Rapid transient protein production by the coat protein-deficient cucumber mosaic virus vector: non-packaged CMV system, NoPaCS.

KEY MESSAGE: We developed a non-packaged CMV system (NoPaCS) for CMV-agroinfection with a virus-inescapable transgenic plant platform, enabling rapid, high production of a large-sequence target protein. For rapidly producing high levels of a desirable protein, many plant virus vectors have been developed. However, there is always a concern that such recombinant viruses may escape into the environment. Especially for insect-transmissible viruses, certain measures must be taken. We here developed a new cucumber mosaic virus (CMV) RNA 3-based vector that is not transmitted by aphids because we deleted the coat protein (CP) gene responsible for aphid transmission and replaced it with a foreign gene. Transgenic Nicotiana benthamiana plants expressing CMV RNA 1 (CR1Tg) were found to be the most suitable platform for producing a recombinant protein using the CMV vector. By agroinfiltrating CR1Tg plants with the RNA 2 construct and the CMV vector harboring the green fluorescence protein (GFP) gene instead of the CP gene, we achieved a high yield of GFP (e.g., ~ 750 mg/kg FW) throughout the bacteria-infiltrated tissues at 2-3 days after infiltration. Furthermore, with this CMV-agroinfection system, a large gene such as the β-glucuronidase (GUS) gene can be expressed because the viral RNAs are not necessarily encapsidated for replication. The system is designated "non-packaged CMV system (NoPaCS)".