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Significance Abscisic acid (ABA) is a phytohormone that plants utilize to coordinate responses to abiotic stress, modulate seed dormancy, and is central to plant development in several contexts. Chemicals that activate or block ABA signaling are useful as research tools and as potential agrochemical leads. Many successes have been reported for ABA activators (agonists), but existing ABA blockers (antagonists) are limited by modest in vivo activity. Here we report antabactin (ANT), a potent ABA blocker developed using “click chemistry”–based diversification of a known ABA activator. Structural studies reveal, ANT disrupts signaling by stabilizing ABA receptors in an unproductive form. ANT can accelerate seed germination in multiple species, making it a chemical tool for improving germination. 

Summary Abscisic acid (ABA) receptors belong to theSTARTdomain superfamily, which encompasses ligand‐binding proteins present in all kingdoms of life.STARTdomain proteins contain a central binding pocket that, depending on the protein, can couple ligand binding to catalytic, transport or signaling functions. In Arabidopsis, the best characterizedSTARTdomain proteins are the 14PYR/PYL/RCAR ABAreceptors, while the other members of the superfamily do not have assigned ligands. To address this, we used affinity purification of biotinylated proteins expressed transiently inNicotiana benthamianacoupled to untargetedLC‐MSto identify candidate binding ligands. We optimized this method usingABA–PYLinteractions and show thatABAco‐purifies with wild‐typePYL5 but not a binding site mutant. TheK_dofPYL5 forABAis 1.1 μm, which suggests that the method has sufficient sensitivity for many ligand–protein interactions. Using this method, we surveyed a set of 37STARTdomain‐related proteins, which resulted in the identification of ligands that co‐purified withMLBP1 (At4G01883) orMLP165 (At1G35260). Metabolite identification and the use of authentic standards revealed thatMLBP1 binds to monolinolenin, which we confirmed using recombinantMLBP1. Monolinolenin also co‐purified withMLBP1 purified from transgenic Arabidopsis, demonstrating that the interaction occurs in a native context. Thus, deployment of this relatively simple method allowed us to define a protein–metabolite interaction and better understand protein–ligand interactions in plants. 

Drought causes crop losses worldwide, and its impact is expected to increase as the world warms. This has motivated the development of small-molecule tools for mitigating the effects of drought on agriculture. We show here that current leads are limited by poor bioactivity in wheat, a widely grown staple crop, and in tomato. To address this limitation, we combined virtual screening, x-ray crystallography, and structure-guided design to develop opabactin (OP), an abscisic acid (ABA) mimic with up to an approximately sevenfold increase in receptor affinity relative to ABA and up to 10-fold greater activity in vivo. Studies inArabidopsis thalianareveal a role of the type III receptorPYRABACTIN RESISTANCE-LIKE 2for the antitranspirant efficacy of OP. Thus, virtual screening and structure-guided optimization yielded newly discovered agonists for manipulating crop abiotic stress tolerance and water use. 

Early abscisic acid signaling involves degradation of clade A protein phosphatases type 2C (PP2Cs) as a complementary mechanism to PYR/PYL/RCAR-mediated inhibition of PP2C activity. At later steps, ABA induces up-regulation of PP2C transcripts and protein levels as a negative feedback mechanism. Therefore, resetting of ABA signaling also requires PP2C degradation to avoid excessive ABA-induced accumulation of PP2Cs. It has been demonstrated that ABA induces the degradation of existing ABI1 and PP2CA through the PUB12/13 and RGLG1/5 E3 ligases, respectively. However, other unidentified E3 ligases are predicted to regulate protein stability of clade A PP2Cs as well. In this work, we identified BTB/POZ AND MATH DOMAIN proteins (BPMs), substrate adaptors of the multimeric cullin3 (CUL3)-RING-based E3 ligases (CRL3s), as PP2CA-interacting proteins. BPM3 and BPM5 interact in the nucleus with PP2CA as well as with ABI1, ABI2, and HAB1. BPM3 and BPM5 accelerate the turnover of PP2Cs in an ABA-dependent manner and their overexpression leads to enhanced ABA sensitivity, whereas bpm3 bpm5 plants show increased accumulation of PP2CA, ABI1 and HAB1, which leads to global diminished ABA sensitivity. Using biochemical and genetic assays, we demonstrated that ubiquitination of PP2CA depends on BPM function. Given the formation of receptor-ABA-phosphatase ternary complexes is markedly affected by the abundance of protein components and ABA concentration, we reveal that BPMs and multimeric CRL3 E3 ligases are important modulators of PP2C coreceptor levels to regulate early ABA signaling as well as the later desensitizing-resetting steps.