The effects of Ca2+ during the elicitation of shikonin derivatives in Onosma paniculatum cells

A fungal elicitor extracted fi om Aspergillus oryzae (Ahlb.) Cobn mycelia promoted the production of shikonin derivatives in Onosma paniculatum Bur et Franch cell suspension cultures. Elicitor treatment also increased Ca2+ concentration in RM, medium, which could be measured earlier than the elicited increase of shikonin formation. Several reagents known to induce Ca2+-influx and increase the intracellular-free Ca2+ level, such as the addition of Ca (NO3)(2). 4H(2)O, the Ca2+ ionophore A(23187), and abscisic acid (ABA), appreciably suppressed the elicitor-promoted shikonin formation in Onosma cells. In contrast, the decrease of intracellular-free Ca2+ level by the specific Ca2+-chelator ethylene glycol bis (beta-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA) or the Ca2+-channel blocker, verapamil, enhanced the biosynthesis of shikonin even in the absence of elicitor. Treatment of cells with trifluoperazine (TFP) also stimulated shikonin formation in Onosma cell cultures. A rapid and transient drop of fi ee Ca2+ level in one protoplast was directly determined after the addition of elicitor to Onosma cell cultures. The inhibitory effect on shikonin formation by ABA was largely on account of its ability to restore the intracellular Ca2+ level lowered by the elicitor. These results suggest that Ca2+ play a significant role in an early stage of the elicitation process of Onosma cells. The rapid drop of cytoplasmic Ca2+ carries the elicitor signal and in turn regulates the biosynthesis of shikonin derivatives.