Seed-Specifically Overexpressed Arabidopsis Cytochrome P450 85A2 Promotes Vegetative and Reproductive Growth and Development of Arabidopsis thaliana

Arabidopsis Cytochrome P450 85A2 (AtCYP85A2) was introduced to Arabidopsis thaliana seeds using a seed-specific promoter, pAt5g54000. GUS (β-Glucuronidase) activity and RT-PCR analysis demonstrated that AtCYP85A2 overexpression only occurred in seeds of a transgenic plant, pAt5g54000-AtCYP85A2::Col-0. A crude enzyme solution prepared pAt5g54000-AtCYP85A2::Col-0 seeds successfully catalyzed the conversion of castasterone (CS) to brassinolide (BL), which was not detected in wild-type seeds. Furthermore, a higher level of CS and BL was detected in pAt5g54000-AtCYP85A2::Col-0 seeds compared to untransformed seeds, thus demonstrating that seed-specific overexpression of AtCYP85A2 efficiently encoded a bi-functional enzyme for brassinosteroids 6-oxidase/brassinolide synthase to generate CS and BL in seeds of pAt5g54000-AtCYP85A2::Col-0. Compared to the wild type, pAt5g54000-AtCYP85A2::Col-0 produced substantially larger seeds with a high concentration of nutrients due to an enhancement in brassinosteroids signaling. Additionally, pAt5g54000-AtCYP85A2::Col-0 exhibited superior seed germination, seedling and rosette plant growth, and flower and silique formation, indicating that seed-specific AtCYP85A2 expression activates overall vegetative and reproductive growth and development in A. thaliana.