Uniconazole [S-3307; (E)-1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-1-penten-3-ol], a synthetic plant-growth retardant, inhibited the differentiation of isolated mesophyll cells of Zinnia elegans L. into tracheary elements (TEs) but had no effect on cell division when it was added to the culture medium at a concentration of 3.4-mu-M. In the presence of uniconazole, none of the cytological events characteristic of the processes of TE differentiation, such as aggregation of actin filaments, bundling of microtubules or localized thickening and lignification of secondary walls, was observed. Uniconazole was effective when it was added to the medium within 36 h after the start of culture. Brassinosteroids (0.2 nM brassinolide or 2-mu-M homobrassinolide), but not gibberellin A3, counteracted the inhibitory effect of uniconazole on TE differentiation. Brassinosteroids were most effective when they were added to cultures between 24 and 30 h after the start of culture. Exogenously applied brassinosteroids promoted TE differentiation. It is suggested that the synthesis of brassinosteroids is essential for the differentiation of the cells into TEs and that uniconazole inhibits this differentiation through its inhibitory effect on the biosynthesis of brassinosteroids.