Enhanced Synthesis of Foreign Nuclear Protein Stimulates Viral Reproduction via the Induction of γ-Thionin Expression

Plants are a promising platform for recombinant protein production. Here we propose a novel approach to increase the level of viral vector-mediated recombinant protein synthesis. This approach is based on the hypothesis that antiviral protection is weakened during the antibacterial cellular response. We suggested that introduced to the cell foreign nuclear localized proteins, including effectors such as bacterial nucleomodulins, can interfere with the import of cellular nuclear proteins and launch antibacterial defense reactions, creating favorable conditions for cytoplasmic virus reproduction. Here, we performed synthesis of an artificial nuclear protein-red fluorescent protein (mRFP) fused with a nuclear localization sequence (NLS)-in plant cells as a mimetic of a bacterial effector. Superproduction of mRFP:NLS induced Nicotiana benthamiana gamma-thionin (Nb gamma Thio) mRNA accumulation. Both NLS-containing protein synthesis and increased Nbr gamma Thio expression stimulated reproduction of the viral vector based on the genome of crucifer-infecting tobacco mosaic virus (crTMV) in N. benthamiana leaves. We isolated the Nbr gamma Thio gene promoter (Pr-gamma Thio) and showed that Pr-gamma Thio activity sharply increased in response to massive synthesis of GFP fused with NLS. We conclude that NLS-induced Pr-gamma Thio activation and increased accumulation of Nbr gamma thio mRNA led to the stimulation of GFP expression from crTMV: GFP vector in the transient expression system.

Automated summary

Plants can be used as a promising platform for producing recombinant proteins. This study proposes a novel method to enhance the synthesis of recombinant proteins using viral vectors. The approach suggests that introducing foreign nuclear proteins, including bacterial effectors, can interfere with the import of cellular nuclear proteins and trigger antibacterial defense reactions, thereby creating favorable conditions for cytoplasmic virus reproduction. In this study, an artificial nuclear protein mRFP:NLS was synthesized in plant cells, mimicking a bacterial effector. The synthesis of mRFP:NLS resulted in the accumulation of Nb gamma Thio mRNA and stimulated the reproduction of a viral vector based on the genome of crucifer-infecting tobacco mosaic virus (crTMV) in plant leaves. The study also identified a gene promoter, Pr-gamma Thio, which showed increased activity in response to the increased synthesis of GFP fused with NLS.