ライフサイエンスコーパス: TMEM16A

1 nnel (CaCC) anoctamin 1 (ANO1, also known as TMEM16A).

2 expressed, purified, and reconstituted human TMEM16A.

3 ng CFTR, ClC-2, ClC-3, CLCA, Bestrophin, and TMEM16A.

4 involve changes in pH and chloride flux via TMEM16A.

5 is necessary for apoCaM preassociation with TMEM16A.

6 Understanding how TMEM16A/16B are regulated by Ca(2+) is critical for defi

7 esults reveal multidimensional regulation of TMEM16A/16B by preassociated apoCaM and introduce ChIMP

8 ee calmodulin (apoCaM) is preassociated with TMEM16A/16B channel complexes.

9 ation of TMEM16A in HPV-positive HNSCC makes TMEM16A a poor therapeutic target in HPV-positive HNSCC,

10 Ano1 (Tmem16a), a Ca(2+)-activated Cl(-) channel, is an ion ch

11 Best2, 3, or 4, or transmembrane member 16A (TMEM16A), a member of another channel family.

12 We determined that Tmem16a, a member of an evolutionarily conserved family

13 but reduced the EC(50) for Ca(2+)-dependent TMEM16A activation.

14 The TMEM16A activators increased CaCC conductance in human s

15 Increase in TMEM16A activity occurred within minutes of exposure to

16 show that a low proton concentration reduces TMEM16A activity while maximum activation is obtained wh

17 CA1 as the first secreted direct modifier of TMEM16A activity, delineating a unique mechanism to incr

18 if impaired the ability of VWA to potentiate TMEM16A activity, suggesting that CLCA1-TMEM16A interact

19 50% CFTR activity and approximately 50% non-Tmem16a activity.

20 acidification and provide evidence that ANO1/TMEM16A acts to attenuate this pH shift.

21 TMEM16A also was required for peripheral blood vessel co

22 TMEM16A (also known as anoctamin 1, ANO1) is thought to

23 Transmembrane protein 16A (TMEM16A), also called anoctamin 1 (ANO1), is a calcium-a

24 ABSTRACT: Transmembrane protein 16A (TMEM16A), also known as ANO1, the pore-forming subunit o

25 on of recently discovered transmembrane 16A (TMEM16A), also termed Anoctamin 1, chloride (Cl(-)) chan

26 on of the calcium-activated chloride channel TMEM16A, also known as ANO1, in VSMCs, intermediate cell

27 TMEM16A alternative splicing provides a mechanism for tu

28 nd pharmacological tools to demonstrate that TMEM16A, an evolutionarily conserved calcium-activated c

29 lic acid arylamides (AACTs) as inhibitors of TMEM16A and analysis of 48 synthesized analogs (10ab-10b

30 Our results demonstrate that both Tmem16A and Best2 generate Ca(2+)-activated Cl(-) curren

31 Heterologous expression of Tmem16A and Best2 transcripts in HEK293 cells produced C

32 the Ca(2+)-gated Cl(-) currents generated by Tmem16A and Best2, members from two distinct families of

33 enous CLCA1 increases cell surface levels of TMEM16A and cellular binding experiments indicate CLCA1

34 However, both large and small BECs express TMEM16A and exhibit Ca(2+)-activated Cl(-) efflux in res

35 includes the Ca(2+)-activated Cl(-) channels TMEM16A and TMEM16B and a small-conductance, Ca(2+)-acti

36 TMEM16A and TMEM16B are calcium-activated chloride chann

37 TMEM16A and TMEM16B are molecular components of the phys

38 Having recently identified TMEM16A and TMEM16B as CaCCs, we further show that TMEM1

39 nexpected role for the putative pore-loop of TMEM16A and TMEM16B channels in the control of the whole

40 We show that TMEM16A and TMEM16B display fast and slow gating.

41 er chimeras within the putative pore-loop of TMEM16A and TMEM16B led to the identification of a regio

42 currents carried via transmembrane proteins TMEM16A and TMEM16B regulate diverse processes including

43 Calcium-activated chloride channels TMEM16A and TMEM16B support important physiological proc

44 Moreover, mouse TMEM16A and TMEM16B yield CaCCs in Axolotl oocytes and m

45 Interestingly, the tracheas of both the Tmem16a(-/-) and the CFTR(-/-) mice exhibited similar co

46 Although the founding members ANO1 (TMEM16A) and ANO2 (TMEM16B) are Ca(2+)-activated Cl(-) c

47 nductance regulator (CFTR), anoctamin-1(ANO1/TMEM16A) and the glycine receptor (GlyR), revealed that

48 c markers-namely, alpha-amylase, cystatin C, TMEM16A, and NKCC1.

