The potential role of two LEAFY orthologs in the chasmogamous/cleistogamous mixed breeding system of Viola pubescens (Violaceae)

Many plants, including most species in Viola, produce both open, outcrossing chasmogamous and closed, self-pollinating cleistogamous flowers. The chasmogamous/cleistogamous mixed breeding system is considered an evolutionarily successful reproductive strategy, but the underlying molecular basis remains largely unknown. The LEAFY (LFY) gene in Arabidopsis is responsible for the initiation of floral meristems and the regulation of flower development; therefore, we sought to identify LFY orthologs in Viola pubescens Aiton with the goal of understanding possible differences in the regulatory genetics of flower development and the molecular genetics underlying chasmogamous/cleistogamous mixed breeding systems. A genomic library of V. pubescens was constructed to identify LFY orthologs, and reverse transcription polymerase chain reaction was employed to study gene expression in both flower types. In addition, overexpression studies of the orthologs were carried out in Arabidopsis to explore their role in the breeding system of Viola. Two LFY orthologs, VpLFY1 and VpLFY2, were isolated from a genomic library of V. pubescens. Both genes were expressed in both flower types, especially in young floral buds. However, over-expression of VpLFY2, but not VpLFY1, caused precocious flowering in Arabidopsis, indicating a potentially unique role of each ortholog. The results suggest that VpLFY1 and VpLFY2 function together in flower development of V. pubescens. The different effects caused by VpLFY1 and VpLFY2 in Arabidopsis indicate that the two orthologs may have unique, gene-specific properties that might contribute to the flower type in violets.