BACKGROUND: Viola philippica and V. prionantha develop chasmogamous (CH) flowers under ≤ 12-h daylight conditions and cleistogamous (CL) flowers under long daylight (>
12-h daylight) conditions (LD), whereas V. cornuta develops CH flowers regardless of the daylight conditions. APETALA3 (AP3) is a major floral B-function gene that regulates the organ identity and development of stamens and petals. Evolutionary changes in AP3 orthologous genes might involve in the dimorphic flower formation. In the present study, we compared AP3 orthologous genes among three Viola species. RESULTS: The AP3 sequences were highly conserved, and obligate AP3-PISTILLATA heterodimers were universally formed. However, the floral expression of VphAP3 in V. philippica and VprAP3 in V. prionantha changed in response to the photoperiod. Their expression was significantly higher under 12-h daylight conditions than under 16-h daylight conditions. In contrast, VcoAP3 expression in the floral buds of V. cornuta was comparable among photoperiods. In accordance with these variations in expression, correlated sequence divergences were observed in the putative regulatory regions of Viola AP3 orthologous genes. CONCLUSIONS: Developmental inhibition of petals and stamens may result from AP3 downregulation by LD, which thereby induces CL flowers. Our study provides insight into the molecular basis underlying the developmental evolution of environmentally dependent mating systems in dimorphic CL plants.