Ca2+/Calmodulin-Dependent Protein Kinase II γ-Dependent Serine727 Phosphorylation Is Required for TMEM16A Ca2+-Activated Cl- Channel Regulation in Cerebrovascular Cells
Ca2+/Calmodulin-Dependent Protein Kinase II γ-Dependent Serine727 Phosphorylation Is Required for TMEM16A Ca2+-Activated Cl- Channel Regulation in Cerebrovascular Cells
复制标题
CaMKIIγ 依赖性 Serine727 磷酸化是脑血管细胞中 TMEM 16A Ca2 激活 Cl 通道调节所必需的
DOI：
10.1253/circj.cj-17-0585
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发表时间：
2018-03-01
影响因子：
3.3
通讯作者：
Guan, Yong-Yuan
中科院分区：
文献类型：
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作者：
Lin, Cai-Xia;Lv, Xiao-Fei;Guan, Yong-Yuan
背景：TMEM16A是Ca2+激活的氯离子通道（CaCCs）的关键成分，并介导高血压脑血管重构中基底动脉平滑肌细胞（BASMC）的增殖。CaMKII是CaCC的负调节因子，四种CaMKII亚型（α、β、γ和δ）在脉管系统中表达；然而，CaMKII亚型影响tmem16a相关的CaCC和BASMC增殖的方式和方式尚不清楚。方法和结果：膜片钳和小干扰RNA （siRNA）敲除不同CaMKII亚型表明，只有CaMKII γ抑制BASMCs中原生Ca2+激活的氯离子电流（I-Cl.(Ca)）。TMEM16A过表达可激活TMEM16A Cl-电流，抑制血管紧张素II （Ang II）诱导的basmc细胞增殖。共免疫沉淀和拉下实验表明CaMKII γ和TMEM16A蛋白之间存在相互作用。通过CaMKII γ在TMEM16A丝氨酸残基上的磷酸化调节TMEM16A的cl电流。TMEM16A中的Serine525和Serine727突变为丙氨酸，只有Ser727 （S727A）突变逆转了TMEM16A Cl-电流对CaMKII γ的抑制。拟磷突变S727D显著降低TMEM16A Cl（-）电流，逆转TMEM16A介导的BASMC增殖抑制，模拟CaMKII γ对TMEM16A的抑制作用。CaMKII γ异构体含量显著增加，同时TMEM16A和I-Cl含量显著降低。钙对注入angii小鼠基底动脉增生性重塑的影响。结论：CaMKII γ对TMEM16A中丝氨酸727的磷酸化提供了调节TMEM16A CaCC活性和Ang ii诱导的BASMC增殖的新机制。
Background: TMEM16A is a critical component of Ca2+-activated chloride channels (CaCCs) and mediates basilar arterial smooth muscle cell (BASMC) proliferation in hypertensive cerebrovascular remodeling. CaMKII is a negative regulator of CaCC, and four CaMKII isoforms (alpha, beta, gamma and delta) are expressed in vasculature; however, it is unknown which and how CaMKII isoforms affect TMEM16A-associated CaCC and BASMC proliferation.Methods and Results: Patch clamp and small interfering RNA (siRNA) knockdown of different CaMKII isoforms revealed that only CaMKII gamma inhibited native Ca2+-activated chloride currents (I-Cl.(Ca)) in BASMCs. The TMEM16A overexpression evoked TMEM16A Cl- current and inhibited angiotensin II (Ang II)-induced proliferation in BASMCs. The co-immunoprecipitation and pull-down assay indicated an interaction between CaMKII gamma and TMEM16A protein. TMEM16A Cl-current was modulated by CaMKII gamma phosphorylation at serine residues in TMEM16A. Serine525 and Serine727 in TMEM16A were mutated to alanine, and only mutation at Ser727 (S727A) reversed the CaMKII gamma inhibition of the TMEM16A Cl- current. Phosphomimetic mutation S727D markedly decreased TMEM16A Cl(-)current and reversed TMEM16A-mediated suppression of BASMC proliferation, mimicking the inhibitory effects of CaMKII gamma on TMEM16A. A significant increase in CaMKII gamma isoform content was observed in parallel to the decrease of TMEM16A and I-Cl.Ca in basilar artery proliferative remodeling in Ang II-infused mice.Conclusions: Serine 727 phosphorylation in TMEM16A by CaMKII gamma provides a new mechanism for regulating TMEM16A CaCC activity and Ang II-induced BASMC proliferation.