Department of Physiology, College of Medicine, Seoul National University, 103 Daehak-ro, Jongno-gu, Seoul, 03080, Republic of Korea.
Department of Physiology, Dongguk University College of Medicine, 123 Dongdae-ro, Gyeongju, 38066, Republic of Korea.
Biochem Biophys Res Commun. 2018 Sep 18;503(4):2348-2354. doi: 10.1016/j.bbrc.2018.06.160. Epub 2018 Jul 10.
Anoctamin 6 (ANO6/TMEM16F) is a recently identified membrane protein that has both phospholipid scramblase activity and anion channel function activated by relatively high [Ca]. In addition to the low sensitivity to Ca, the activation of ANO6 Cl conductance is very slow (>3-5 min to reach peak level at 10 μM [Ca]), with subsequent inactivation. In a whole-cell patch clamp recording of ANO6 current (I), disruption of the actin cytoskeleton with cytochalasin-D (cytoD) significantly accelerated the activation kinetics, while actin filament-stabilizing agents (phalloidin and jasplakinolide) commonly inhibited I. Inside-out patch clamp recording of ANO6 (I) showed immediate activation by raising [Ca]. We also found that intracellular ATP (3 mM MgATP in pipette solution) decelerated the activation of I, and also prevented the inactivation of I. However, the addition of cytoD still accelerated both activation and inactivation of I. We conclude that the actin cytoskeleton and intracellular ATP play major roles in the Ca-dependent activation and inactivation of I, respectively.
Anoctamin 6（ANO6/TMEM16F）是一种最近被发现的膜蛋白，它具有磷脂翻转酶活性和阴离子通道功能，由相对较高的[Ca]激活。除了对 Ca 的低敏感性外，ANO6 Cl 电导的激活非常缓慢（在 10μM [Ca]下达到峰值水平需要>3-5 分钟），随后失活。在 ANO6 电流（I）的全细胞膜片钳记录中，用细胞松弛素-D（cytoD）破坏肌动蛋白细胞骨架会显著加速激活动力学，而肌动蛋白丝稳定剂（鬼笔环肽和 jasplakinolide）通常会抑制 I。通过升高 [Ca]，可以立即对内质网片钳记录中的 ANO6（I）进行激活。我们还发现细胞内 ATP（在管内溶液中为 3mM MgATP）会减缓 I 的激活，并且还可以防止 I 的失活。然而，添加 cytoD 仍然可以加速 I 的激活和失活。我们得出结论，肌动蛋白细胞骨架和细胞内 ATP 分别在 I 的 Ca 依赖性激活和失活中起主要作用。
