title: A potyvirus vector efficiently targets recombinant proteins to chloroplasts, mitochondria and nuclei in plant cells when expressed at the amino terminus of the polyprotein.
abstract: Plant virus-based expression systems allow quick and efficient production of recombinant proteins in plant biofactories. Among them, a system derived from tobacco etch virus (TEV; genus potyvirus) permits coexpression of equimolar amounts of several recombinant proteins. This work analyzed how to target recombinant proteins to different subcellular localizations in the plant cell using this system. We constructed TEV clones in which green fluorescent protein (GFP), with a chloroplast transit peptide (cTP), a nuclear localization signal (NLS) or a mitochondrial targeting peptide (mTP) was expressed either as the most amino-terminal product or embedded in the viral polyprotein. Results showed that cTP and mTP mediated efficient translocation of GFP to the corresponding organelle only when present at the amino terminus of the viral polyprotein. In contrast, the NLS worked efficiently at both positions. Viruses expressing GFP in the amino terminus of the viral polyprotein produced milder symptoms. Untagged GFPs and cTP and NLS tagged amino-terminal GFPs accumulated to higher amounts in infected tissues. Finally, viral progeny from clones with internal GFPs maintained the extra gene better. These observations will help in the design of potyvirus-based vectors able to coexpress several proteins while targeting different subcellular localizations, as required in plant metabolic engineering.
authors.affiliation: Instituto de Biología Molecular y Celular de Plantas (Consejo Superior de Investigaciones Científicas-Universidad Politécnica de Valencia), Valencia, Spain.
authors.forename: Eszter
authors.initials: E
authors.lastname: Majer
authors.affiliation: Instituto de Biología Molecular y Celular de Plantas (Consejo Superior de Investigaciones Científicas-Universidad Politécnica de Valencia), Valencia, Spain.
authors.forename: José-Antonio
authors.initials: JA
authors.lastname: Navarro
authors.affiliation: Instituto de Biología Molecular y Celular de Plantas (Consejo Superior de Investigaciones Científicas-Universidad Politécnica de Valencia), Valencia, Spain. jadaros@ibmcp.upv.es.
authors.forename: José-Antonio
authors.initials: JA
authors.lastname: Daròs
mesh_terms: D002467:Cell Nucleus / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D005822:Genetic Vectors / Q000235:genetics*; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000821:virology; D020815:Polyproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017800:Potyvirus / Q000235:genetics*; D021382:Protein Sorting Signals / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics / Q000821:virology
chemical_list: D008164:Luminescent Proteins; D020815:Polyproteins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins
keywords: Potyvirus vector; Recombinant protein; Signal peptides; Subcellular localization; Tobacco etch virus