Oxidation modulates LINGO2-induced inactivation of large conductance, Ca2+-activated potassium channels

Ca2+ and voltage-activated K+ (BK) channels are ubiquitous ion channels that can be modulated by accessory proteins, including beta, gamma, and LINGO1 BK subunits. In this study, we utilized a combination of site-directed mutagenesis, patch clamp electrophysiology, and molecular modeling to investigate if the biophysical properties of BK currents were affected by coexpression of LINGO2 and to examine how they are regulated by oxidation. We demonstrate that LINGO2 is a regulator of BK channels, since its coexpression with BK channels yields rapid inactivating currents, the activation of which is shifted -30 mV compared to that of BK alpha currents. Furthermore, we show the oxidation of BK:LINGO2 currents (by exposure to epifluorescence illumination or chloramine-T) abolished inac-tivation. The effect of illumination depended on the presence of GFP, suggesting that it released free radicals which oxidized cysteine or methionine residues. In addition, the oxidation ef-fects were resistant to treatment with the cysteine-specific reducing agent DTT, suggesting that methionine rather than cysteine residues may be involved. Our data with synthetic LINGO2 tail peptides further demonstrate that the rate of inactivation was slowed when residues M603 or M605 were oxidized, and practically abolished when both were oxidized. Taken together, these data demonstrate that both methionine residues in the LINGO2 tail mediate the effect of oxidation on BK:LINGO2 channels. Our molecular modeling suggests that methionine oxidation reduces the lipophilicity of the tail, thus preventing it from occluding the pore of the BK channel.

Automated summary

In this study, the researchers found that LINGO2 plays a role in modulating BK channels. Coexpression of LINGO2 and BK channels resulted in rapid inactivating currents with shifted activation compared to BK alpha currents. The oxidation of BK:LINGO2 currents abolished inactivation, and this effect was resistant to treatment with a reducing agent specific for cysteine residues. The molecular modeling further revealed that methionine oxidation reduces the lipophilicity of the LINGO2 tail, preventing it from occluding the pore of BK channels.