ABSTRACT: The life cycle of flowering plants alternates between a diploid sporophytic and a haploid gametophytic generation. After fertilization of each the egg and central cells by one male gamete, the development of both fertilization products occurs coordinated with the maternally derived seed coat and carpel tissues forming the fruit. The reproduction program is likely to involve the concerted activity of many genes. To identify genes with specific functions during reproduction, we have analyzed the expression profile of more than 22,000 genes present on the Arabidopsis ATH1 microarray during three stages of flower and fruit development. We found 1,886 genes regulated during reproductive development and 1,043 genes that were specifically expressed during reproduction. When compared to cells from an Arabidopsis suspension culture, S-phase genes were underrepresented and G2 and M-phase genes were strongly enriched in the set of specific genes, indicating that important functions during reproduction are exerted in the G2 and M phases of the cell cycle. Many potential signaling components, such as receptor-like protein kinases, phosphatases, and transcription factors, were present in both groups of genes. Members of the YABBY, MADS box, and Myb transcription factor families were significantly overrepresented in the group of specific genes, revealing an important role of these families during reproduction. Furthermore, we found a significant enrichment of predicted secreted proteins smaller than 15 kD that could function directly as signaling molecules or as precursors for peptide hormones. Our study provides a basis for targeted reverse-genetic approaches aimed to identify key genes of reproductive development in plants. During evolution, both animals and plants developed distinct pathways of sexual reproduction. The haploid life cycle starts with the formation of the meiotic products, for instance arrested primary oocytes and spermatocytes in mammals or micro- and megaspores in plants. The haploid phase ends when a sperm cell fuses with an egg cell to form the diploid zygote, which initiates embryo development. In animals, gametes are derived from a population of germ cells committed early during development to their reproductive fate and which usually rest until the onset of sexual maturity. By contrast, stem cells in shoot apical meristems of flowering plants divide continuously during postembryonic development. Only after formation of vegetative organs, the meristem produces specialized reproductive organs in which meiosis occurs, the stamens and carpel-borne ovules of the flower (for review, see Bowman and Eshed, 2000; Carles and Fletcher, 2003). As early as 1790, Goethe suggested that floral organs are derived from leaves and adapted to their specialized tasks (Goethe, 1790). Animal gametes differentiate directly from the products of meiosis, whereas in plants the meiotic products (spores) typically undergo two or three mitotic divisions to give rise to the male or female gametophytes, respectively (Drews et al., 1998; Grossniklaus and Schneitz, 1998). In several plant species, including Arabidopsis, the female gametophyte consists of seven cells: three antipodal cells, two synergid cells, one egg cell, and one central cell that contains two polar nuclei (Drews et al., 1998; Grossniklaus and Schneitz, 1998). The mature male gametophyte comprises two sperm cells contained within a vegetative cell (McCormick, 1993; da Costa-Nunes and Grossniklaus, 2003). During fertilization one sperm cell nucleus fuses with the egg cell nucleus, giving rise to the diploid embryo, whereas the other sperm cell fuses with the homodiploid central cell nucleus, generating the triploid endosperm (Drews and Yadegari, 2002). Subsequent seed and fruit development are highly coordinated processes between the embryo and endosperm, as well as the maternally derived testa (seed coat) and carpels. Because of the significant economical importance of flowers, seeds, and fruits, the complex processes involved in plant reproduction are not only of academic interest but also of high economic relevance. Many genes have been identified genetically that have a function in plant reproduction. The establishment of floral organ identity and initiation of seed development have been studied extensively, and many regulators of these processes have been identified (McElver et al., 2001; Zik and Irish, 2003; for review, see Chaudhury et al., 2001). High-throughput RNA profiling technologies can now complement the genetic and molecular approaches to provide new insights into plant reproduction. In particular, oligonucleotide-based microarrays can produce reliable, high-quality data (Lockhart et al., 1996; Hennig et al., 2003) to establish new biological knowledge on the transcriptional programs that are active during developmental processes (Spellman et al., 1998; Harmer et al., 2000; Becker et al., 2003; Honys and Twell, 2003). Here, we report a comprehensive Affymetrix GeneChip analysis of the Arabidopsis transcriptome at key steps of reproductive development. We identified 1,043 genes that were specifically expressed during reproduction. Among those genes were many potential signaling components, such as receptor-like protein kinases, phosphatases, and transcription factors. In addition, we found a significant enrichment of predicted secreted proteins smaller than 15 kD that could function directly as signaling molecules or as precursors for peptide hormones. Our study provides the basis for targeted reverse-genetic approaches aimed to identify important regulators of reproductive development in plants. Experimenter name = Lars Hennig Experimenter phone = +41 1 632 22 44 Experimenter fax = +41 1 632 10 44 Experimenter department = Plant Science C Experimenter institute = Institute of Plant Sciences and Zurich-Basel Experimenter address = Swiss Federal Institute of Technology Experimenter address = ETH Center Experimenter address = Zurich Experimenter zip/postal_code = CH-8092 Experimenter country = Switzerland Keywords: organism_part_comparison_design;
