Abstract
Lithospermum erythrorhizon produces shikonin derivatives, red naphthoquinone pigments, in the roots. In the cell cultures of L. erythrorhizon, stimulators of shikonin biosynthesis (Cu2+, oligogalacturonide, and MJ) as well as its inhibitors (NH4+, 2,4-D, and light) have been identified. Shikonin molecule is formed in vivo from geranyldiphosphate derived via mevalonate pathway and p-hydroxybenzoate via shikimate pathway. However, the biosynthetic reactions responsible for the formation of naphthalene ring are not clarified yet.
Utilizing the dark-inducibility of shikonin synthesis, we applied PCR-select cDNA subtraction technique to L. erythrorhizon cell cultures to isolate cDNAs encoding enzymes involved in the naphthalene frame formation. Out of 240 clones whose expressions were enhanced in the dark, we have found two candidate genes for the naphthalene-forming enzyme, i.e. catechol polyphenol oxidase-like protein and a member of berberine-bridge enzyme family, adding to preciously identified oxidoreductase genes. Characterization of these genes will be presented.
