Virus Vector-Mediated Rapid Protein Production in Nicotiana benthamiana: Effects of Temperature and Photosynthetic Photon Flux Density on Hemagglutinin Accumulation

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Virus vector-mediated transgene expression enables rapid pharmaceutical protein production using plants, such as hemagglutinin (HA), a vaccine antigen of influenza virus. Here, effects of air temperature and photosynthetic photon flux density (PPFD) from the transgene introduction to harvest on recombinant HA content in Nicotiana benthamiana leaves were investigated. A deconstructed tobamovirus-based vector system (magnICON) was used for transient expression of HA. Plants were incubated at a PPFD of 100 (LP) or 300 (HP) μmol m−2 s−1 and an air temperature of 20 (LT) or 25 (HT) °C for 6 d after transgene introduction. At the harvest, high-level accumulation of intact HA of approximately 75 kDa was detected in leaves. Leaf HA content per unit dry weight in LT was three-times greater than that in HT. There was no significant difference in HA content between LP and HP regardless of temperature. The greater HA content in LT was positively correlated with a greater total soluble protein content, suggesting that not only HA but overall soluble proteins including HA were greater in LT than in HT. Thus, air temperature for plant incubation after transgene introduction can be a crucial environmental factor for HA productivity in virus vector-mediated transient expression systems.