The genomes of vascular plants encode more than 1000 protein kinases. The long-term goal is to define all the regulatory phosphorylation sites in crop plant proteomes. Such sites represent potential control points for all aspects of plant growth, including responses to biotic and abiotic stress. The focus is to use bioinformatics to predict a subset of potentially important sites of phospho-regulation in the rice proteome, and then use a peptide-based experimental strategy to start validating those predictions. The four Specific Aims are: 1. Improve bioinformatics tools to identify motifs involved in phospho-signaling in plants. 2. Conduct a clustering analysis of all Ser and Thr containing sub-sequences in plant proteomes. 3. Design peptide array platform to profile kinase activities in vitro. 4. Identify peptides that can disrupt in vivo signaling pathways (more than 50 will be tested). The uniqueness and strength of the proposed research rests upon the underlying rationale that a computational comparison of two plant proteomes can be used as a 'noise filter' to identify important sequence motifs that are fundamental to signaling pathways conserved between monocots and dicots. This filtering provides the critical foundation for implementing 'peptide-disruptors' as a powerful new approach to understanding and manipulating the dynamics of the plant cell phosphorylome. DNA constructs for all genes found to encode a 'peptide disruptor' or function as a reporter for a particular kinase activity will be deposited at the ABRC at Ohio State University, and will be freely available.
