title: High-level transient expression of ER-targeted human interleukin 6 in Nicotiana benthamiana.
abstract: Tobacco plants can be used to express recombinant proteins that cannot be produced in a soluble and active form using traditional platforms such as Escherichia coli. We therefore expressed the human glycoprotein interleukin 6 (IL6) in two commercial tobacco cultivars (Nicotiana tabacum cv. Virginia and cv. Geudertheimer) as well as the model host N. benthamiana to compare different transformation strategies (stable vs. transient expression) and subcellular targeting (apoplast, endoplasmic reticulum (ER) and vacuole). In T(0) transgenic plants, the highest expression levels were achieved by ER targeting but the overall yields of IL6 were still low in the leaves (0.005% TSP in the ER, 0.0008% in the vacuole and 0.0005% in the apoplast). The apoplast variant accumulated to similar levels in leaves and seeds, whereas the ER-targeted variant was 1.2-fold more abundant in seeds and the vacuolar variant was 6-fold more abundant in seeds. The yields improved in subsequent generations, with the best-performing T(2) plants producing the ER-targeted IL6 at 0.14% TSP in both leaves and seeds. Transient expression of ER-targeted IL6 in leaves using the MagnICON system resulted in yields of up to 7% TSP in N. benthamiana, but only 1% in N. tabacum cv. Virginia and 0.5% in cv. Geudertheimer. Although the commercial tobacco cultivars produced up to threefold more biomass than N. benthamiana, this was not enough to compensate for the lower overall yields. The recombinant IL6 produced by transient and stable expression in plants was biologically active and presented as two alternative bands matching the corresponding native protein.
authors.affiliation: Department of Agrobiotechnology, Agricultural and Environmental Faculty, University of Rostock, Rostock, Germany.
authors.forename: Henrik
authors.initials: H
authors.lastname: Nausch
authors.forename: Heike
authors.initials: H
authors.lastname: Mikschofsky
authors.forename: Roswitha
authors.initials: R
authors.lastname: Koslowski
authors.forename: Udo
authors.initials: U
authors.lastname: Meyer
authors.forename: Inge
authors.initials: I
authors.lastname: Broer
authors.forename: Jana
authors.initials: J
authors.lastname: Huckauf
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D018628:Gene Dosage; D015870:Gene Expression; D023061:Gene Order; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics* / Q000378:metabolism*; D042541:Intracellular Space / Q000378:metabolism; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified; D021381:Protein Transport; D012639:Seeds / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism*; D014170:Transformation, Genetic; D019076:Transgenes
chemical_list: D015850:Interleukin-6