MscS inactivation and recovery are slow voltage-dependent processes sensitive to interactions with lipids

Abstract

Mechanosensitive channel MscS, the major bacterial osmolyte release valve, shows a characteristic adaptive behavior. With a sharp onset of activating tension, the channel population readily opens, but under prolonged action of moderate near-threshold tension, it inactivates. The inactivated state is non-conductive and tension-insensitive, which suggests that the gate gets uncoupled from the lipid-facing domains. The kinetic rates for tension-driven opening-closing transitions are 4-6 orders of magnitude higher than the rates for inactivation and recovery. Here we show that inactivation is augmented and recovery is slowed down by depolarization. Hyperpolarization, conversely, impedes inactivation and speeds up recovery. We then address the question of whether protein-lipid interactions may set the rates and influence voltage dependence of inactivation and recovery. Mutations of conserved arginines 46 and 74 anchoring the lipid-facing helices to the inner membrane leaflet to tryptophans do not change the closing transitions, but instead change the kinetics of both inactivation and recovery and essentially eliminate their voltage-dependence. Uncharged polar substitutions (S or Q) for these anchors produce functional channels but increase the inactivation and reduce the recovery rates. The data suggest that it is not the activation and closing transitions, but rather the inactivation and recovery pathways that involve substantial rearrangements of the protein-lipid boundary associated with the separation of the lipid-facing helices from the gate. The discovery that hyperpolarization robustly assists MscS recovery indicates that membrane potential can regulate osmolyte release valves by putting them either on the ‘ready’ or ‘standby’ mode depending on the cell’s metabolic state.

Polymer-extracted structure of the mechanosensitive channel MscS reveals the role of protein-lipid interactions in the gating cycle

“…The opening and closing transitions were found to be essentially frictionless, non-perturbing, and non-dissipative (Çetiner et al, 2023), indicating that lipid re-arrangement is not possible. The slow inactivation and recovery transitions on the other hand are strongly voltage-dependent, which suggests that inactivation is specifically associated with the rearrangement of the protein-lipid boundary (Britt et al, 2024).…”

“…The opening-closing transitions are sharply tension-dependent and can be very fast (Boer et al, 2011). In contrast, the inactivation and recovery transitions are slow and sensitive to the lipid environment and the nature of lipid-anchoring sidechains (Britt et al, 2024).…”

Mechanisms of PIEZO Channel Inactivation

“…Inactivation is a general property of most types of ion channels enabling filtering out repetitive or prolonged stimuli by blocking the flow of ions via a mechanism other than the closing of the channel [ 13 , 14 , 15 , 16 , 17 ]. Structural studies reveal diverse mechanisms for inactivation, while most of the inactivation processes involve conformational changes in one or multiple inactivation gates located within the pore region.…”