Effect of Rol Genes on Polyphenols Biosynthesis in Artemisia annua and Their Effect on Antioxidant and Cytotoxic Potential of the Plant

Abstract

Flavonoids are famous for their antioxidant capacity and redox potential. They can combat with cell aging, lipid peroxidation, and cancer. In the present study, Artemisia annua hybrid (Hyb8001r) was subjected to qualitative and quantitative analysis of flavonoids through HPLC. Rol genes transgenics of A. annua were also evaluated for an increase in their flavonoid content along with an increase in antioxidant and cytotoxic potential. This was also correlated with the expression level of flavonoids biosynthetic pathway genes as determined by real-time qPCR. Phenylalanine ammonia-lyase and chalcone synthase genes were found to be significantly more highly expressed in rol B (four to sixfold) and rol C transgenics (3.8-5.5-fold) than the wild-type plant. Flavonoids detected in the wild-type A. annua through HPLC include rutin (0.31 mg/g DW), quercetin (0.01 mg/g DW), isoquercetin (0.107 mg/g DW) and caffeic acid (0.03 mg/g DW). Transgenics of the rol B gene showed up to threefold increase in rutin and caffeic acid, sixfold increase in isoquercetin, and fourfold increase in quercetin. Whereas, in the case of transgenics of rol C gene, threefold increase in rutin and quercetin, 5 fold increase in isoquercetin, and 2.6-fold increase in caffeic acid was followed. Total phenolics and flavonoids content was also found to be increased in rol B (1.5-fold) and rol C (1.4-fold) transgenics as compared to the wild-type plant along with increased free radical scavenging activity. Similarly, the cytotoxic potential of rol gene transgenics against MCF7, HeLA, and HePG2 cancer cell lines was found to be significantly enhanced than the wild-type plant of A. annua. Current findings support the fact that rol genes can alter the secondary metabolism and phytochemical level of the plant. They increased the flavonoids content of A. annua by altering the expression level of flavonoids biosynthetic pathway genes. Increased flavonoid content also enhanced the antioxidant and cytotoxic potential of the plant.

Enhanced stress tolerance in transformed Ajuga bracteosa Wall. ex Benth. regenerants by upregulated gene expression of metabolic pathways

“…Our findings are supported by previous reports which demonstrated that rolB and C genes increased the amount of artemisinin and its derivatives in Artemisia annua by overexpressing the amorphadiene synthase (ADS), aldehyde dehydrogenase 1 (ALDH1), cytochrome P450, (CYP71AV1), PAL, cholane synthase (CHS), and trichomespecific fatty acyl-CoA reductase 1(TAFR1) genes (Dilshad et al, 2015(Dilshad et al, , 2016. Furthermore, Kayani et al (2016aKayani et al ( , 2017 found that rolABC-positive hairy root clones of A. bracteosa produce enhanced phytoecdysteroid content (up to 4449 μg g −1 ) compared to control hairy roots.…”

Neuroprotective, antidiabetic and antioxidant effect of Hedera nepalensis and lupeol against STZ + AlCl3 induced rats model

“…After dissection of rats, their midbrain region of CNS was dissected and washed thrice with cold saline and samples were quickly subjected to storage at −70 °C until estimation of dopamine and serotonin was done by HPLC as reported earlier [ 36 , 37 ]. Briefly, tissues were homogenized in phosphate buffer and then subjected to centrifugation at 14000 rpm for 10 min at 4 °C.…”

Artemisinin and Total Flavonoid Content in In Vitro Cultivted Artemisia Annua L. "Hairy" Root Culture

“…Recently it was shown by Dilshad et al (2016) that transgenic plants of Artemisia annua carrying rol B gene demonstrated threefold increase in rutin and caffeic acid, sixfold increase in isoquercetin, and fourfold increase in quercetin as compared to the wild-type plants. They suggested that flavonoids content increase was caused by altering the expression level of the genes of flavonoids biosynthetic pathway.…”