49 Recently, TMEM16A (ANO1) and -B were shown to be critical componen

50 ly members to be functionally characterized, TMEM16A (ANO1) and TMEM16B (ANO2), form Ca(2+)-activated

51 TMEM16A (ANO1) functions as a calcium-activated chloride

52 TMEM16A (ANO1) is a calcium-activated chloride channel (

53 1 (CLCA1) modulates the activity of the CaCC TMEM16A/Anoctamin 1 (ANO1) by directly engaging the chan

54 cells in a paracrine fashion, and endogenous TMEM16A/Anoctamin1 conducts the currents.

55 cell-attached patches that were inhibited by TMEM16A antibodies and were of similar amplitude to reco

56 to those expressed in native cells, yet only Tmem16A appears to be a critical component of the acinar

57 Recent studies have unequivocally identified TMEM16A as a glandular epithelial CaCC.

58 Our data identify Tmem16a as a novel regulator of epithelial and smooth mu

59 The identification of ANO1/TMEM16A as the likely calcium-dependent chloride channel

60 as an expression system, we have identified TMEM16A as the Xenopus oocyte CaCC.

61 have developed antibodies specific for mouse TMEM16A, as evidenced by the absence of immunoreactivity

62 tes two distinct Ca(2+)-dependent effects on TMEM16A, as revealed by expression of dominant-negative

63 the chondrogenic mesenchyme does not express Tmem16a at any time, we propose that the cartilage ring

64 cid 623, which enables voltage activation of TMEM16A at non-saturating [Ca(2+) ]i .

65 nt with the hypothesis that CLCA1 stabilizes TMEM16A at the cell surface by preventing its internaliz

66 TR or the calcium-activated chloride channel TMEM16A attenuated the proinflammatory cytokines interle

67 but not P2Y receptor-mediated activation of TMEM16A attenuates IL-8 secretion in respiratory epithel

68 e pore-loop of TMEM16A with that of TMEM16B (TMEM16A-B channels) reduced the currents by approximatel

69 TMEM16F of the TMEM16 family that includes TMEM16A/B Ca(2+)-activated Cl(-) channels (CaCCs) is lin

70 TMEM16A belongs to a family of integral membrane protein

71 eported calcium-activated chloride channels (TMEM16A, Bestrophin-1, ClC2, and SLC26A9), both features

72 tance, providing further evidence for direct TMEM16A binding.

73 Direct and reversible inhibition of TMEM16A by 10bm was demonstrated by patch-clamp analysis

74 e intestinal calcium-activated Cl(-) channel TMEM16A by a voltage-independent inhibition mechanism wi

75 tion by the structure determination of mouse TMEM16A by cryo-electron microscopy and a complementary

76 tic potential of the selective activation of TMEM16A by the CLCA1 VWA domain in loss-of-function chlo

77 Noise analysis showed that protons regulate TMEM16A by tuning its open probability without modifying

78 on of two residues in the pore region of the TMEM16A Ca(2+)-activated Cl(-) channel convert it into a

79 voltage-dependent Ca(2+) channels (VDCCs) or TMEM16A Ca(2+)-activated Cl(-) channels significantly ch

80 efflux and osmotic cell shrinkage by opening TMEM16A Ca(2+)-activated Cl(-) channels.

81 compounds revealed compounds that activated TMEM16A CaCC conductance without increasing cytoplasmic

82 Our data suggest that TMEM16A CaCC-mediated Cl(-) secretion appears to be nece

83 TMEM16A-CaCC blockers, including those identified here,

84 ing intense interest in the mechanism behind TMEM16A-CaCC calcium-dependent gating, comprehensive sur

85 in the dimerization domain affect functional TMEM16A-CaCC channel expression, as expected from its cr

86 creen to identify small-molecule blockers of TMEM16A-CaCC channels.

87 versus anion selectivity of TMEM16F-SCAN and TMEM16A-CaCC channels.

88 We show that inhibition of TMEM16A-CaCC significantly impairs mucus secretion in pr

89 Furthermore, inhibition of TMEM16A-CaCC significantly reduces mouse and human ASM c

90 ed here permit pharmacological dissection of TMEM16A/CaCC function and are potential development cand

91 We conclude that TMEM16A carries nearly all CaCC current in salivary glan

92 ts in arterial myocytes that were blocked by TMEM16A channel inhibitory antibodies, RNAi-mediated sel

93 hibitory antibodies, RNAi-mediated selective TMEM16A channel knockdown, removal of extracellular calc

94 d by nonselective cation channels stimulates TMEM16A channels to induce myogenic constriction.

