• Open Access

Plant recombinant erythropoietin attenuates inflammatory kidney cell injury

Authors

  • Andrew J. Conley,

    1. Department of Biology, University of Western Ontario, London, ON, Canada, N6A 5B7
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  • Kanishka Mohib,

    1. Departments of Medicine and Microbiology and Immunology, University of Western Ontario, London, ON, Canada, N6A 5A5
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  • Anthony M. Jevnikar,

    Corresponding author
    1. Departments of Medicine and Microbiology and Immunology, University of Western Ontario, London, ON, Canada, N6A 5A5
    2. Transplantation Immunology Group, Lawson Health Research Institute, London, ON, Canada, N6A 5A5
      * Correspondence (fax (519)663-8808; e-mail: jevnikar@uwo.ca)
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  • Jim E. Brandle

    1. Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, London, ON, Canada, N5V 4T3
    2. Vineland Research and Innovation Centre, Vineland Station, ON, Canada, L0R 2E0
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* Correspondence (fax (519)663-8808; e-mail: jevnikar@uwo.ca)

Summary

Human erythropoietin (EPO) is a pleiotropic cytokine with remarkable tissue-protective activities in addition to its well-established role in red blood cell production. Unfortunately, conventional mammalian cell cultures are unlikely to meet the anticipated market demands for recombinant EPO because of limited capacity and high production costs. Plant expression systems may address these limitations to enable practical, cost-effective delivery of EPO in tissue injury prevention therapeutics. In this study, we produced human EPO in tobacco and demonstrated that plant-derived EPO had tissue-protective activity. Our results indicated that targeting to the endoplasmic reticulum (ER) provided the highest accumulation levels of EPO, with a yield approaching 0.05% of total soluble protein in tobacco leaves. The codon optimization of the human EPO gene for plant expression had no clear advantage; furthermore, the human EPO signal peptide performed better than a tobacco signal peptide. In addition, we found that glycosylation was essential for the stability of plant recombinant EPO, whereas the presence of an elastin-like polypeptide fusion had a limited positive impact on the level of EPO accumulation. Confocal microscopy showed that apoplast and ER-targeted EPO were correctly localized, and N-glycan analysis demonstrated that complex plant glycans existed on apoplast-targeted EPO, but not on ER-targeted EPO. Importantly, plant-derived EPO had enhanced receptor-binding affinity and was able to protect kidney epithelial cells from cytokine-induced death in vitro. These findings demonstrate that tobacco plants may be an attractive alternative for the production of large amounts of biologically active EPO.

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