BPS2026 - PIP2 activates SK channels by a similar mechanism across all known isoforms

Small conductance Ca 2+ -activated K + (SK or KCa2) channels are unique since they are gated solely by intracellular Ca 2+ . They are known to play critical roles in atrial repolarization and are upregulated in atrial arrhythmias and heart failure, highlighting their clinical significance. Previous studies demonstrated that phosphatidylinositol 4,5-bisphosphate (PIP2) in the plasma membrane contributed to the channel activation, and a potential activation binding was proposed in SK2, yet the mechanistic role of PIP2 regulation of cardiac SK channels remains incompletely understood. Recent cryogenic-electron microscopic structures of the SK4-calmodulin (CaM) complex (PDB: 6CNM, 6CNN, and 6CNO) and SK2-CaM complex (PDB: 8V2G) enabled us to generate nearly full-length SK3-CaM homology models in various conformational states. We then performed several replicas of 1-2-μs-long unbiased molecular dynamics (MD) simulations of the SK3-CaM models in a hydrated 1-palmitoyl-2-oleoylphosphatidylcholine (POPC):PIP2 lipid membrane. We started simulations with 1 PIP2 molecule bound to each of the 4 activation binding sites to understand its effect in a saturated environment. Our computational study revealed important structural information of how the channel is regulated by PIP2. Specifically, amino acid residue R544, localized in the S6 transmembrane segment in proximity to the intracellular hydrophobic gate, forms a salt bridge with S6 residue E547 from the adjacent subunit. Both residues are conserved in all known isoforms of SK channels. Our findings reinforce our previous theory that PIP2 binding to R544 or equivalent residue in all SK isoforms disrupts the R544:E547 salt bridge increasing the flexibility of the pore-lining segment S6 and contributing to the activation of the channel. Additionally, we discovered that the intracellular gate residue V539 sidechains cannot adopt parallel orientation to each other for K + ions to enter the pore, which widens our understanding of ion channel conduction mechanisms.