Title : Behind Black: Unfolding the hidden secrets of plant melanins
Melanin is considered to be one of the earliest eukaryotic pigments to have arisen during evolution. They are synthesized from phenolic precursors, which are oxidized by polyphenol oxidase into quinone, which is subsequently polymerized. Based on the monomers, three classes of melanin are recognized: eumelanins, pheomelanins and allomelanins. Eumelanin is produced by oxidative polymerization of tyrosine or phenylalanine into L-3,4-dihydroxyphenylalanine, which is converted into dopachrome and then to melanin. It is the predominant form found in animals and microorganisms. The pheomelanins are also formed from tyrosine, but contain sulfur. It found only in certain yellow, orange or reddish hair and feathers. Plant and fungal melanins, classified as allomelanins. It is the most heterogeneous group: their precursors are particularly varied. In plants, melanins are widely presented in seeds where it protect embryo against impurity. Much of the research directed at its biochemistry and synthesis has been directed at animals and micro-organisms, with little focus as yet placed on plants. The pigment responsible for the black spike formed by certain varieties of barley has long been suspected to be a melanin. The black spike trait is under monogenic control of the Blp gene mapped to chromosome 1H. This simple mode of inheritance has facilitated the breeding of lines which are near isogenic for Blp. The objective of the present study was to exploit these near isogenic lines to reveal the molecular and cellular basis of melanin formation in barley. The pigment was extracted and purified from the spike of the Blp-carrying near isogenic line and subjected to a series of solubility tests, which indicated that the pigment was indeed melanin. This conclusion was supported by an analysis based on Fourier transform infrared spectroscopy. A comparison at the microscopic level between immature grain harvested at key developmental stages from the two near isogenic lines showed that the pigment is accumulated in the pericarp and husk within organelles derived from chloroplasts. We have demonstrated that, just as in animal cells, plant cells synthesize melanin in a membrane-delimited organelle. The comparative RNA-seq analysis of the near isogenic lines in hulls and grain pericarp revealed the influence of the Blp locus on expression more than thousand genes. Among them, the genes belonging phenylpropanoid and fatty acids biosynthesis pathways were the most represented and upregulated in black-colored line. To study the relationships between chlorophyll and melanin biosynthesis the hybrid i:BwBlpalm line characterized simultaneously by partial chlorophyll insufficiency and accumulation of melanins in spike were developed. Distinct dynamics of melanin accumulation was observed in the developed line in comparison to the black-colored parental one. Although the key genes responsible for melanin accumulation remains to be identified, in the current study the great progress has been achieved in understanding of the melanogenesis in plant species. The study was supported by RSF grants ?16-04-00086 and ?19-76-00018.