Structure and activity of SLAC1 channels for stomatal signaling in leaves

Abstract

Significance Stomatal openings in plant leaves admit CO2 and release water vapor between plant and atmosphere. Environmental stimuli such as darkness or dryness lead to closure, whereas light or low CO2 stimulate opening. Each stoma is defined by a pair of guard cells, and its pore aperture is controlled by ion channels that regulate the turgor pressure in these cells. SLAC1 anion channels are of central importance here, and environmental factors affect channel activity by controlling SLAC1 phosphorylation. Our studies define multiple sites of regulatory phosphorylation in the context of an atomic structure of the SLAC1 channel, providing a mechanistic understanding of the fine-tuning of channel activity and thereby of stomatal apertures in response to the environment. Stomata in leaves regulate gas exchange between the plant and its atmosphere. Various environmental stimuli elicit abscisic acid (ABA); ABA leads to phosphoactivation of slow anion channel 1 (SLAC1); SLAC1 activity reduces turgor pressure in aperture-defining guard cells; and stomatal closure ensues. We used electrophysiology for functional characterizations of Arabidopsis thaliana SLAC1 (AtSLAC1) and cryoelectron microscopy (cryo-EM) for structural analysis of Brachypodium distachyon SLAC1 (BdSLAC1), at 2.97-Å resolution. We identified 14 phosphorylation sites in AtSLAC1 and showed nearly 330-fold channel-activity enhancement with 4 to 6 of these phosphorylated. Seven SLAC1-conserved arginines are poised in BdSLAC1 for regulatory interaction with the N-terminal extension. This BdSLAC1 structure has its pores closed, in a basal state, spring loaded by phenylalanyl residues in high-energy conformations. SLAC1 phosphorylation fine-tunes an equilibrium between basal and activated SLAC1 trimers, thereby controlling the degree of stomatal opening.