https://doi.org/10.1111/pbi.70089，Plant Biotechnology Journal,2025

Plants have emerged as powerful biofactories for producing proteins of interest. Compared to mammalian-based expression system, plant biofactories offer unique advantages in terms of higher scalability, lower production costs, reduced risk of contamination by human pathogens and faster expression rate (Chung  et al ., 2022; Eidenberger  et al ., 2023). Over the past three decades, plant-based expression platforms have been successfully used to generate a wide array of recombinant products, including vaccine antigens, therapeutic antibodies and bioactive proteins. The recent pandemic of Ebola and SARS-CoV-2 has further demonstrated the potential of plant biofactories as an alternative protein production system, enabling rapid responses to global health crises and providing high-quality products (Capell  et al ., 2020; Zeitlin  et al ., 2011).

A critical consideration for plant biofactories is enhancing the yield of the target protein. While transgenic systems are useful for large-scale production of single vaccines or therapeutics, they require a time-consuming process and may suffer from low yields due to RNA silencing. Alternatively, engineered plant viral vectors have been developed to efficiently deliver genes of interest into plants, facilitating rapid expression of vaccines, monoclonal antibodies or other therapeutic proteins (Abrahamian  et al ., 2020). However, plant antiviral responses, which manifest at the DNA, RNA and protein levels can significantly impact the yield of target proteins. It is speculated that modulating antiviral host factors could enhance plant viral vector infection and, consequently, improve overall protein expression efficiently. Previously, key components of the conserved antiviral RNA silencing pathway were attractive targets that had been knocked out to improve the productivity of transient expression. However, CRISPR-based knockout of  RDR6  in  Nicotiana benthamiana  resulted in abnormal and sterile flowers similar to  Arabidopsis  ∆ rdr6  mutant, limiting the utility of these engineered plants (Matsuo and Atsumi, 2019). Therefore, identification of antiviral host factors whose knockout can facilitate plant viral vector infection without imposing growth penalties would be ideal for improving target protein production.

Cell wall is a rigid outer layer that provides mechanical support and acts as a physical barrier against biotic and abiotic stresses. In this study, we found that the mRNA level of a glycine-rich cell wall structural protein ( NbGRP ) was upregulated by threefold in response to tomato brown rugose fruit virus (ToBRFV) infection (Figure S1a, Table S1). Sequence analysis showed that NbGRP encodes 132 amino acids, including a potential signal peptide (SP) from residues 1 to 25 at the N-terminus and a glycine-rich domain conserved in the class II GRP (Figure S1b). Confocal microscopy revealed that the NbGRP protein fused to yellow fluorescent protein (NbGRP-YFP) displayed a punctate distribution pattern at the cell wall boundaries (Figure 1a), suggesting localization of NbGRP at plasmodesmata (PD). Furthermore, NbGRP-YFP colocalized with the cell wall marker AtPDLP1 tagged with cyan fluorescent protein (AtPDLP1-CFP) at punctate spots along the cell wall (Figure 1b). After plasmolyzing the cells with 10% NaCl, the fluorescence of NbGRP-YFP remained in the cell wall (Figure 1c), confirming that NbGRP is a PD-localized protein.

Plant Biotechnology Journal, IF=10.1

https://onlinelibrary.wiley.com/doi/10.1111/pbi.70089