Effect of cytokinin on alkaloid accumulation in periwinkle callus cultures transformed with a light-inducible ipt gene

The effect of cytokinins on accumulation of indole alkaloids in periwinkle callus cultures was investigated. Firstly, it was found that exogenously-applied cytokinin increased the ajmalicine and serpentine content of untransformed callus culture obtained from cotyledons. Secondly, periwinkle cotyledons were transformed with the isopentenyl transferase (ipt) gene under the control of a light-inducible promoter and two transformed callus lines were used in order to investigate whether endogenously-produced cytokinin could also increase the alkaloid production. We found that the ipt-transgenic tissues accumulated higher levels of isopentenyl transferase transcripts as well as zeatin riboside, even under non-inductive condition, but lower concentration of alkaloids compared to that of untransformed tissues. A 28 kDa polypeptide whose accumulation was previously found to be associated with alkaloid production in a periwinkle cell suspension was also present in the non-transformed tissue and its level was increased in parallel to the CK-enhanced alkaloid production. Neither light induction condition, nor exogenous cytokinin treatment led to the increase of the 28 kDa polypeptide accumulation in the transformed tissues. All these data show that endogenously-produced cytokinin does not mimic the effect of exogenously-applied cytokinin on the alkaloid production in periwinkle calli.