Elevated thiol levels affect virus infection in Cucurbita pepo plants regenerated via somatic embryogenesis

Thiols are known to play crucial roles in plant defense responses, e.g. against pathogens. In the present study, the efficiency of these molecules was evaluated in regenerated Cucurbita pepo L. subsp. pepo var. styriaca Greb. plants in response to compatible Zucchini yellow mosaic virus infection. Thiol contents were modulated by applying the cysteine precursor L-2-oxothiazolidine-4-carboxylic acid (OTC) to callus cells. Cytological and biochemical methods were used for determination of glutathione within callus cells and leaves of plants regenerated via somatic embryogenesis. Reduced glutathione levels (GSH) significantly increased in callus cells one week after 1.0 mM OTC treatment by about 56% and resulted in enhanced GSH levels throughout the investigation period of 13 weeks. Three weeks after ZYMV infection untreated plants regenerated from callus culture exhibited stunted growth forms, yellowing, mosaic and blistering on the leaves whereas 68% of 1.0 mM OTC pretreated regenerated plants were characterized by a complete absence of symptoms. Treatment with OTC stimulated the synthesis, conversion and translocation of thiols during advanced ZYMV infection. Compared to untreated infected leaves, a graduate increase in cysteine (cys) and γ-glutamyl-cysteine (γ-glucys) contents was measured in 0.5 mM and 1.0 mM OTC treated older leaves, whereas total glutathione (tGSH) decreased significantly after infection. In OTC treated younger leaves cys and tGSH concentrations remained at control levels and were unaffected by ZYMV. In contrast, OTC treatment caused a dramatic induction of γ-glucys, up to 191% in younger infected leaves. The results show that altered levels of thiols by OTC were accompanied by enhanced tolerance against ZYMV, suppression of ZYMV induced symptoms as well as delayed symptom development.