Synthetic Agonists of Abscisic Acid Receptors and Their Metabolomic Effects on Plants

Land plants respond to multiple abiotic stresses, including drought, through the signaling cascade induced by the phytohormone abscisic acid (ABA). Endogenous production, or externally applied ABA has a major function of eliciting guard cell closure, ultimately lowering transpiration and increasing drought tolerance in plants. Accordingly, the mechanisms of action in which ABA facilitates this response have been popular targets for agricultural research and applications. However, ABA has a multitude of responses aside from stomatal closure that are important for plant's survival to abiotic stress. In response to limited water availability, the phytohormone is known to maintain primary root growth while decreasing shoot growth, increase osmolyte accumulation, inhibit seed germination, and is involved in substantial crosstalk with other phytohormones. These responses are dependent on a core ABA signaling pathway comprised of three components: the ABA receptors known as the PYRABACTIN RESISTANT/PYRABACTIN RESISTANT-LIKE/REGULATORY COMPONENT OF ABA RECEPTORs (PYR/PYL/RCARs), the clade A protein phosphatase 2Cs (PP2Cs), and the sucrose nonfermenting related subfamily 2 (SnRK2s).The ABA receptors are the first interactors with ABA within the pathway, and cluster into three clades (I, II, III) based on sequence identity. Additionally, these clades can be further grouped based on oligomeric preference; clades I and II are preferentially monomeric, while clade III is preferentially dimeric. While some research has been done on non-redundant functions of the PYL proteins, many of the ABA responses remain uncharacterized with respect to the differential contributions of the different receptors. Additionally, most of the published ABA-receptor agonists are either direct ABA analogs displaying pan-agonist activity, or are primarily active only on the dimeric subgroup of the receptors. It would thus be potentially beneficial to develop agonists that show preferential activation of the monomeric receptors, allowing for temporal activation and subsequent analysis of their biological relevance.In order to identify compounds with novel selectivities, preferably on the monomeric receptors, I used high-throughput virtual screening to evaluate compounds unbiased to previous, active scaffolds. This resulted in a series of chemically similar hits which showed potent activity on the monomeric receptors, and translated to some in vivo responses. This potent, monomeric-specific scaffold was optimized using structure-aided design, improving the in vivo responses. Using this probe molecule I provide data that suggest that monomeric and dimeric ABA receptors may differentially control metabolomic and transcriptional responses, but are adequate in seed germination inhibition and primary root elongation