Securing industrial molecular farming: NbXTH30-mediated cell wall fortification enhances insect resistance in the biofactory crop Nicotiana benthamiana

The rapid commercialization of Nicotiana benthamiana as a scalable biofactory for molecular farming necessitates large-scale cultivation, which in turn elevates the agronomic risk of significant yield loss due to pest outbreaks. The plant cell wall is a crucial primary structural barrier against herbivores, yet the molecular mechanisms underlying cell wall-mediated defense, particularly against piercing-sucking insects, remain poorly characterized. In this study, we demonstrate that aphid infestation of N. benthamiana triggers a broad defense response and significant cell wall thickening. Through expression profiling, we identified a Xyloglucan endotransglucosylase/hydrolase gene ( NbXTH30 ) as a key insect-responsive gene, which plays an important role in modifying cell wall architecture. Functional characterization revealed that NbXTH30 enhances plant resistance to aphids by disrupting insect feeding behavior. Further enzymatic analysis confirmed that NbXTH30 functions as an active xyloglucan endotransglucosylase (XET), contributing to cell wall reinforcement. The conservation of this mechanism was underscored by the fact that the tomato ortholog similarly conferred resistance against piercing-sucking insects. Our findings delineate a defense mechanism, from the induction of NbXTH30 to XET-mediated cell wall thickening and ultimately enhanced insect resistance, providing both fundamental insights into plant structural immunity and a promising genetic target for breeding insect-resistant crops. • Aphid infestation induces cell wall fortification and NbXTH30 expression. • NbXTH30 is a functional XET that modifies cell walls and increases wall thickness. • NbXTH30 confers aphid resistance by impeding stylet penetration and feeding. • The tomato XTH30 ortholog similarly restricts aphids via wall thickening. • This defines a conserved structural defense mechanism with breeding potential.