Glutathione Contribution in Interactions between Turnip mosaic virus and Arabidopsis thaliana Mutants Lacking Respiratory Burst Oxidase Homologs D and F

Abstract

Respiratory burst oxidase homologs (Rbohs) play crucial and diverse roles in plant tissue-mediated production of reactive oxygen species during the development, growth, and response of plants to abiotic and biotic stress. Many studies have demonstrated the contribution of RbohD and RbohF in stress signaling in pathogen response differentially modulating the immune response, but the potential role of the Rbohs-mediated response in plant–virus interactions remains unknown. The present study analyzed, for the first time, the metabolism of glutathione in rbohD-, rbohF-, and rbohD/F-transposon-knockout mutants in response to Turnip mosaic virus (TuMV) infection. rbohD–TuMV and Col-0–TuMV interactions were characterized by susceptible reaction to TuMV, associated with significant activity of GPXLs (glutathione peroxidase-like enzymes) and induction of lipid peroxidation in comparison to mock-inoculated plants, with reduced total cellular and apoplastic glutathione content observed at 7–14 dpi and dynamic induction of apoplast GSSG (oxidized glutathione) at 1–14 dpi. Systemic virus infection resulted in the induction of AtGSTU1 and AtGSTU24, which was highly correlated with significant downregulation of GSTs (glutathione transferases) and cellular and apoplastic GGT (γ-glutamyl transferase) with GR (glutathione reductase) activities. On the contrary, resistant rbohF–TuMV reactions, and especially enhanced rbohD/F–TuMV reactions, were characterized by a highly dynamic increase in total cellular and apoplastic glutathione content, with induction of relative expression of AtGGT1, AtGSTU13, and AtGSTU19 genes. Moreover, virus limitation was highly correlated with the upregulation of GSTs, as well as cellular and apoplastic GGT with GR activities. These findings clearly indicate that glutathione can act as a key signaling factor in not only susceptible rbohD reaction but also the resistance reaction presented by rbohF and rbohD/F mutants during TuMV interaction. Furthermore, by actively reducing the pool of glutathione in the apoplast, GGT and GR enzymes acted as a cell first line in the Arabidopsis–TuMV pathosystem response, protecting the cell from oxidative stress in resistant interactions. These dynamically changed signal transductions involved symplast and apoplast in mediated response to TuMV.

Glutathione-the “master” antioxidant in the regulation of resistant and susceptible host-plant virus-interaction

“…This increase correlated with a significant reduction in the amount and expression of PVY NTN and the induction of the HR response. A similar pattern of stable increase in levels of GSH and GSSG was observed in Arabidopsis mutant plants with increased resistance ( rbohF ) and HR-like reaction ( rbohD/F ) infected by TuMV ( Otulak-Kozieł et al., 2023 ), which was also associated with decreased expression of the virus. Singh et al.…”

“…(2022a) and Otulak-Kozieł et al. (2022b ; 2023) detected modulation of glutathione levels during investigations of infections caused by PVY NTN and TuMV on susceptible and resistant (with hypersensitive response or hypersensitive-like, HR or HR-like) respectively on potato and rbohF and rbohD/f mutants of Arabidopsis ( Figure 2 ). Resistant potato plants infected by PVY NTN exhibited a dynamic increase in the content of glutathione during both resistance and, to some extent, susceptible reactions.…”

“…Therefore, maintaining a precisely controlled level of ROS is crucial, as overproduction could disrupt plant defense/tolerance responses and lead to direct and serious damage to cells. Elements crucial for ROS control, cell protection, and known antiviral responses include the glutathione cycle, especially the levels of GSH and GSSG forms, along with glutathione-associated enzymes such as glutathione transferases (GSTs), glutathione reductases (GRs), and glutathione peroxidases (GPXs).The involvement of glutathione or glutathione treatment in resistance reactions to plant viruses has been reported for various virus types, including potato virus Y-PVY ( Otulak-Kozieł et al., 2022a ) on different cultivar of potato, tobacco mosaic virus — TMV on tobacco ( Gullner et al., 1999 ; Király et al., 2012 ; Künstler et al., 2019a , b ; Zhu et al., 2021 ), turnip mosaic virus-TuMV ( Otulak-Kozieł et al., 2022b , 2023 ) on Arabidopsis and obuda pepper virus - pepper interaction ( Kalapos et al., 2021 ). Generally, it is suggested that elevated glutathione or its external supplementation improves resistance or tolerance against plant viruses.…”

“…The GRs (EC 1.8.1.7) are integral to plant antioxidant defense systems against pathogens, participating in both enzymatic and nonenzymatic oxidation-reduction processes of the cell ( Clarke et al, 2022 ). GRs depend on NADPH levels to transform GSSG to GSH, and through it, they maintain a high ratio of GSH/GSSG in the cell and contribute to the response against plant viruses ( Otulak-Kozieł et al., 2023 ). The plant glutathione peroxidase-like enzymes (GPXL) family consists of multiple isoenzymes with distinct subcellular locations, exhibiting different tissue-specific expression patterns and involvement in various types of stress.…”

Role of NPR1 in Systemic Acquired Stomatal Immunity

“…It is reported that calcium-dependent protein kinase CPK5 contributes to priming, which could enhance basal resistance without the enhancement of SAR in an Arabidopsis sard1-1 mutant [29]. CPK5 is involved in RBOHD-mediated ROS signaling [30], and glutathione can act as a key signaling factor through redox homeostasis [31].…”