Abstract
Main conclusion
An edible plant was tested as a host for the production of secretory monoclonal IgA against Shiga toxin 1 (Stx1). The lettuce-derived IgA completely protected Vero cells from Stx1.
Abstract
Secretory immunoglobulin A (SIgA) is thought to control mucosal infections and thus it may be applicable to oral passive immunotherapy. Edible plants are candidate hosts for producing oral formulations with SIgA against pathogenic agents. We previously established a recombinant IgA specific for the B subunit of Shiga toxin 1 (Stx1B) consisting of the Fab fragment of Stx1B-specific monoclonal IgG and the Fc region of IgA (hyIgA). Here, we developed transgenic lettuce (Lactuca sativa) that produces hyIgA in a secretory form (S-hyIgA). An Arabidopsis-derived light-harvesting complex II (LHCB) promoter was used for the expression of all four transgenes (hyIgA heavy, light and j chains, and secretory component). Agrobacterium-mediated transformation was carried out to introduce genes into lettuce leaf discs by means of a single vector harboring all four transgenes. Consistent with the tissue specificity of the LHCB promoter, the expression of hyIgA transgenes was observed in leaf and stem tissues, which contain chloroplasts, at the mRNA and protein levels. The leaves produced hyIgA in a more than tenfold higher yield as compared with stems. The lettuce-derived S-hyIgA was found to bind to Stx1B in a dose-dependent manner by means of ELISA. A leaf extract of the transgenic lettuce completely neutralized the cytotoxicity of Stx1 against Vero cells, which are highly susceptible to Stx1. In conclusion, we established a transgenic lettuce producing a secretory form of hyIgA that can bind bacterial toxin. The results indicate that edible practical plants containing S-hyIgA will provide a possible means for immunotherapy for food poisoning.
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Abbreviations
- AmS:
-
Ammonium sulfate
- DIG:
-
Digoxigenin
- EHEC:
-
Enterohemorrhagic Escherichia coli
- FBS:
-
Fetal bovine serum
- FW:
-
Fresh weight
- Gb3 :
-
Globotriaosylceramide
- HRP:
-
Horseradish peroxidase
- Hc:
-
Heavy chain
- hyIgA:
-
Hybrid-IgG/IgA
- Jc:
-
Joining chain
- Lc:
-
Light chain
- LHCB:
-
Light-harvesting complex II
- mAb:
-
Monoclonal antibody
- MS:
-
Murashige and Skoog
- PBS:
-
Phosphate-buffered saline
- pIgR:
-
Polymeric immunoglobulin receptor
- SC:
-
Secretory component
- S-hyIgA:
-
Secretory hybrid-IgG/IgA
- SIgA:
-
Secretory IgA
- Stx1:
-
Shiga toxin 1
- Stx1A:
-
A subunit of Stx1
- Stx1B:
-
B subunit of Stx1
- T-DNA:
-
Transfer-DNA
- TSP:
-
Total soluble protein
- UBQ:
-
Ubiquitin extension protein
- WT:
-
Wild-type
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Acknowledgements
This work was supported by Japan Society for the Promotion of Science (JSPS) KAKENHI Grant Numbers JP15H04660 and JP25670063 to YI; JP25·10915 to KN as well as by a research grant from the University of Shizuoka. We thank Dr. Hiroshi Asao (Nara Prefecture Agricultural Research and Development Center) for guidance for transformation of lettuce; and Mr. N.J. Halewood for language editing services.
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Nakanishi, K., Matsuda, M., Ida, R. et al. Lettuce-derived secretory IgA specifically neutralizes the Shiga toxin 1 activity. Planta 250, 1255–1264 (2019). https://doi.org/10.1007/s00425-019-03215-1
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DOI: https://doi.org/10.1007/s00425-019-03215-1