting plantibody, the cytotoxicity of Stx1 was inhibited. The toxin neutralization was also demonstrated by means of several assays including Stx1-induced phosphatidylserine translocation on the plasma membrane, caspase-3 activation and 180 base-pair DNA ladder formation due to inter-nucleosomal cleavage. These results indicate that edible plants containing hybrid-IgG/IgA against Stx1B have the potential to be used for immunotherapy against Stx1-caused food poisoning.
authors: Laboratory of Microbiology and Immunology, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka City, Shizuoka, Japan.
authors: Katsuhiro
authors: K
authors: Nakanishi
authors: Sanshiro
authors: S
authors: Narimatsu
authors: Shiori
authors: S
authors: Ichikawa
authors: Yuki
authors: Y
authors: Tobisawa
authors: Kohta
authors: K
authors: Kurohane
authors: Yasuo
authors: Y
authors: Niwa
authors: Hirokazu
authors: H
authors: Kobayashi
authors: Yasuyuki
authors: Y
authors: Imai
mesh_terms: D019138:Antibodies, Blocking / Q000235:genetics / Q000276:immunology; D015295:Antigens, Tumor-Associated, Carbohydrate / Q000276:immunology / Q000378:metabolism; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D016923:Cell Death / Q000276:immunology; D015870:Gene Expression; D023061:Gene Order; D007070:Immunoglobulin A / Q000235:genetics / Q000276:immunology*; D007074:Immunoglobulin G / Q000235:genetics / Q000276:immunology*; D046549:Plantibodies / Q000235:genetics / Q000276:immunology*; D030821:Plants, Genetically Modified; D011485:Protein Binding / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D022622:Shiga Toxin 1 / Q000276:immunology* / Q000378:metabolism
chemical_list: D019138:Antibodies, Blocking; D015295:Antigens, Tumor-Associated, Carbohydrate; C472766:Gb3 antigen; D007070:Immunoglobulin A; D007074:Immunoglobulin G; D046549:Plantibodies; D011994:Recombinant Proteins; D022622:Shiga Toxin 1