95 Furthermore, whole-cell currents mediated by TMEM16A channels were approximately six times larger tha

96 Membrane stretch activates arterial myocyte TMEM16A channels, leading to membrane depolarization and

97 and were of similar amplitude to recombinant TMEM16A channels.

98 losing M-type potassium channels and opening TMEM16A chloride channels, resulting in the production o

99 lecule TMEM16A inhibitors that fully blocked TMEM16A chloride current with an IC(50) < 10 muM, withou

100 ivators were identified that fully inhibited TMEM16A Cl(-) conductance, providing further evidence fo

101 intraluminal pH and test the idea that ANO1/TMEM16A contributes to luminal pH balance.

102 tivated chloride channels (CaCCs) encoded by TMEM16A control neuronal signalling, smooth muscle contr

103 physical interactions between calmodulin and TMEM16A could not be detected in copurification experime

104 2+) stores, did not alter swelling-activated TMEM16A currents.

105 high degree of similarity with heterologous TMEM16A currents.

106 TMEM16A deletion in IAS-SMCs abolishes the effects of mo

107 hat all mice homozygous for a null allele of Tmem16a died within one month of birth and exhibited sev

108 r proton concentrations ([H(+) ]o ) on mouse TMEM16A expressed in HEK-293 cells using whole-cell and

109 We demonstrate that the lower levels of TMEM16A expression in HPV-positive tumors can be attribu

110 which is attributable to an upregulation of TMEM16A expression in PASMCs.

111 We also show that TMEM16A expression is decreased in HPV-positive HNSCC at

112 The TMEM16A expression pattern established in this study thu

113 Members of the Anoctamin (Ano)/TMEM16A family have recently been identified as essentia

114 cript in addition to other paralogues of the Tmem16a family.

115 PV-negative cell lines are more dependent on TMEM16A for survival than HPV-positive cell lines.

116 TMEM16A forms a dimer with two pores.

117 TMEM16A forms calcium-activated chloride channels (CaCCs

118 be useful as pharmacological tools to study TMEM16A function and as potential drug development candi

119 rders, and for pharmacological dissection of TMEM16A function.

120 Downregulation of TMEM16A gene expression in primary cultures of rat PASMC

121 report that tissue specific knockout of the TMEM16A gene in mouse intestine and airways not only eli

122 mily members linked to tracheomalacia (mouse TMEM16A), gnathodiaphyseal dysplasia (human TMEM16E), ab

123 Mice lacking vascular TMEM16A had lower systemic blood pressure and a decrease

124 w gating, but, mutating 448 AVK451 to RSQ in TMEM16A has little effect.

125 d Cl(-) channel (CaCC), Anoctamin 1 (Ano1 or TMEM16A), has been implicated in vital physiological fun

126 TMEM16A, however, appeared to contribute little to unsti

127 For example, TMEM16A ICl activates slowly with a non-mono-exponential

128 Protons regulate TMEM16A in a voltage-independent manner, regardless of c

129 data demonstrate fundamentally new roles of TMEM16A in differentiated epithelial cells: TMEM16A prov

130 calcium-activated chloride channels such as TMEM16a in GPCR-activation of itch nerve terminals.

131 fore, we suspect that the down-regulation of TMEM16A in HPV-positive HNSCC makes TMEM16A a poor thera

132 ctures of the transmembrane domains of mouse TMEM16A in nanodiscs and in lauryl maltose neopentyl gly

133 Here, we show a functional role for TMEM16A in tumor growth.

134 ishes the Ca(2+) dependence of reconstituted TMEM16A, in a manner similar to what was reported for th

135 e constructs) abrogated the growth effect of TMEM16A, indicating a role for mitogen-activated protein

136 Mechanistically, TMEM16A-induced cancer cell proliferation and tumor grow

137 ent than the most potent previously reported TMEM16A inhibitor 4 (Ani9), and >10-fold improved metabo

138 ES, and that this is blocked by the specific TMEM16A inhibitor T16inh-A01.

139 rast to the airway and intestinal cells, all TMEM16A inhibitors fully blocked CaCC current in salivar

140 TMEM16A inhibitors have been proposed for treatment of d

141 our novel chemical classes of small molecule TMEM16A inhibitors that fully blocked TMEM16A chloride c

142 osed functions of CaCC, we hypothesized that TMEM16A inhibitors would negatively regulate both epithe

143 iate TMEM16A activity, suggesting that CLCA1-TMEM16A interactions are Mg(2+)- and metal ion-dependent

144 The calcium-activated chloride channel TMEM16A is a member of a conserved protein family that c

145 We conclude that ANO1/TMEM16A is a significant pathway in pancreatic acinar ce

146 In the gastrointestinal (GI) tract, TMEM16A is absent from smooth muscle cells, but present

147 physiological pH, E623 is un-protonated and TMEM16A is activated when intracellular calcium increase

148 gative HNSCC and that this overexpression of TMEM16A is associated with decreased patient survival.

