Regulation of the Tocopherol Biosynthetic Pathway by AREB۵ in Cannabis sativa

Introduction: The oil extracted from hemp (Cannabis sativa) seeds can be used in human nutrition, and several components of hemp oil such as tocopherols benefit human health. Increasing the content of natural antioxidants like tocopherols in the oil seed is achieved through the metabolic engineering of key enzymes or regulatory transcription factors involved in tocopherol biosynthesis. Although there is significant information regarding the genes involved in the tocopherol biosynthesis pathway, the molecular mechanisms regulating its transcription remain unclear. Materials and Methods: In this study, using fluorescent HPLC, the contents of vitamin E isomers (α, δ, and γ) were measured at four developmental stages (germination, milky, doughy, and mature) of cannabis seed. Based on previous study, candidate TFs involved in tocopherol biosynthesis were identified. RT-qPCR was used to evaluate the expression of candidate transcription factors in different seed stages. Statistical analyses were performed using two-way ANOVA and Pearson’s correlation coefficients. Results: High-performance liquid chromatography (HPLC) identified three tocopherol isomers (α, γ, δ), with γ-tocopherol being predominant. The highest tocopherol concentration (۱۲.۵ µg/g) was observed at the seed maturation stage. Expression analysis of transcription factors (AREB۵, NAC۰۷۲, and PIF۳) revealed significant upregulation of AREB۵ during seed maturation, correlating strongly with tocopherol levels (r²: ۰.۸۶– ۰.۹۴). These findings suggest AREB۵ pivotal role in tocopherol biosynthesis and provide insights for future metabolic engineering in hemp. Conclusions: This study offers insights for ongoing biofortification efforts to breed and/or engineer vitamin E in cannabis and other oilseeds.

کلیدواژه ها:

Hemp ، HPLC ، Seed Development ، Transcription factor ، Vitamin E