Production of Biopharmaceuticals in Nicotiana benthamiana—Axillary Stem Growth as a Key Determinant of Total Protein Yield

Abstract

Data are scarce about the influence of basic cultural conditions on growth patterns and overall performance of plants used as heterologous production hosts for protein pharmaceuticals. Higher plants are complex organisms with young, mature, and senescing organs that show distinct metabolic backgrounds and differ in their ability to sustain foreign protein expression and accumulation. Here, we used the transient protein expression host Nicotiana benthamiana as a model to map the accumulation profile of influenza virus hemagglutinin H1, a clinically promising vaccine antigen, at the whole plant scale. Greenhouse-grown plants submitted to different light regimes, submitted to apical bud pruning, or treated with the axillary growth-promoting cytokinin 6-benzylaminopurine were vacuum-infiltrated with agrobacteria harboring a DNA sequence for H1 and allowed to express the viral antigen for 7 days in growth chamber under similar environmental conditions. Our data highlight the importance of young leaves on H1 yield per plant, unlike older leaves which account for a significant part of the plant biomass but contribute little to total antigen titer. Our data also highlight the key contribution of axillary stem leaves, which contribute more than 50% of total yield under certain conditions despite representing only one-third of the total biomass. These findings underline the relevance of both considering main stem leaves and axillary stem leaves while modeling heterologous protein production in N. benthamiana. They also demonstrate the potential of exogenously applied growth-promoting hormones to modulate host plant architecture for improvement of protein yields.

Seasonal Weather Changes Affect the Yield and Quality of Recombinant Proteins Produced in Transgenic Tobacco Plants in a Greenhouse Setting

“…These observations were consistent with a previous study which reported higher protein expression in summer batches based on plant maturation up to a certain point (Sack et al, 2015). Others have reported the effects of light regimes (Goulet et al, 2019) and that short-term heat exposure (37°C) can boost transient protein expression (Norkunas et al, 2018).…”

Current updates and research on plant-based vaccines for coronavirus disease 2019

“…The Quebec City-headquartered biopharma company is employing VLPs which are grown in Nicotiana benthamiana , a tobacco related plant species [ 24 25 26 ]. N. benthamiana is commonly used in biopharmaceutical production due to its ability to express gene sequences that are heterologous in nature [ 27 ]. The biotech company is utilizing genetically modified A. tumefaciens which are able to infect plants by integrating their DNA into the plant cells to produce the VLP [ 28 29 ].…”

“…Researchers are confident in utilizing N. benthamiana to produce COVID-19 vaccine, as it also has contributed in other vaccine production previously such as Ebola [ 72 ]. Other than that, N. benthamiana is preferred by the researches for its rapid growth rate and capable of heterologous gene sequence expression, which is some of the few prominent factors to be selected as a bio-factory for vaccine production [ 27 ]. Newcotiana's plan would be involving the introduction of the desired genes into the plants leaves and the sap which will lead the plant to produce the antigens required for the COVID-19 vaccine and by this, the chances of producing huge amounts of protein would be higher in comparison to other methods [ 72 ].…”

Innovation in plant-based transient protein expression for infectious disease prevention and preparedness

“…For example, the selected application of plant hormones has been used to rebalance the proteome from an antibacterial response to favour recombinant protein accumulation [45]. Hormones have also been used to modify plant architecture, increasing biomass and recombinant protein yield [46]. The over expression of recombinant proteins puts enormous stress on the cell and the co-expression of (b) Modulation of the pH in the secretory pathway via the co-expression of the Influenza M2 ion channel.…”