A Minimal Cysteine Motif Required to Activate the SKOR K+ Channel of Arabidopsis by the Reactive Oxygen Species H2O2*

Abstract

Reactive oxygen species (ROS) are essential for development and stress signaling in plants. They Reactive oxygen species (ROS) 3 are major signals in virtually every aspect of eukaryotic cell biology (1, 2). In animals, they are important regulators of cell division and muscle contraction among others (3), and in plants they are essential for development (4) and stress signaling, including drought (5, 6) and defense against pathogens (7). Within proteins, ROS target a small subset of amino acids, notably cysteine (Cys) residues, chemically modifying these amino acids and thereby altering protein structure and function (8). Nonetheless, the targeted residues and associated motifs are often poorly defined, the effects wide ranging and protein-specific, thus confounding molecular analyses. Indeed, among the few well documented examples, ROS modifications in mammalian ryanodine-receptor and BK channels have different consequences depending on the positions of the residues targeted (3, 9).For development and signaling in plants, the activities of membrane ion channels are essential, often including their regulation by ROS. For example, ROS affect non-selective cation channels during Fucus development (10); they regulate Ca 2ϩ channels and Ca 2ϩ -based signaling (5) as well as voltage-sensitive K ϩ channels of guard cells (11) that are important for stomatal movement; they contribute in responses to drought and pathogen defense (12); and they have been implicated in targeting K ϩ efflux during programmed cell death (13). However, until now virtually nothing has been known of the molecular targets for ROS in plants, nor has a site of action been identified with an ion channel protein.SKOR is one of two K ϩ channels found in Arabidopsis thaliana that rectify strongly outward, thereby mediating K ϩ efflux from the cell. It is expressed within the xylem parenchyma of the root where it facilitates K ϩ loading into the xylem (14), thereby contributing directly to K ϩ homeostasis and indirectly, through charge balance, to the transport of other solutes throughout the plant (12,15). Like other members of the Kvlike channel superfamily (16), the functional channel assembles from four monomers with each SKOR monomer incorporating six transmembrane ␣-helices. The first four ␣-helices form a positively charged voltage-sensor complex or "paddle" that moves within the membrane in response to voltage and couples this movement to the channel gate. The fifth and sixth ␣-helices line the aqueous pore through the membrane and assemble in a diaphragm or "gate" at the inner membrane surface, which opens/closes to regulate ion flux through the channel. Here we report that K ϩ current through the heterologously expressed SKOR is modulated by H 2 O 2 , thereby identifying the K ϩ channel as a potential target for ROS, and we show that a single Cys within the voltage sensor complex is essential for its ROS sen-

ROS, Calcium, and Electric Signals: Key Mediators of Rapid Systemic Signaling in Plants

“…Allen et al, 2000;Foreman et al, 2003;Demidchik et al, 2007) and molecular levels (e.g. the Ca 2+ -permeable Stelar K + Outward Rectifier or Annexin1; Garcia-Mata et al, 2010;Richards et al, 2014; Fig. 1), although to date the relevant channels related to systemic signaling have not been identified.…”

Intracellular Redox Compartmentation and ROS-Related Communication in Regulation and Signaling

“…However, although the details of the signaling pathways are being elucidated (Wrzaczek et al, 2015), information on the identity of the initial targets that are affected by ROS is still relatively scarce. These targets, which are presumably subject to ROS-induced oxidation, may include lipids or sensitive Cys residues on proteins such as ion channels (Garcia-Mata et al, 2010).…”

Arabidopsis Annexin1 Mediates the Radical-Activated Plasma Membrane Ca2+- and K+-Permeable Conductance in Root Cells

“…The molecular identities of ROS-regulated Ca 2+ -permeable channels in plants have remained obscure, despite their manifest significance in growth, adaptation, and defense (Dodd et al, 2010). The Ca 2+ -permeable Stelar K + Outward K + Rectifier PM channel (Gaymard et al, 1998) has been shown to be activated by hydrogen peroxide in a heterologous expression system (Garcia-Mata et al, 2010), but the effects of ROS on its Ca 2+ permeation, ability to affect [Ca 2+ ] cyt , and where it lies in relation to NADPH oxidase activity are unknown. Maize ANN33/ANN35 have previously been shown to form a hyperpolarization-activated macroscopic Ca 2+ conductance in PLBs containing malondialdehyde (to mimic peroxidized PM; Laohavisit et al, 2010).…”