AUTHOR=Zhang Haiyan , Shi Yuqin , Sun Mengdan , Hu Xuezhi , Hao Mengyu , Shu Yu , Zhou Xue-Rong , Hu Qiong , Li Chao , Mei Desheng 

TITLE=Functional Differentiation of BnVTE4 Gene Homologous Copies in α-Tocopherol Biosynthesis Revealed by CRISPR/Cas9 Editing

JOURNAL=Frontiers in Plant Science

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2022.850924

DOI=10.3389/fpls.2022.850924

ISSN=1664-462X

ABSTRACT=<p>Tocopherols are essential nutrients for human health known as vitamin E. Vitamin E deficiency can have a profound effect on human health, including the central nervous system and cardiovascular and immune protection. Multiple enzymatic steps are involved in the conversion between different forms of tocopherols. Among them, γ-tocopherol methyltransferase encoded by gene <italic>VTE4</italic> catalyzes the conversion of γ- to α-tocopherol or δ- to β-tocopherol isoforms. However, the gene copies and their functional contribution of <italic>VTE4</italic> homologs in <italic>Brassica napus</italic> were not elucidated. To this end, different mutation combinations of four putative <italic>BnVTE4</italic> homologous copies were generated by using CRISPR/Cas9 genome editing technology. Editing of those <italic>BnVTE4</italic> homologs led to a significant change of the α-tocopherol content and the ratio between α- and γ-tocopherol compared with wide-type control. Analysis of the different combinations of <italic>BnVTE4</italic>-edited homologs revealed that the contribution of the <italic>BnVTE4</italic> individual gene displayed obvious functional differentiation in α-tocopherol biosynthesis. Their contribution could be in order of <italic>VTE4.C02-2</italic> (BnaC02G0331100ZS) &gt; <italic>VTE4.A02-1</italic> (BnaA02G0247300ZS) &gt; <italic>VTE4.A02-2</italic> (BnaA02G0154300ZS). Moreover, the <italic>VTE4.A02-1</italic> and <italic>VTE4.A02-2</italic> copies might have severe functional redundancies in α-tocopherol biosynthesis. Overall, this study systemically studied the different effects of <italic>BnVTE4</italic> homologs, which provided a theoretical basis for breeding high α-tocopherol content oilseed rape.</p>