DPP6A Confers Redox Sensitivity to Kv4 Channel Inactivation

Abstract

Reactive oxygen species (ROS) are important modulators of excitability and may play a critical role in the etiology of many neurodegenerative disorders. Previous studies have shown that the fast N-type inactivation of Kv1.4 channels is suppressed by ROS-mediated cysteine oxidation whereas Kv4 channel N-type inactivation is not. However, in native neurons Kv4 channel subunits form large macromolecular complexes with KChIP and DPL proteins, and fast N-type inactivation is conferred to Kv4 channel complexes by a specific isoform of DPL proteins, either DPP10a or DPP6a. To investigate whether the DPP6a-mediated fast inactivation is regulated by ROS, tert- butyl hydroperoxide (tBHP) was applied to oocytes expressing Kv4.2+KChIP3a+DPP6a channels. tBHP (1 mM) application dramatically increases the peak current amplitude by ∼ 44% and slowed inactivation kinetics. The effects of tBHP are reversed by the reducing agent dithiothreitol (DTT, 10 mM). In contrast, ternary complex channels containing another DPP6 isoform (DPP6K) are not affected by tBHP, indicating the importance of the DPP6a variable N-terminal domain for the tBHP effect. Alignment of N-terminal sequences from DPP6a and DPP10a with Kv1.4 reveals a common cysteine residue in position 13 (Cys-13), which in Kv1.4 is critical for the redox-regulation of N-type inactivation. Substituting Cys-13 of DPP6a with serine (DPP6a/C13S) results in a loss of regulation by tBHP, consistent with a similar critical role for DPP6a Cys-13. To test if other cysteines in the channel are also required for this regulation, we switched to mutant Kv4.1 (C11xA) and KChIP3a (KChIP3a/del2-59) constructs that remove most intracellular cysteine residues. Channels composed of C11xA, KChIP3a/del2-59, and normal DPP6a show disrupted regulation by tBHP, suggesting that ROS likely induces an intersubunit disulfide linkage to regulate DPP6a-mediated N-type inactivation. This work was supported by a grant from the National Institute of Health (R01 GM090029).