Overexpression of ThMYC4E Enhances Anthocyanin Biosynthesis in Common Wheat

Abstract

The basic helix-loop helix (bHLH) transcription factor has been inferred to play an important role in blue and purple grain traits in common wheat, but to date, its overexpression has not been reported. In this study, the bHLH transcription factor ThMYC4E, the candidate gene controlling the blue grain trait from Th. Ponticum, was transferred to the common wheat JW1. The positive transgenic lines displayed higher levels of purple anthocyanin pigments in their grains, leaves and glumes. Stripping the glumes (light treatment) caused white grains to become purple in transgenic lines. RNA-Seq and qRT-PCR analysis demonstrated that the transcript levels of structural genes associated with anthocyanin biosynthesis were higher in transgenic wheat than the wild-type (WT), which indicated that ThMYC4E activated anthocyanin biosynthesis in the transgenic lines. Correspondingly, the anthocyanin contents in grains, roots, stems, leaves and glumes of transgenic lines were higher than those in the WT. Metabolome analysis demonstrated that the anthocyanins were composed of cyanidin and delphinidin in the grains of the transgenic lines. Moreover, the transgenic lines showed higher antioxidant activity, in terms of scavenging DPPH radicals, in the ethanol extracts of their grains. The overexpression of ThMYC4E sheds light on the traits related to anthocyanin biosynthesis in common wheat and provide a new way to improve anthocyanin content.

Anthocyanin Biosynthesis and a Regulatory Network of Different-Colored Wheat Grains Revealed by Multiomics Analysis

“…In the BSR-seq data, no SNVs were detected in TraesCS2A02G409400.1 ( MYC1 ) in 29p, 29b, and 29w. However, in 29b, TraesCS4D02G224600.1 (which was named MYC4 in this study) was detected with many SNVs and was a homologue of HvMYC4H and ThMYC4E . ,, Of those SNVs, 16 nonsynonymous SNVs were verified by Sanger sequencing (Figure o,p). In addition, the variable splicing mode of the gene in blue wheat differed from the reference database (Figure n,o); this was the key domain, bHLH_MYC_N, that is bonded to the MYB transcription factor.…”

“…Moreover, F3′5′H 2 was also upregulated and its high expression was necessary to the cluster involved in the blue aleurone of wheat because ThMYC4E overexpression in common wheat only enhances F3′H expression, producing purple grains instead of blue grains. 21 In the BSR-seq analysis, TaMYC4 and F3′5′H 2 were both located in candidate regions related to blue grains, because they were closely located on chromosome 4D. No MYB transcription factors with significant changes in expression were found nearby, but in the candidate regions related to blue grains, another MYB TraesCS2A02G456600.1 (REVEILLE 7) was upregulated in XY22b and could be associated with anthocyanin accumulation in blue grains.…”

“…In barley, the Blx1 locus was involved in blue aleurone and was identified as a trigenic cluster containing HvMYB4H , HvMYC4H, and HvF35H . , In XY22b, an MYC on chromosome 4D ( TaMYC4 ), which was homologous with ThMYC4E , was found to be upregulated and could play an important role in forming blue grains. Moreover, F3′5′H 2 was also upregulated and its high expression was necessary to the cluster involved in the blue aleurone of wheat because ThMYC4E overexpression in common wheat only enhances F3′H expression, producing purple grains instead of blue grains . In the BSR-seq analysis, TaMYC4 and F3′5′H 2 were both located in candidate regions related to blue grains, because they were closely located on chromosome 4D.…”

“…In a variety of plants, MYBs and bHLHs regulate the accumulation of anthocyanins by affecting the expression of anthocyanin biosynthesis genes . In wheat with purple pericarps, TaPpm1/Tamyb7D (R2R3-MYB) and TaPpb1/TaMyc1 (bHLH) coregulate anthocyanin biosynthesis. , TsMYC2 and HvMYC4H were associated with blue grains in triticale and barley. , The blue grain genes of hexaploid wheat could come from Blue aleurone 1 ( Ba1 ) and Blue aleurone 2 ( Ba2 ) in Thinopyrum ponticum and Triticum monococcum , while the gene ThMYC4E was identified as the candidate gene of Blue aleurone 1 . ,− …”

Wheat grain phenolics: a review on composition, bioactivity, and influencing factors

“…103 The overexpression of bHLH transcription factor ThMYC4E activates anthocyanin biosynthesis in the transgenic lines, which enhances excess anthocyanin content formation. 104 Progeny analysis from crosses between red-, purple-, and bluegrained wheat varieties showed that it can be an effective way to increase anthocyanin content in wheat grain by combining the different genetic backgrounds. 105…”