A network of phosphatidylinositol 4,5-bisphosphate binding sites regulates gating of the Ca ²⁺ -activated Cl ⁻ channel ANO1 (TMEM16A)

Significance Membrane proteins dwell in a sea of phospholipids that not only structurally stabilize the proteins by providing a hydrophobic environment for their transmembrane segments but also dynamically regulate protein function. While many cation channels are known to be regulated by phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 ], relatively little is known about anion channel regulation by phosphoinositides. Using a combination of patch-clamp electrophysiology and atomistic molecular-dynamics simulations, we have identified several PI(4,5)P 2 binding sites in ANO1 (TMEM16A), a Cl − channel that performs myriad physiological functions from epithelial fluid secretion to regulation of electrical excitability. These binding sites form a band at the cytosolic interface of the membrane that we propose constitute a network to dynamically regulate this highly allosteric protein.