Cooperative Mechanisms Of Calcium-dependent Gating In Calcium-activated Chloride Channels

Calcium-activated chloride channel?CaCCs?is widely distributed in various tissues of spinal and non-spinal animals.CaCCs involves in many of the physiological processes,such as epithelial secretion,sensory transduction and adaptation,regulation of smooth muscle contraction,control of neuronal and cardiac excitability,and nociception.The gating of CaCCs is dual regulated by calcium and voltage.Study on CaCCs?TMEM16A?channel gating mechanism not only helps to understand its physiological effect,but also sheds lights on drug design targeting at CaCCs.In this paper,we combined site-directed mutagenesis,patch clamp with confocal laser scanning microscope and studied the Ca²⁺ dependent gating mechanism of TMEM16 A channels.The main contents are as follows:?1?.There are two Ca²⁺ sensitive domains with different calcium affinity in TMEM16 A.One with lower Ca²⁺ affinity locates in the first intracellular loop,another with higher Ca²⁺ affinity is in the center of the sixth,seventh and eighth transmembrane helices.There is a cooperation between the two Ca²⁺ sensitive domains.?2?.The size of side chains of Ca²⁺ binding residues in the first intracellular loop play key roles in Ca²⁺ dependent activation of CaCCs.?3?.The divalent cations,Ca²⁺ and Sr²⁺,can activate TMEM16 A channels and may share common mechanism to activate TMEM16 A.?4?.Chitosan oligosaccharide?COS?can directly activate TMEM16 A in a Ca²⁺ non-dependent way.The monomers with different degree of polymerization of COS shows different binding affinity.The EC50 of different monomers for CaCCs are 35.72±3.633 ?M,29.03±3.696 ?M,36.18±6.036 ?M,38.43±2.916 ?M,43.90±4.985 ?M,respectively.?5?.The chitosan oligosaccharide activates TMEM16 A channel by different mechanism of calcium.Here,we explored the gating mechanism of calcium activated CaCCs and drew some conclusions.However,there are still open questions as follows: the dynamics of calcium dependent activation of CaCCs,the structural basis of chitosan oligosaccharide binding with CaCCs and its dynamical properties.