Despite the essential role of plants to support animal life on earth, our understanding of developmental processes in plants is still fragmentary. Similar to animals, plants use a small number of signaling systems, such as the signaling circuitries triggered by the plant hormones cytokinin and auxin, for many and diverse developmental processes. Typically, plant hormones are small organic molecules, elusive to in situ detection. Therefore, reporter genes, which label all signal-receiving cells in vivo, have proved powerful tools to dissect fundamental developmental processes. To analyze a long-postulated function of cytokinin during embryonic pattern formation, I have constructed a novel synthetic sensor that labels the cytokinin-signaling cells in planta. Combined with targeted genetic manipulations I have discovered an antagonistic relation between auxin and cytokinin that is required for embryonic stem-cell specification. The aim of this project is to characterize the genetic regulatory network downstream of cytokinin signaling that governs embryonic stem-cell specification. In addition, taking advantage of the unique tools I have generated, I novel cytokinin functions during female gametophyte development will be analyzed. The proposed studies will be conducted in Arabidospsis thaliana, a dicot reference plant that offers excellent genetic, genomic, imaging, and tissue culture amenability. In short, the specific aims are to:(a) Identify and characterize the cytokinin target genes in the nascent embryonic root meristem(b) Describe the role of cytokinin signaling in gametophyte developmentData gleaned from this studies will afford a point of reference and toolbox to study cytokinin signaling in other developmental contexts, such as leaf senescence, stem-cell pool maintenance in the shoot, shoot and root branching, grain size control, defense against pathogens, just to name a few. Given the conservation of the cytokinin signaling mechanism in higher plants, the value of the proposed experiments will extend beyond the reference plant Arabidopsis thaliana to include crop plants such as rice, wheat, or maize.