BrTTG1 regulates seed coat proanthocyanidin formation through a direct interaction with structural gene promoters of flavonoid pathway and glutathione S-transferases in Brassica rapa L

Wenju Zhao; Xiaojuan Li; Junqin Wen; Quanhui Li; Shuanling Bian; Yanjing Ren

doi:10.3389/fpls.2024.1372477

BrTTG1 regulates seed coat proanthocyanidin formation through a direct interaction with structural gene promoters of flavonoid pathway and glutathione S-transferases in Brassica rapa L

Front Plant Sci. 2024 Apr 4:15:1372477. doi: 10.3389/fpls.2024.1372477. eCollection 2024.

Authors

Wenju Zhao¹, Xiaojuan Li¹, Junqin Wen^{1

2}, Quanhui Li^{1

2}, Shuanling Bian¹, Yanjing Ren^{1

2}

Affiliations

¹ Qinghai University, Academy of Agriculture and Forestry Sciences of Qinghai Province, Laboratory of Research and Utilization of Germplasm Resources in Qinghai-Tibet Plateau, Qinghai, Xining, China.
² Key Laboratory of Germplasm Resources Protection and Genetic Improvement of the Qinghai-Tibet Plateau in Ministry of Agriculture and Rural, Qinghai, Xining, China.

Abstract

Introduction: Seed coat color is a significant agronomic trait in horticultural crops such as Brassica rapa which is characterized by brown or yellow seed coat coloration. Previous Brassica rapa studies have shown that BrTTG1 is responsible for seed coat proanthocyanidin formation, which is dependent on the MYB-bHLH-WD40 complex, whereas some studies have reported that TRANSPARENT TESTA GLABRA 1 (TTG1) directly interacts with the structural gene promoters of the flavonoid pathway.

Methods: Herein, the brown-seeded inbred B147 and ttg1 yellow-seeded inbred B80 mutants were used as plant materials for gene expression level analysis, gene promoter clone and transient overexpression.

Results: The analysis identified eleven structural genes involved in the flavonoid biosynthesis pathway, which are potentially responsible for BrTTG1- dependent seed coat proanthocyanidin formation. The promoters of these genes were cloned and cis-acting elements were identified. Yeast one-hybrid and dual-luciferase assays confirmed that BrTTG1 directly and independently interacted with proCHS-Bra008792, proDFR-Bra027457, proTT12-Bra003361, proTT19-Bra008570, proTT19-Bra023602 and proAHA10-Bra016610. A TTG1-binding motif (RTWWGTRGM) was also identified. Overexpression of TTG1 in the yellow-seed B. rapa inbred induced proanthocyanidin accumulation by increasing the expression levels of related genes.

Discussion: Our study unveiled, for the first time, the direct interaction between TTG1 and the promoters of the flavonoid biosynthesis pathway structural genes and glutathione S-transferases in Brassica rapa. Additionally, we have identified a novel TTG1-binding motif, providing a basis for further exploration into the function of TTG1 and the accumulation of proanthocyanidins in seed coats.

Keywords: BrTTG1; Brassica rapa L.; flavonoid biosynthesis pathway; gene overexpression; proanthocyanidins; promoters.

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was supported by the National Natural Science Foundation of China (31960602), Project of Qinghai Provincial Science and Technology Department (2021-ZJ-763), and National Key Research and Development Program (2022YFD1602400).