Structure reveals a regulation mechanism of plant outward-rectifying K⁺ channel GORK by structural rearrangements in the CNBD-Ankyrin bridge.

Abstract

Guard cells, which regulate stomatal apertures in plants, possess a sophisticated mechanism for regulating turgor pressure. The outward-rectifying "K⁺_out" channel GORK, expressed in guard cells of the plant <i>Arabidopsis thaliana</i>, is a central component that promotes stomatal closure by releasing K⁺ to the extracellular space, thereby lowering turgor pressure. To date, the structural basis underlying the regulation of the K⁺ transport activity of GORK is unclear. Using cryo-EM, we determined the structures of the GORK outward-rectifying K⁺ channel with a resolution of 3.16 to 3.27 Å in five distinct conformations that differ significantly in their C-terminal cyclic nucleotide binding domain (CNBD) and ankyrin repeat (ANK) domain. The C-linker connects the transmembrane domains to the C-terminal domains, i.e., CNBD, CNBD-Ankyrin bridge, and ANK. The structural changes and interactions in the C-linker determine whether the closed state of GORK is closer to the preopen state or in a more removed state from the open state of the channel. In particular, interconversion in the short sequence within the CNBD-Ankyrin bridge plays a decisive role in this determination. This region forms an α-helix in the preopened state, while it adopts a nonhelical structure in further distant closed states. The dynamics of the cytosolic region strongly suggest that the K⁺ channel activity of GORK is regulated by cytosolic signaling factors during stomatal closure.