Recombinant mouse granulocyte-macrophage colony-stimulating factor is glycosylated in transgenic tobacco and maintains its biological activity

Abstract

The granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine with many important applications and, due to its immunostimulatory properties, could also be used as a vaccine adjuvant. A simple strategy to produce recombinant mouse GM-CSF (mGM-CSF) in transgenic Nicotiana tabacum plants was used in this study. The mGM-CSF cDNA followed by the sequence encoding endoplasmic reticulum retention signal (KDEL) was cloned into the ImpactVector under the control of the strong promoter from the gene encoding a small subunit of Rubisco. In transgenic plants the accumulation level of recombinant mGM-CSF varied in the individual transformants from 8 to 19 microg/g of fresh leaf tissue, which makes up to 0.22% of total soluble protein. In most analyzed plants, the apparent molecular weight of the recombinant protein was larger than predicted due to its N-glycosylation, presumably in 2 sites. The recombinant plant-produced murine GM-CSF retained its biological activity as confirmed in vitro in proliferation assay using a mouse cell line, which is growth-dependent on GM-CSF.