Coexpression of human perforin improves yeast-mediated delivery of DNA and mRNA to mammalian antigen-presenting cells.
Previous studies underlined the capacity of recombinant yeast as efficient vehicle for the targeted delivery of functional nucleic acids as well as proteinaceous antigens to mammalian antigen-presenting cells (APCs). To improve this yeast-mediated cargo transport into APCs, we investigated the impact of coexpression of the human membrane-perturbing protein perforin in comparison with bacterial listeriolysin O (LLO) on the yeast-based delivery of DNA, mRNA and proteins to mammalian APCs. In contrast to LLO, a cholesterol-dependent pore-forming toxin of Listeria, intracellular expression of human perforin in Saccharomyces cerevisiae had no impact on yeast cell viability, while its coexpression significantly increased translocation of ovalbumin and subsequent activation of ovalbumin-specific T lymphocytes. Likewise, perforin improved the expression of the model antigen enhanced green fluorescent protein after yeast-mediated DNA and mRNA delivery, whereas LLO was only able to enhance DNA delivery. Taken together, our data show that human perforin, besides bacterial hemolysins, represents a promising means to improve the yeast-mediated delivery of functional nucleic acids and proteins to mammalian APCs.
Department of Molecular and Cell Biology, Saarland University, Saarbrücken, Germany.|Medical Faculty, Junior Research Group for Virology and Immunology, Institute of Virology and Center of Human and Molecular Biology, Saarland University, Homburg Saar, Germany.
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Walch-Rückheim
Department of Molecular and Cell Biology, Saarland University, Saarbrücken, Germany.
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R
Kiefer
Faculty for Clinical Medicine Mannheim of the University Heidelberg, Institute for Medical Microbiology and Hygiene, Mannheim, Germany.
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Geginat
Department of Molecular and Cell Biology, Saarland University, Saarbrücken, Germany.
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MJ
Schmitt
Department of Molecular and Cell Biology, Saarland University, Saarbrücken, Germany.
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Breinig
D000818:Animals; D000938:Antigen-Presenting Cells / Q000166:cytology / Q000378:metabolism*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004247:DNA / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D010047:Ovalbumin; D054353:Perforin / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D013601:T-Lymphocytes
D001427:Bacterial Toxins; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; C510468:PRF1 protein, human; D012333:RNA, Messenger; D054353:Perforin; D049452:Green Fluorescent Proteins; D010047:Ovalbumin; D004247:DNA; C038705:hlyA protein, Listeria monocytogenes