Discovery and Structural Basis of Zoledronic Acid as a Potent SPS-Targeting Herbicide

Solanesyl diphosphate synthase (SPS) is essential for plastoquinone-9 (PQ-9) biosynthesis and has emerged as a promising target for herbicide discovery. The commercial SPS herbicides bind to the dimer interface of the protein; however, substrate-competitive SPS inhibitors are still rare, particularly in agronomically relevant weed and crop orthologs. Here, we show that zoledronic acid (ZOL) inhibits Arabidopsis thaliana SPS1 with an IC₅₀ value of 2.55 μM in an Mg²⁺-dependent manner. Crystal structures of ZOL-bound SPS from the weed Aegilops tauschii (AeSPS) and the crop Sorghum bicolor (SbSPS) revealed that the bisphosphonate moiety is anchored in the catalytic pocket through an Mg²⁺-water coordination network and conserved aspartate-rich motifs, showing substrate-competitive inhibition. ZOL induced chlorosis, root-growth inhibition, and reactive oxygen species (ROS) accumulation in A. thaliana. Transcriptomic analyses further showed that ZOL and aclonifen affected shared antioxidant-related processes, but with markedly different response magnitudes. ZOL showed good herbicidal activity against S. viridis, D. sanguinalis, V. persica, and C. album at 1000 g a.i./ha. Together, this work establishes ZOL as a mechanistically distinct SPS inhibitor and provides a structural basis for the development of competitive SPS-inhibiting herbicides.