Abstract
The small genomes of plant viruses can be easily engineered to transform these pathogens into powerful biotechnological tools able to induce the production of large amounts of recombinant proteins in the host plant, which is converted in a biofactory. Plant virus biotechnology approaches have traditionally focused on viral species that induce the production of huge amounts of some of their proteins, such us Tobacco mosaic virus (TMV) or Cowpea mosaic virus (CPMV). However, the amazing wealth of genome architectures and expression strategies found in plant viruses open many other possibilities. One of them is the use of potyviruses as viral vectors. Potyviruses (genus Potyvirus) make one of the largest groups of plant viruses with around 200 different species known to date. Their genome is composed of an about 10,000-nt long plus-strand RNA molecule that is bound at the 5¿ end to a genome-linked viral (VPg) protein and is polyadenylated at the 3¿ end. This genomic RNA is encapsidated in elongated and flexuous virions. What makes potyviruses special is their gene expression strategy. Basically, potyviruses express a single polyprotein that is processed by three viral proteases. The highly specific NIa protease (NIaPro) performs most cleavages in the viral polyprotein based on a seven amino acid recognition motif. We set up a potyvirus-derived vector system in which the viral cistron encoding the RNA-dependent RNA polymerase (NIb) is replaced by a cassette to produce several proteins. The cDNAs encoding for these proteins are flanked with sequences corresponding to the NIaPro cleavage site. This way, the viral vector produces equimolar amounts of several proteins in plants that co-express the viral NIb. Insertion of the recombinant cDNA at the most amino terminal end of the viral polyprotein has allowed us to target heterologous proteins to specific organelles, such as chloroplasts and mitochondria. Using these potyvirus vectors we have produced antibodies, transcription factors and metabolic enzymes.
