Virus-mediated delivery of the green fluorescent protein to the endoplasmic reticulum of plant cells

Abstract

The green fluorescent protein (GFP) was targeted to the endoplasmic reticulum (ER) of living plant cells using a virus-based expression system. The signal peptide from the storage protein, patatin was fused to the amino-terminus of the GFP while the ER retention signal KDEL was fused to the carboxy-terminus. The chimaeric gfp cDNA was inserted into a potato virus X-based expression vector and in-vitro transcripts, representing the recombinant viral genome, were inoculated on to Nicotiana benthamiana and N. clevelandii plants. In virus-infected cells, the GFP was targeted both to the cortical ER and to a mobile system of ER elements which underwent streaming in the cytoplasm. In addition, a population of GFP-containing inclusions was apparent. These inclusions were motile but remained closely associated with elements of the ER. Staining of the ER with membrane potential-sensitive dyes confirmed that the GFP had been targeted to the ER. The utility of virus-mediated delivery systems in studies of the plant endomembrane system is discussed.