Features of Activity of the Phenylpropanoid Biosynthesis Pathway in Melanin-Accumulating Barley Grains

Abstract

Barley (Hordeum vulgare L.) grain pigmentation is caused by two types of phenolic compounds: anthocyanins (which are flavonoids) give a blue or purple color, and melanins (which are products of enzymatic oxidation and polymerization of phenolic compounds) give a black or brown color. Genes Ant1 and Ant2 determine the synthesis of purple anthocyanins in the grain pericarp, whereas melanins are formed under the control of the Blp1 gene in hulls and pericarp tissues. Unlike anthocyanin synthesis, melanin synthesis is poorly understood. The objective of the current work was to reveal features of the phenylpropanoid biosynthesis pathway functioning in melanin-accumulating barley grains. For this purpose, comparative transcriptomic and metabolomic analyses of three barley near-isogenic lines accumulating anthocyanins, melanins, or both in the grain, were performed. A comparative analysis of mRNA libraries constructed for three stages of spike development (booting, late milk, and early dough) showed transcriptional activation of genes encoding enzymes of the general phenylpropanoid pathway in all the lines regardless of pigmentation; however, as the spike matured, unique transcriptomic patterns associated with melanin and anthocyanin synthesis stood out. Secondary activation of transcription of the genes encoding enzymes of the general phenylpropanoid pathway together with genes of monolignol synthesis was revealed in the line accumulating only melanin. This pattern differs from the one observed in the anthocyanin-accumulating lines, where — together with the genes of general phenylpropanoid and monolignol synthesis pathways — flavonoid biosynthesis genes were found to be upregulated, with earlier activation of these genes in the line accumulating both types of pigments. These transcriptomic shifts may underlie the observed differences in concentrations of phenylpropanoid metabolites analyzed in the grain at a late developmental stage by high-performance liquid chromatography. Both melanin-accumulating lines showed an increased total level of benzoic acids. By contrast, anthocyanin-accumulating lines showed higher concentrations of flavonoids and p-coumaric and ferulic acids. A possible negative effect of melanogenesis on the total flavonoid content and a positive influence on the anthocyanin content were noted in the line accumulating both types of pigments. As a conclusion, redirection of metabolic fluxes in the phenylpropanoid biosynthesis pathway occurs when melanin is synthesized.

Transcriptional convergence after repeated duplication of an amino acid transporter gene leads to the independent emergence of the black husk/pericarp trait in barley and rice

“…Out of the 11 candidate genes mapped in Hor13821, only HvBlp was highly and specifically expressed in the black-coloured husk/pericarp tissues in BNIL at S3 stage: BS3, while the other 10 genes either were barely expressed or displayed no significant difference between BINL and YNIL (Figure 1d; Table S2). In addition, we analysed another published transcriptome datasets of different black-and yellow-coloured barley lines (Table S2; Glagoleva et al, 2022) and confirmed that HvBlp was uniquely transcribed in the black barley genotype. Next, we performed qRT-PCR on HvBlp in 100 black and 100 yellow inbred lines.…”

“…(a) The transcriptional expression of Blp1 and Blp2 homologous genes in the spikes of barley NILs BLP (black lemma and pericarp), PLP (purple lemma and pericarp), BP (black and purple) and of cv. Bowman (BW) at the booting stage (1), late milk stage (2), and early dough stage (3) (Glagoleva et al ., 2022 ). (b) The transcriptional expression of Blp1 and Blp2 homologous genes in the husk/pericarp tissues of BNIL and YNIL at green (early milk stage, S1) and black grain stages (late milk, S3).…”

“…Black‐coloured rice and barley have attracted enormous human attention not only for their distinctive appearance but also due to their exceptional nutritional values as healthy foods (Ge et al ., 2021 ; Shen et al ., 2016 ). The accumulation of melanins, usually accompanied by enriched phenolics and flavonoids, exhibits strong antioxidant activity with proven beneficial effects on human health (Ge et al ., 2021 ; Glagoleva et al ., 2022 ). In addition to the antioxidant effects, the accumulation of melanins is also believed to increase the mechanical strength of seeds, protecting them from insect and mechanical damage (Jocković et al ., 2020 ), and microbial pathogen infections (Choo et al., 2015 ; Glagoleva et al., 2020 ).…”

“…In barley, the black grain trait has long been known to be controlled by a single dominant locus Blp on chromosome 1H (Costa et al ., 2001 ). Several previous studies have attempted to identify the Blp gene (Bungartz et al ., 2016 ; Glagoleva et al ., 2017 , 2022 ; Jia et al ., 2017 ; Long et al ., 2019 ; Shoeva et al ., 2016 ) and have narrowed it to an extremely small genetic region, containing only 21 annotated genes (Long et al ., 2019 ). However, the corresponding candidate gene is still hiding in mystery.…”

Melanin found in wheat spike husks

“…Dark-colored barley seeds have higher contents of phenolic compounds and lignin than uncolored ones. It was suggested that melanin biosynthesis genes may be connected to phenylpropanoid-derived biosynthesis pathways [9].…”