9722774 Taylor Using mutants that are unable to synthesize flavonols preliminary research has established an essential role of flavanols in reproduction. Although mutant, flavanol-deficient maize and petunia pollen is unable to germinate or produce a functional pollen tube, the defect can be corrected by the exogenous application of low concentrations of kaempferol, a flavonol aglycone to an in vitro suspension of pollen. This results in a rapid (2-5 min) pollen tube outgrowth that is indistinguishable from that of wild-type pollen. The rapid response of pollen to low doses of kaempferol indicates that this flavanol has a role in signal transduction. In order to understand how flavanols signal pollen germination it is essential to identify the proteins that interact with kaempferol. By using highly specific affinity probes it is feasible to identify the proteins that bind to kaempferol and thereby elucidate the downstream components of the signal transduction pathway. To gain new expertise in the synthesis and use of labeled flavanols as tools the following objectives will be accomplished as career advancement activities; 1, To synthesize and evaluate radiolabeled photoaffinity probes for flavonol (kaempferol) receptors. 2, To prepare kaempferol-polymer conjugates for affinity chromatography. 3, To make caged kaempferol to study the intracellular sites of action. 4, To adapt the existing yeast two-hybrid system to detect protein-protein interactions mediated by kaempferol. In order to become proficient in use of bioactive small molecule chemistry in biological research I will conduct sabbatical research and training in the laboratory of Dr. Thomas Wandless of the Department of Chemistry at Stanford University. As a consequence of the sabbatical research and training I will have expanded opportunities to investigate biological problems involving the signaling and regulation of pathways by small bioactive molecules. Many small molecules signal the onset of biological processes. How variou s molecules induce new physiological and developmental activity remains some of the most important biological processes that must be elucidated. Flavanols are essential to growth of pollen tubes and reproductive activity in plants. Mutants unable to produce flavanols are arrested in pollen growth that can be almost immediately restored with the exposure of the mutant pollen to low doses of the flavanol kaempferol. This grant will fund sabbatical training and research devoted toward developing tools and techniques that will allow the PI to investigate how flavanols interact with specific cellular proteins that result in signaling reproductive growth in pollen. ***
