Biology subjects

Ichino, T.

Publications and source records attributed to Ichino, T..

4 recordsLinked to original sources

bioRxiv2026

Accumulation of two distinct alleles causes astringency loss in cultivated polyploid persimmon

Persimmon fruit accumulates high levels of proanthocyanidins (PAs), causing extreme astringency. In pollination-constant and non-astringent (PCNA) cultivars, PA accumulation ceases early in fruit development. Although the PCNA trait is recessively controlled by the ASTRINGENCY (AST) locus, its causal gene has remained unknown. Using haplotype-resolved forward genetics in a hexaploid genetic background, we identified the previously uncharacterized TANNIN (TAN) as the causal gene. Long-read sequencing revealed that PCNA originated through the accumulation of independently derived ast alleles that cause transcriptional deficiency of TAN. These findings resolve the complex genetic basis of natural astringency loss in persimmon and provide a framework for molecular breeding. Further investigation of TAN function would uncover mechanisms underlying massive accumulation of PAs in persimmon fruit and insight into specialized metabolite sequestration.

plant biology↗

bioRxiv2025

Membrane systems for lipid secretion in the hairy roots of Lithospermum erythrorhizon observed by electron microscopy

Many plant-derived lipophilic compounds benefit human life. For example, shikonin derivatives--lipophilic red pigments from the medicinal plant Lithospermum erythrorhizon--exhibit wound-healing and antimicrobial effects. These compounds are excreted from root epidermal cells and accumulate in the root bark. Our understanding of how lipophilic compounds are transported in plant cells remains limited, and L. erythrorhizon hairy roots serve as a good model for studying the lipid secretion. To investigate this, the shikonin secretion was induced in cultured hairy roots, which were then fixed via chemical fixation or high-pressure freezing followed by freeze-substitution and observed by transmission electron microscopy. Electron micrographs from chemically fixed samples showed multivesicular bodies (MVBs) surrounding electron-dense bodies--presumed to contain shikonin derivatives--and fusing with the plasma membrane. Similarly, freeze-substituted samples revealed MVBs fused with the plasma membrane. Additionally, some MVBs contacted plastids containing enzymes for shikonin precursor biosynthesis. These findings suggest MVBs play key roles in both synthesis and secretion of shikonin derivatives. HighlightThis study reveals that multivesicular bodies mediate both synthesis and secretion of shikonin derivatives in the hairy roots of Lithospermum erythrorhizon.

cell biology↗

bioRxiv2025

CRISPR/Cas9-mediated genome editing reveals the involvement of a polyphenol oxidase in the shikonin-specific biosynthesis in Lithospermum erythrorhizon

Shikonin, a 1,4-naphthoquinone derivative produced by several Boraginaceae species, exhibits unique pharmacological properties and is used as a natural dye. The regulatory factors of shikonin production have been demonstrated using a cell culture system of Lithospermum erythrorhizon. Among these factors, copper is known to be the strongest enhancer of shikonin production. Although shikonin biosynthesis has been studied for over 40 years, the steps of naphthalene ring formation are still unknown, as is the reason for the effect of copper. In this study, we explored candidate genes associated with shikonin production using a PCR-select subtraction experiment. Polyphenol oxidase (PPO), a dicopper-dependent oxidoreductase, was highlighted because it showed synchronous expression with shikonin production. Transcriptome analysis of hairy roots and cultured cells of this plant revealed that, of the five PPO genes expressed in L. erythrorhizon, only PPO1 showed a strong correlation with shikonin production. Next, we generated genome-edited hairy roots of LePPO1 using CRISPR/Cas9-mediated mutagenesis to analyze its impact on shikonin derivative and other specialized metabolite production. The results showed that shikonin content was markedly reduced in all LePPO1-ge lines. Interestingly, the content of deoxyshikonofuran, a hydroquinone derivative and shunt product that branches after GHQ-3''-OH in the shikonin biosynthetic pathway, remained unaffected in the LePPO1-ge lines. These findings suggest that LePPO1 participates in naphthalene ring formation and explain why a copper ion is crucial for shikonin biosynthesis.

biochemistry↗

bioRxiv2021

Excretion of triacylglycerol as a matrix lipid facilitating apoplastic accumulation of a lipophilic metabolite shikonin.

Plants produce a large variety of lipophilic metabolites, many of which are secreted by cells and accumulated in apoplasts. The mechanism of secretion remains largely unknown, because hydrophobic metabolites, which may form oil droplets or crystals in cytosol, inducing cell death, cannot be directly secreted by transporters. Moreover, some secondary metabolic lipids react with cytosolic components leading to their decomposition. Lipophilic metabolites should thus be solubilized by matrix lipids and compartmentalized by membrane lipids. The mechanism of lipophilic metabolite secretion was assessed using shikonin, a red naphthoquinone lipid, in Lithospermum erythrorhizon. Cell secretion of shikonin also involved the secretion of about 30% of triacylglycerol (TAG), composed predominantly of saturated fatty acids. Shikonin production was associated with the induction of large amounts of the membrane lipid phosphatidylcholine. Together with in vitro reconstitution, these findings suggest a novel role for TAG as a matrix lipid for the secretion of lipophilic metabolites.

plant biology↗