Calmodulin‐dependent activation of the epithelial calcium‐dependent chloride channel TMEM16A

Abstract

TMEM16A (anoctamin 1, Ano1), a member of a family of 10 homologous proteins, has been shown to form an essential component of Ca2+‐activated CU channels. TMEM16A‐null mice exhibit severe defects in epithelial transport along with tracheomalacia and death within 1 mo after birth. Despite its outstanding physiological significance, the mechanisms for activation of TMEM16A remain obscure. TMEM16A is activated on increase in intracellular Ca2+, but it is unclear whether Ca2+ binds directly to the channel or whether additional components are required. We demonstrate that TMEM16A is strictly membrane localized and requires cytoskeletal interactions to be fully activated. Despite the need for cytosolic ATP for full activation, phosphorylation by protein kinases is not required. In contrast, the Ca2+ binding protein calmodulin appears indispensable and interacts physically with TMEM16A. Openers of small‐ and intermediate‐conductance Ca2+‐activated potassium channels known to interact with calmodulin, such as 1‐EBIO, DCEBIO, or riluzole, also activated TMEM16A. These results reinforce the use of these compounds for activation of electrolyte secretion in diseases such as cystic fibrosis.—Tian, Y., Kongsuphol, P., Hug, M., Ousingsawat, J., Witzgall, R., Schreiber, R., Kunzelmann, K. Calmodulin‐dependent activation of the epithelial calcium‐dependent chloride channel TMEM16A. FASEB J. 25, 1058–1068 (2011). http://www.fasebj.org

Preassociated apocalmodulin mediates Ca 2+ -dependent sensitization of activation and inactivation of TMEM16A/16B Ca 2+ -gated Cl − channels

“…Additionally, whether TMEM16A binds Ca 2+ -CaM is controversial-some biochemical pull-down studies support an interaction (31,32), whereas others do not (40). Notably, pull-down methods can be notoriously inconsistent with regards to confirming apoCaM/Ca 2+ -CaM association with ion channels even when these are functionally known to occur (35,41).…”

Anoctamin 1 (Tmem16A) Ca 2 + -activated chloride channel stoichiometrically interacts with an ezrin–radixin–moesin network

“…The actin cytoskeleton and its associated regulatory proteins could be implicated in Ano1 function by modulating Ano1 channel gating, directing the trafficking Ano1 to the apical membrane, or assembling Ano1 into signaling networks. It has been reported that both depolymerization of the actin cytoskeleton with cytochalasin-D and stabilization of the actin network with phalloidin suppresses Ano1 currents (42). A number of ion channels have been shown to interact with the actin cytoskeleton (43).…”

Role of volume-regulated and calcium-activated anion channels in cell volume homeostasis, cancer and drug resistance

“…45 They show outward-rectification and a similar permeability sequence as described for VRAC (SCN ->I ->NO 3 ->Br ->Cl ->gluconate), they are activated by osmotic cell swelling in the presence of extracellular Ca 2C40 and requires cellular ATP. 22,43,44,46 However, VRAC and ANO1/6 activities can be distinguished by (i) their sensitivity to strongly buffered intracellular Ca 2C , i.e. VRAC can be activated whereas ANO1 cannot, (ii) activation/deactivation at positive potentials, i.e., VRAC deactivates 9,41,47 whereas ANO1/6 activates ( Fig.…”

“…126 Two putative CaM binding sites were identified on ANO1, which led to the proposal that Ca 2C sensitivity is mediated by calmodulin. 46,127 Recent papers did however present strong evidence against the possible role of calmodulin. 128,129 The most direct evidence that the Ca 2C sensitivity is intrinsic and not dependent on calmodulin comes from the observation that purified ANO1 reconstituted in liposomes recapitulates the functional properties of ANO1.…”