Molecular insights into the activation mechanism of the slow anion channel AtSLAH3

The plant slow anion channel SLAH3 regulates essential physiological processes, including stomatal closure and stress responses, yet the structural mechanism underlying its activation remains poorly understood. Here, we investigated the activation mechanism of SLAH3 by determining cryo-electron microscopy (cryo-EM) structures of the wild-type (WT) channel and gain-of-function mutants identified through a targeted mutagenesis screen of pore-lining residues. Notably, the L400A mutant was captured in a dilated, open-like conformation, in contrast to the auto-inhibited WT channel. Structural comparisons revealed concerted rearrangements of the pore-lining transmembrane helices and intracellular regulatory elements. The observations point to an activation model in which displacement of the intracellular auto-inhibitory module relieves pore occlusion, triggering conformational changes that propagate from the intracellular loop to pore-lining segments, ultimately widening the entrance for anion transport. Therefore, our study provides a structural framework for understanding SLAH3 regulation and advances our understanding of nitrate signaling and stress adaptation in plants.