Multiple transcription factors of Arabidopsis thaliana that are activated by LEAFY COTYLEDON 2 regulate triacylglycerol biosynthesis.

ABSTRACT

Plant triacylglycerols (TAG) are used in food and various industrial feedstocks. LEAFY COTYLEDON 2 (LEC2), a master positive regulator of TAG biosynthesis, regulates a complex network of transcription factors (TFs) during seed development. Aside from WRINKLED1 (WRI1), the TFs regulated by LEC2 related to TAG biosynthesis have not yet been identified. Previously, we identified 25 seed-expressing TFs that were upregulated in Arabidopsis leaves that overexpressed senescence-induced LEC2. In this study, each of the 25 TFs was transiently expressed in the leaves of Nicotiana benthamiana to identify unknown TFs that regulate TAG biosynthesis. The TAG content of the transformed leaves was analyzed using thin layer chromatography and gas chromatography. We observed that five TFs, ARABIDOPSIS RESPONSIVE REGULATOR 21 (ARR21), AINTEGUMENTA-LIKE 6 (AIL6), APETALA2/ETHYLENE RESPONSIVE FACTOR 55 (ERF55), WRKY DNA-BINDING PROTEIN 8 (WRKY8), and ARABIDOPSIS NAC DOMAIN CONTAINING PROTEIN 38 (ANAC038) increased TAG synthesis in the leaves. Among these, the promoters of AIL6, ERF55, WRKY8, and ANAC038 contain RY motifs, which are LEC2-binding sites activated by LEC2. AIL6 overexpression in Arabidopsis increased the total fatty acid (FA) content in seeds and altered the FA composition, with increases in 160, 181, and 182 and decreases in 180, 183, and 201 compared with those in the wild type (WT). AIL6 overexpression activates several FA and TAG biosynthesis genes. Therefore, our study successfully identified several new TFs regulated by LEC2 in TAG biosynthesis and showed that AIL6 increased the TAG content in seeds.