The ga3ox1b mutation reveals the crosstalk between gibberellin and other phytohormones in controlling the growth and development of female flowers in Cucurbita pepo

Abstract

Cucurbita pepo is a monoecious species with female and male unisexual flowers on the same plant. The large size of its flowers makes this species a model for studying the regulation of sex determination and sex-associated developmental traits. Because male flowers appear at earlier stages of plant development than female flowers, the plant has evolved a system in which female flowers grow and open much faster than male flowers in order to synchronize male and female flowering for proper pollination and fertilization. In this paper, we show that this differential growth of female and male flowers is controlled by gibberellin (GA). We identify and characterize a loss-of-function mutation in the GA biosynthetic gene CpGA3ox1B, which results in a deficiency of active GA in various vegetative and reproductive organs. The dwarf phenotype of the ga3ox1b mutant is similar to that of other plants deficient in GA, but surprisingly, the GA mutation affects only the development of female flowers, which are much smaller, do not open, and are completely sterile. However, male flowers develop normally, reaching the same size and fertility as male WT flowers. In the WT, female flowers have higher expression of CpGA3ox1B than male flowers, causing them to accumulate more active GA and grow faster than male flowers. The mutant dwarf plant and flower phenotypes were rescued by GA3, but not ethylene or JA, indicating that the ga3ox1b flower did not reach the appropriate size to become sensitive to ethylene and JA, the hormones controlling petal expansion and flower opening in later stages of female flower development. The GA mutation does not affect ethylene-regulated sex determination at the earliest floral meristem, but it does affect the growth of the corolla and pistil once the meristem is determined to be a female flower. The transcriptomic changes and differential accumulation of other phytohormones in the WT and ga3ox1b female organs revealed that the function of GA depends on its interaction with other phytohormones, including auxin and cytokinin as GA synergists and ethylene, ABA, and JA as GA antagonists in the development of female floral organs. Completely opposite transcriptomic changes in the ovary of ga3ox1b and the ethylene-insensitive mutant etr2b demonstrate the mutual exclusion of ethylene and GA signaling at different stages of female flower development and highlight the importance of several transcription factors, such as MYB62, for the integration of ethylene and GA in the regulation of both sex determination and flower organ growth and development in C. pepo.