A binding site for phosphoinositides described by multiscale simulations explains their modulation of voltage gated sodium channels

Voltage gated sodium channels (Nav) are membrane proteins which open to facilitate the inward flux of sodium ions into excitable cells. In response to stimuli, Nav channels transition from the resting, closed state to an open, conductive state, before rapidly inactivating. Dysregulation of this functional cycle due to mutations causes diseases including epilepsy, pain conditions and cardiac disorders, making Nav channels a significant pharmacological target. Phosphoinositides are important lipid cofactors for ion channel function. The phosphoinositide PI(4,5)P2 decreases Nav1.4 activity by increasing the difficulty of channel opening, accelerating fast inactivation and slowing recovery from fast inactivation. Using multiscale molecular dynamics simulations, we show that PI(4,5)P2 binds stably to inactivated Nav at a conserved site within the DIV S4-S5 linker, which couples the voltage sensing domain (VSD) to the pore. As the Nav C-terminal domain is proposed to also bind here during recovery from inactivation, we hypothesise that PI(4,5)P2 prolongs inactivation by competitively binding to this site. In atomistic simulations, PI(4,5)P2 reduces the mobility of both the DIV S4-S5 linker and the DIII-IV linker, responsible for fast inactivation, slowing the conformational changes required for the channel to recover to the resting state. We further show that in a resting state Nav model, phosphoinositides bind to VSD gating charges, which may anchor them and impede VSD activation. Our results provide a mechanism by which phosphoinositides alter the voltage dependence of activation and the rate of recovery from inactivation, an important step for the development of novel therapies to treat Nav-related diseases. SignificanceVoltage-gated sodium channels form pores in the membrane to mediate electrical activity in nerve and muscle cells. They play critical roles throughout the human body and their dysfunction leads to diseases including epilepsy, cardiac arrhythmias and pain disorders. Membrane lipids called phosphoinositides have recently been shown to reduce the activity of a voltage-gated sodium channel, but the molecular basis of this mechanism is not known. Here we use simulations to reveal where these lipids bind to the channels and how they reduce channel activity by making it harder for the pores to open and slower to subsequently recover to the closed resting state.