Identification of gametophytic mutations affecting female gametophyte development in Arabidopsis

The female gametophyte (embryo sac or megagametophyte) plays a critical role in sexual reproduction of angiosperms. It is the structure that produces the egg cell and central cell which, following fertilization, give rise to the seed's embryo and endosperm, respectively. In addition, the female gametophyte mediates a host of reproductive processes including pollen tube guidance, fertilization, and the induction of seed development. Several major events occur during megagametogenesis, including syncitial nuclear divisions, cellularization, nuclear migration and fusion, and cell death. While these events have been described morphologically, the molecules regulating them in the female gametophyte are largely unknown. We discuss a genetic screen based on reduced seed set and segregation distortion to identify mutations affecting megagametogenesis and female gametophyte function. We report on the isolation of four mutants (fem1, fem2, fem3, and fem4) and show that the four mutations map to different locations within the genome. Additionally, we show that the fem1 and fem2 mutations affect only the female gametophyte, while the fem3 and fem4 mutations affect both the female and male gametophyte. We analyzed female gametophyte development in these four mutants as well as in the gfa2, gfa3, gfa4, gfa5, and gfa7 mutants. We found that the fem2, fem3, gfa4, and gfa5 mutants abort development at the one-nucleate stage, while the fem1, fem4, gfa2, gfa3, and gfa7 mutants are affected in processes later in development such as polar nuclei fusion and cellularization. The establishment of a genetic screen to identify mutants and the development of a rapid procedure for analyzing mutant phenotypes represent a first step in the isolation of molecules that regulate female gametophyte development and function.