Combating photooxidative stress in green hairy roots of Daucus carota cultivated under light irradiation

Abstract

The light-dependent generation of active oxygen species, which can disrupt normal metabolic process of plant, is termed as photo-oxidative stress. Plants are equipped with enzymatic and non-enzymatic antioxidative defence system to reduce the effect of such stress. Hairy root culture of Daucus carota when cultivated under continuous illumination (250μmolm−2s−1) turned green. To know the reason behind that and photo-oxidative stress response in green hairy roots, activities of several antioxidant enzymes were measured. When compared with normal hairy roots, green hairy roots showed an enhanced superoxide dismutase (SOD) activity. Treatment with a SOD inhibitor diethyldithiocarbamate led to suppression of SOD activity in a concentration-dependent manner in green hairy roots. Interestingly, SOD-suppressed root showed three-fold enhanced caffeic acid glucoside accumulation in the soluble fraction as compared to untreated ones. While ascorbate peroxidase activity showed marginal increase in green hairy roots, a decrease in the activities of guaiacol peroxidase and catalase were observed. SDS-PAGE of crude protein profile from green hairy roots showed a distinct band, which was absent in normal hairy roots. MALDI-TOF-MS/MS analysis of the extracted protein confirmed it as the large subunit of RuBisCO. RT-PCR based expression analysis of betaine aldehyde dehydrogenase showed enhanced transcript levels in green hairy roots as compared to normal hairy roots, whereas reverse trends were observed with the transcripts accumulation for phenylalanine ammonia-lyase and chalcone synthase. These findings corroborate with the in vitro BADH activities in hairy roots, and thus indicate an important role of this stress enzyme in combating photo-oxidative stress in green hairy roots upon continuous light exposure.