Localized High Expression of Type-A Response Regulator and Cytokinin Oxidase/dehydrogenase Genes in Relation to Forchlorfenuron-induced Changes in Flower Morphology in Torenia fournieri Lind.

Abstract

Several morphological changes in flowers can be induced in torenia (Torenia fournieri L.) by applying forchlorfenuron (CPPU) to flower buds. We investigated the temporal and spatial distributions of the cytokinin response in CPPU-treated flower buds, which is indicated by type-A response regulator (RR) and cytokinin oxidase (CKX) gene expression. The quantitative real-time PCR analysis showed that the expression of both T. fournieri RR1 (TfRR1) and TfCKX5 was induced from 1 day after CPPU treatment in the sepal, petal, stamen, and pistil, and maintained at a high level until 5 days after the treatment, when the earliest morphological changes due to CPPU treatment were observed. In situ hybridization analysis showed weak expression of both genes in the stamen and pistil through all floral stages of non-treated flower buds. However, when CPPU was applied at the sepal development stage, expression of both genes was strongly induced in the abaxial side of the stamen primordia, which is the site of initiation of the wide paracorolla. When CPPU was applied during the early stage of corolla development, high expression of those genes was observed in the stamen, basal, and middle part of the petal, which is the site of initiation of the narrow paracorolla. Those high levels of expression became more strongly localized to the paracorolla initiation site once the paracorolla primordia formed. When CPPU was applied during the middle corolla development stage, strong expression of those genes was detected in the middle to apical parts of the petal, which is the site of changes in the distribution pattern of the vascular bundles and the resultant serrated margin. These results suggest that long-term elevation of cytokinin signaling caused by CPPU treatment induces changes in flower morphology, and the paracorolla and serrated margin of the petal are induced by localized high levels of cytokinin signaling at the site of those morphological changes within flower buds.