Structure-Function of TMEM16 Ion Channels and Lipid Scramblases

Abstract

The TMEM16 protein family comprises two novel classes of structurally conserved but functionally distinct membrane transporters that function as Ca²⁺-dependent Cl^- channels (CaCCs) or dual functional Ca²⁺-dependent ion channels and phospholipid scramblases. Extensive functional and structural studies have advanced our understanding of TMEM16 molecular mechanisms and physiological functions. TMEM16A and TMEM16B CaCCs control transepithelial fluid transport, smooth muscle contraction, and neuronal excitability, whereas TMEM16 phospholipid scramblases mediate the flip-flop of phospholipids across the membrane to allow phosphatidylserine externalization, which is essential in a plethora of important processes such as blood coagulation, bone development, and viral and cell fusion. In this chapter, we summarize the major methods in studying TMEM16 ion channels and scramblases and then focus on the current mechanistic understanding of TMEM16 Ca²⁺- and voltage-dependent channel gating as well as their ion and phospholipid permeation.

Keywords: Anoctamin; CaCC; CaPLSase; Gating; Permeation; Scramblase; TMEM16.

Fig. 6.1

TMEM16 protein family. (a) TMEM16 proteins can function as Ca²⁺-activated chloride channels (CaCCs) or Ca²⁺-dependent phospholipid scramblases (CaPLSases). (b) TMEM16 protein family with diverse functionality. Note that the yeast IST2 and C. elegans ANOH-1 and −2 have not been assigned as CaCCs or CaPLSases. (c) TMEM16A CaCC and TMEM16F small-conductance, Ca²⁺-activated nonselective channel (SCAN) at different Ca²⁺ concentrations [2]. (d) A fluorescence assay to monitor CaPLSase activity

Fig. 6.2

Structural basis of TMEM16 proteins. (a) Topology of TMEM16 proteins. (b) Structure of the Ca²⁺-bound fungal nhTMEM16 (PDB 4WIS). (c) Structure of the Ca²⁺-bound mouse TMEM16A showing its overall architecture (left), Ca²⁺ coordinating residues (middle), and conformational change of TM6 upon Ca²⁺ binding. (d, e) Residues that are implicated in controlling phospholipid scrambling in the fungal nhTMEM16 (PDB 4WIS) and mouse TMEM16F (PDB 6QP6). (f) The lipid permeation pathway of the human TMEM16K as viewed from the ER lumen in its closed state (PDB 6R6X) and open state (PDB 5OC9)

Fig. 6.3

Mechanistic models of phospholipid and ion permeation by dual-functional TMEM16 CaPLSases/channels. The proposed models for lipidic pore (a), out-of-the-groove (b), and alternating pore-cavity (c) mechanisms are shown