149 In addition, TMEM16A is expressed in airway smooth muscle cells and t

150 ed calcium-activated chloride channel (CaCC) TMEM16A is expressed in the adult airway surface epithel

151 CaCCs, we further show that TMEM16B but not TMEM16A is important for hippocampal CaCC, laying the gr

152 Here, we show that TMEM16A is located in the apical membranes of epithelial

153 ports suggest that the Ca(2+) sensitivity of TMEM16A is mediated by its association with calmodulin,

154 To test whether TMEM16A is necessary and sufficient to form functional C

155 Our results demonstrate that purified TMEM16A is necessary and sufficient to recapitulate the

156 TMEM16A is opened by voltage-dependent calcium binding a

157 The calcium-activated chloride channel TMEM16A is overexpressed in a variety of cancers, includ

158 Here, we demonstrate that TMEM16A is preferentially overexpressed in HPV-negative

159 ciliogenesis and chloride transport by ANO1/TMEM16A is required for the genesis or maintenance of pr

160 TMEM16A is shown to be essential for proper activation a

161 Based on these results, we propose that TMEM16A is the major constituent of the vascular calcium

162 n, mouse, and rat BECs provide evidence that TMEM16A is the operative channel and contributes to Ca(2

163 The physiological importance of TMEM16A is underscored by the diminished rhythmic contra

164 smembrane proteins with unknown function 16 (TMEM16A) is a calcium-activated chloride channel (CaCC)

165 Anoctamin-1 (ANO1, TMEM16A) is a principal CaCC subunit in many cell types,

166 ecome possible with the discovery that Ano1 (TMEM16a) is an essential subunit of CaCCs.

167 activated Cl ((-)) channel anoctamin-1 (ANO1/TMEM16A) is located in the primary cilium and that block

168 TMEM16A knockdown reduced intravascular pressure-induced

169 denced by the absence of immunoreactivity in TMEM16A knockout mice.

170 ic contraction of gastric smooth muscle from TMEM16A knockout mice.

171 A1 constructs containing the VWA domain, and TMEM16A-like currents were activated.

172 Genetic deletion of TMEM16A markedly reduces the spontaneous activity of IHC

173 ator, Cl(-) channel in BECs and suggest that TMEM16A may be a potential target to modulate bile forma

174 lopment of small-molecule inhibitors against TMEM16A may be clinically relevant for treatment of huma

175 We demonstrate that TMEM16A-mediated control of cytoplasmic Cl(-) regulates

176 ctivated protein kinase (MAPK) activation in TMEM16A-mediated proliferation.

177 Importantly, the residual CaCC activity in Tmem16a(-/-) mice appeared inadequate for normal airway

178 trast, submandibular gland acinar cells from Tmem16A(-/-) mice lacked a Ca(2+)-activated Cl(-) curren

179 Best2(-/-) and Tmem16A(-/-) mice were used to further characterize the

180 VWA-dependent TMEM16A modulation was not modified by the S357N mutatio

181 of CaCCs, but was associated with increased TMEM16A mRNA and protein expression.

182 TMEM16A mRNA was identified in rat and human pulmonary a

183 Furthermore, TMEM16A mutant channels containing double cysteine subst

184 e that the cartilage ring defect observed in Tmem16a mutants is secondary to an expansion of the embr

185 dorsal aspect of the trachea was abnormal in Tmem16a mutants.

186 or of the calcium-activated chloride channel TMEM16A, N-((4-methoxy)-2-naphthyl)-5-nitroanthranilic a

187 Having identified TMEM16A of the transmembrane proteins with unknown funct

188 Direct pharmacological activation of TMEM16A offers a potential therapeutic strategy to reduc

189 ther, our data suggest that CLCA1 stabilizes TMEM16A on the cell surface, thus increasing surface exp

190 r binding experiments indicate CLCA1 engages TMEM16A on the surface of these cells.

