30:	The role of ethylene in tobacco ovule development.

Authors:	De Martinis, Domenico(A)Mariani, Celestina(A)
Affiliations:	(A): Catholic University of Nijmegen (B): ENEA
Presenter:	De Martinis, Domenico , demartinis@casaccia.enea.it

The role of ethylene in plant reproduction has been studied in mainly with regards to pollination and flower senescence. Little is known about the role of ethylene in early ovule development; in orchids it has been suggested that ethylene is required for post-pollination ovary development. However, to date, a clear demonstration that ethylene alone controls ovule development was never provided. We studied the role in plant reproduction of a pistil-specific gene encoding for the "ethylene-forming enzyme" ACC oxidase (ACO) previously cloned from a Nicotiana tabacum (tobacco), pistil cDNA library (Pezzotti M. and Mariani C.). In situ hybridization shows that within the ovary, ACO expression is located preferentially on the ovules since early stages of ovule development, suggesting a role of this member of the ACO gene family in influencing the physiology of the reproductive organs in tobacco. Using an antisense and a co-suppression strategy we successfully down-regulated the ACO enzyme-activity in the ovary of transgenic tobacco. The two sets of transformants showed a similar flower phenotype with a reduced size and, importantly, female sterility. Cytological analysis revealed that in the transgenic plants, ovules did not complete megasporogenesis and did not produce an embryo sac. The phenotype obtained is reversible, and flowers from transgenic plants will produce fully developed and functional ovules if an ethylene source becomes available. The reversibility of the phenotype obtained suggests that exogenous ethylene can reactivate the "machinery" necessary for megasporogenesis and embryo sac formation, probably activating the proper ethylene-responsive genes. Together, these results definitively demonstrate that ethylene is an essential hormone for ovule development.