Accumulation and regulation of anthocyanins in white and purple Tibetan Hulless Barley (Hordeum vulgare L. var. nudum Hook. f.) revealed by combined de novo transcriptomics and metabolomics

Abstract

Background Colored barley, which may have associated human health benefits, is more desirable than the standard white variety, but the metabolites and molecular mechanisms underlying seedcoat coloration remain unclear. Results Here, the development of Tibetan hulless barley was monitored, and 18 biological samples at 3 seedcoat color developmental stages were analyzed by transcriptomic and metabolic assays in Nierumuzha (purple) and Kunlun10 (white). A total of 41 anthocyanin compounds and 4186 DEGs were identified. Then we constructed the proanthocyanin-anthocyanin biosynthesis pathway of Tibetan hulless barley, including 19 genes encoding structural enzymes in 12 classes (PAL, C4H, 4CL, CHS, CHI, F3H, F3’H, DFR, ANS, ANR, GT, and ACT). 11 DEGs other than ANR were significantly upregulated in Nierumuzha as compared to Kunlun10, leading to high levels of 15 anthocyanin compounds in this variety (more than 25 times greater than the contents in Kunlun10). ANR was significantly upregulated in Kunlun10 as compared to Nierumuzha, resulting in higher contents of three anthocyanins compounds (more than 5 times greater than the contents in Nierumuzha). In addition, 22 TFs, including MYBs, bHLHs, NACs, bZips, and WD40s, were significantly positively or negatively correlated with the expression patterns of the structural genes. Moreover, comparisons of homologous gene sequences between the two varieties identified 61 putative SNPs in 13 of 19 structural genes. A nonsense mutation was identified in the coding sequence of the ANS gene in Kunlun10. This mutation might encode a nonfunctional protein, further reducing anthocyanin accumulation in Kunlun10. Then we identified 3 modules were highly specific to the Nierumuzha (purple) using WGCNA. Moreover, 12 DEGs appeared both in the putative proanthocyanin-anthocyanin biosynthesis pathway and the protein co-expression network were obtained and verified. Conclusion Our study constructed the proanthocyanin-anthocyanin biosynthesis pathway of Tibetan hulless barley. A series of compounds, structural genes and TFs responsible for the differences between purple and white hulless barley were obtained in this pathway. Our study improves the understanding of the molecular mechanisms of anthocyanin accumulation and biosynthesis in barley seeds. It provides new targets for the genetic improvement of anthocyanin content and a framework for improving the nutritional quality of barley.

Genome-wide identification of WD40 transcription factors and their regulation of the MYB-bHLH-WD40 (MBW) complex related to anthocyanin synthesis in Qingke (Hordeum vulgare L. var. nudum Hook. f.)

“…f), a common barley species belonging to the wheat family gramineae, is also called hulless barley or naked barley because its lemma and palea are separated during harvesting [ 1 ]. The Qinghai–Tibet Plateau is the main cultivation area of Qingke, and it is distributed in the plateau areas at elevations of 2400–4500 m [ 1 ]. It is one of the main food crops for the people in Tibet.…”

Integrative metabolomic and transcriptomic analyses reveal the mechanisms of Tibetan hulless barley grain coloration

“…The central biosynthetic pathway producing anthocyanin has been well studied previously ( Koes et al., 2005 ). Previous studies have identified multiple anthocyanin biosynthesis genes ( PAL , C4H , 4CL , CHS , CHI , F3H , F3’H , DFR , ANS , GT , and ACT ) showing the significantly higher expression in colored qingke, and their expression was strongly correlated with the content of anthocyanins ( Yao et al., 2022 ). It is important to note that the initial steps to generate anthocyanin have been well-studied.…”

“…Integrating transcriptomic and metabolomic analyses in wheat has revealed that variations in anthocyanin content are highly linked to the differential expression of key structural genes, such as TaCHS and TaANS, in the anthocyanin biosynthesis pathways (Wang et al, 2021). Yao et al (2022) also identified a series of structural genes and transcription factors in anthocyanin pathways showing the differential expression between purple and white qingke seeds.…”

“…Cultivated qingke shows different grain coat colors, such as white, blue, and purple. Recently, the demand for colored qingke grain as a functional food has increased because of its higher levels of bioactive compounds, such as anthocyanins, compared to white qingke ( Bellido and Beta, 2009 ; Yao et al., 2022 ). Anthocyanins have essential health properties that reduce the incidence of cancer, cardiovascular disease, and other chronic diseases ( Hatier et al., 2008 ).…”

“…In recent years, technical advancements in transcriptome and metabolome analyses have provided highly effective ways to identify new genes and metabolites and understand the underlying molecular mechanism of plant organ coloration ( Wang et al., 2020 ; Yao et al., 2022 ). Integrating transcriptomic and metabolomic analyses in wheat has revealed that variations in anthocyanin content are highly linked to the differential expression of key structural genes, such as TaCHS and TaANS , in the anthocyanin biosynthesis pathways ( Wang et al., 2021 ).…”