Dataset Information

Epithelial Chloride Transport by CFTR Requires TMEM16A.

ABSTRACT: Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is the secretory chloride/bicarbonate channel in airways and intestine that is activated through ATP binding and phosphorylation by protein kinase A, but fails to operate in cystic fibrosis (CF). TMEM16A (also known as anoctamin 1, ANO1) is thought to function as the Ca2+ activated secretory chloride channel independent of CFTR. Here we report that tissue specific knockout of the TMEM16A gene in mouse intestine and airways not only eliminates Ca2+-activated Cl- currents, but unexpectedly also abrogates CFTR-mediated Cl- secretion and completely abolishes cAMP-activated whole cell currents. The data demonstrate fundamentally new roles of TMEM16A in differentiated epithelial cells: TMEM16A provides a mechanism for enhanced ER Ca2+ store release, possibly engaging Store Operated cAMP Signaling (SOcAMPS) and activating Ca2+ regulated adenylyl cyclases. TMEM16A is shown to be essential for proper activation and membrane expression of CFTR. This intimate regulatory relationship is the cause for the functional overlap of CFTR and Ca2+-dependent chloride transport.

SUBMITTER: Benedetto R

PROVIDER: S-EPMC5622110 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Epithelial Chloride Transport by CFTR Requires TMEM16A.

Benedetto Roberta R Ousingsawat Jiraporn J Wanitchakool Podchanart P Zhang Yong Y Holtzman Michael J MJ Amaral Margarida M Rock Jason R JR Schreiber Rainer R Kunzelmann Karl K

Scientific reports 20170929 1

Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is the secretory chloride/bicarbonate channel in airways and intestine that is activated through ATP binding and phosphorylation by protein kinase A, but fails to operate in cystic fibrosis (CF). TMEM16A (also known as anoctamin 1, ANO1) is thought to function as the Ca<sup>2+</sup> activated secretory chloride channel independent of CFTR. Here we report that tissue specific knockout of the TMEM16A gene in mouse intestine and airways not ...[more]

PMID: 28963502

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Project description:Niclosamide and benzbromarone have been described as inhibitors of the calcium activated chloride channel, TMEM16A, and on this basis have been considered and tested as clinical candidates for the treatment of airway diseases. However, both compounds have previously demonstrated activity on a range of additional biological targets and it is unclear from the literature to what extent any activity on TMEM16A may contribute to efficacy in these models of airway disease. The aim of the present study was therefore to examine the pharmacology and selectivity of these clinical candidates together with a structurally unrelated TMEM16A blocker, Ani9, in a range of functional assays to better appreciate the putative role of TMEM16A in the regulation of both epithelial ion transport and the development of an airway epithelial mucus secretory phenoptype. Benzbromarone and Ani9 both attenuated recombinant TMEM16A activity in patch clamp studies, whereas in contrast, niclosamide induced a paradoxical potentiation of the TMEM16A-mediated current. Niclosamide and benzbromarone were also demonstrated to attenuate receptor-dependent increases in intracellular Ca2+ levels ([Ca2+]i) which likely contributed to their concomitant attenuation of the Ca2+-stimulated short-circuit current responses of FRT-TMEM16A and primary human bronchial epithelial (HBE) cells. In contrast, Ani9 attenuated the Ca2+-stimulated short-circuit current responses of both cell systems without influencing [Ca2+]i which supports a true channel blocking mechanism for this compound. Additional studies using HBE cells revealed effects of both niclosamide and benzbromarone on global ion transport processes (absorptive and secretory) as well as signs of toxicity (elevated LDH levels, loss of transepithelial resistance) that were not shared by Ani9. Ani9 also failed to influence the IL-13 induced differentiation of HBE towards a goblet cell rich, mucus hypersecreting epithelium, whereas niclosamide and benzbromarone attenuated numbers of both goblet and multiciliated cells, that would be consistent with cellular toxicity. Together these data challenge the description of niclosamide as a TMEM16A blocker and illustrate a range of off-target effects of both niclosamide and benzbromarone which may contribute to the reported activity in models of airway function.

Dataset Information

Epithelial Chloride Transport by CFTR Requires TMEM16A.

Publications

Epithelial Chloride Transport by CFTR Requires TMEM16A.

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