191 93 cells transiently transfected with either TMEM16A or TMEM16B as well as from mouse parotid acinar

192 Heterologous expression of either TMEM16A or TMEM16B resulted in whole-cell anion currents

193 SMCs abolishes the effects of modulators for TMEM16A or VDCCs on a RyR-mediated rise in global [Ca(2+

194 Here we present evidence that TMEM16A (or anoctamin 1), a member of the transmembrane

195 s of TRPP1, TRPC1, TRPC3, TRPC6, TRPM4, ANO1/TMEM16A, or voltage-dependent Ca(2+) (CaV1.2) channels,

196 We found TMEM16A overexpression in 80% of head and neck squamous

197 TMEM16A overexpression significantly promoted anchorage-

198 Out data suggest that TMEM16A plays a general role in arteriolar and capillary

199 Cl(-) channel (CaCC), recently identified as TMEM16A, plays important roles in pulmonary vascular fun

200 polarized BEC monolayers with IL-4 increased TMEM16A protein expression, membrane localization, and t

201 TMEM16A protein levels on the cell surface were increase

202 e pore-loop of TMEM16F transplanted into the TMEM16A protein scaffold did not conduct anions or catio

203 TMEM16A in differentiated epithelial cells: TMEM16A provides a mechanism for enhanced ER Ca(2+) stor

204 INTS: The calcium-activated chloride channel TMEM16A provides a pathway for chloride ion movements th

205 Deleting 448 EAVK451 residues in TMEM16A reduced slow gating.

206 age-independent growth in vitro, and loss of TMEM16A resulted in inhibition of tumor growth both in v

207 TMEM16A RNAi knockdown also inhibited mainly the transie

208 CACNA1C, rs6702619 near PALMD, rs7127129 in TMEM16A, rs11207426 near FGGY, rs17608766 in GOSR2, and

209 External glutamic acid 623 (E623) is key for TMEM16A's ability to respond to external protons.

210 currents were significantly knocked down by TMEM16A siRNA.

211 Further analyses revealed that several TMEM16A splice variants were detected in rat PASMCs and

212 ogical functions, knowledge of the mammalian TMEM16A structure and identification of its pore-lining

213 ium-activated chloride channels that contain TMEM16A subunits.

214 a developmental small-molecule inhibitor of TMEM16A, T16A-inh01 (A01), abrogated tumor cell prolifer

215 activated calcium-dependent chloride channel TMEM16A (TAOS2 or ANO1) has been reported in several mal

216 Small-molecule, TMEM16A-targeted activators may be useful for drug thera

217 levels of the Ca(2+)-activated Cl(-) channel TMEM16A, the major apical Cl(-) efflux pathway in saliva

218 By contrast, expression of TMEM16A, the water channel aquaporin 5 (AQP5), and other

219 In TMEM16A this region has also been suggested to contain r

220 ice homozygous for a null allele of Tmem16a (Tmem16a(tm1Bdh)(/tm1Bdh)) and did not develop subsequent

221 eonatal mice homozygous for a null allele of Tmem16a (Tmem16a(-/-)) to investigate the role of this c

222 irth in mice homozygous for a null allele of Tmem16a (Tmem16a(tm1Bdh)(/tm1Bdh)) and did not develop s

223 ly includes Ca(2+)-activated anion channels (TMEM16A, TMEM16B), a cation channel (TMEM16F) and protei

224 cal tools to investigate the contribution of TMEM16A to CaCC conductance in human airway and intestin

225 ice homozygous for a null allele of Tmem16a (Tmem16a(-/-)) to investigate the role of this channel in

226 When compared with wild-type tracheas, the Tmem16a(-/-) tracheas exhibited a >60% reduction in puri

227 adequate for normal airway hydration because Tmem16a(-/-) tracheas exhibited significant, neonatal, l

228 GI muscles express splice variants of the Tmem16a transcript in addition to other paralogues of th

229 used by a global rise in [Ca(2+)]i via a RyR-TMEM16A-VDCC signalling module sets the basal tone.

230 TMEM16A was found recently to be a calcium-activated Cl(

231 alcium-dependent chloride channel DOG1 (ANO1/TMEM16A), which is strongly and specifically expressed i

232 ic analysis of the tumorigenic properties of TMEM16A, which represents a potentially novel therapeuti

233 lass of CaCC, anoctamin 1 (ANO1), encoded by Tmem16a, which was discovered to be highly expressed in

234 Our results also show that association of TMEM16A with other proteins, such as calmodulin, is not

235 Replacement of the putative pore-loop of TMEM16A with that of TMEM16B (TMEM16A-B channels) reduce

236 ns identified tannic acid as an inhibitor of TMEM16A, with IC(50) approximately 6 muM and approximate

237 conditions determine the open probability of TMEM16A without changing its calcium sensitivity.