Disruption of the methionine cycle and reduced cellular gluthathione levels underlie potex–potyvirus synergism in Nicotiana benthamiana

Summary Infection caused by the synergistic interaction of two plant viruses is typically manifested by severe symptoms and increased accumulation of either virus. In potex–potyviral synergism, the potyviral RNA silencing suppressor helper component proteinase (HCPro) is known to enhance the pathogenicity of the potexvirus counterpart. In line with this, Potato virus X (PVX; genus Potexvirus ) genomic RNA (gRNA) accumulation and gene expression from subgenomic RNA (sgRNA) are increased in Nicotiana benthamiana by Potato virus A (PVA; genus Potyvirus ) HCPro expression. Recently, we have demonstrated that PVA HCPro interferes with the host cell methionine cycle by interacting with its key enzymes S ‐adenosyl‐ l ‐methionine synthetase (SAMS) and S ‐adenosyl‐ l ‐homocysteine hydrolase (SAHH). To study the involvement of methionine cycle enzymes in PVX infection, we knocked down SAMS and SAHH. Increased PVX sgRNA expression between 3 and 9 days post‐infiltration (dpi) and upregulation of (–)‐strand gRNA accumulation at 9 dpi were observed in the SAHH‐silenced background. We found that SAMS and SAHH silencing also caused a significant reduction in glutathione (GSH) concentration, specifically in PVX‐infected plants between 2 and 9 dpi. Interestingly, HCPro expression in PVX‐infected plants caused an even stronger reduction in GSH levels than did SAMS + SAHH silencing and a similar level of reduction was also achieved by knocking down GSH synthetase. PVX sgRNA expression was increased in the GSH synthetase‐silenced background. GSH is a major antioxidant of plant cells and therefore GSH shortage may explain the strong oxidative stress and severe symptoms observed during potex–potyvirus mixed infection.