Involvements of the ABC protein ABCF2 and α-actinin-4 in regulation of cell volume and anion channels in human epithelial cells
Involvements of the ABC protein ABCF2 and α-actinin-4 in regulation of cell volume and anion channels in human epithelial cells
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DOI：
10.1002/jcp.24050
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发表时间：
2012-10-01
影响因子：
5.6
通讯作者：
Okada, Yasunobu
中科院分区：
文献类型：
--
作者：
Ando-Akatsuka, Yuhko;Shimizu, Takahiro;Okada, Yasunobu
在渗透膨胀后，细胞体积被称为调节性体积减少(RVD)的过程来调节。尽管肌动蛋白细胞骨架已知在RVD中起调节作用，但肌动蛋白结合蛋白如何参与RVD过程尚不清楚。在本研究中，肌动蛋白结合蛋白α-肌动蛋白-4(ACTN4)参与了人上皮HEK293T细胞的RVD。ACTN4过表达显著促进RVD，而siRNA介导的内源性ACTN4下调抑制RVD。当细胞受到低渗应激时，100,000×g颗粒中ACTN4的含量增加，这对细胞松弛素D的预处理敏感。蛋白质重叠分析表明，ABCF2是ABC转运蛋白超家族的胞质成员，是ACTN4的结合伙伴。ACTN4-ABCF2的相互作用在低张刺激下明显增强，需要ABCF2的NH2末端区域。ABCF2过表达抑制RVD，而ABCF2下调促进RVD。然后，我们测试了ABCF2是否对容量敏感的外向整流阴离子通道(VSOR)的活性有抑制作用，因为另一种ABC转运体成员CFTR被证明抑制了VSOR的活性，VSOR是参与RVD的氯离子外流的中介。ABCF2过表达可显著降低全细胞VSOR电流，而siRNA介导的ABCF2缺失可显著增强全细胞VSOR电流。因此，本研究表明ACTN4是RVD的增强子，而ABCF2则是VSOR和RVD的抑制者，并提示ACTN4和ABCF2之间的肿胀诱导的相互作用阻止了ABCF2抑制人上皮细胞中VSOR的活性。J.细胞。物理。2012年，227：34983510。(C)2012年威利期刊公司。
After osmotic swelling, cell volume is regulated by a process called regulatory volume decrease (RVD). Although actin cytoskeletons are known to play a regulatory role in RVD, it is not clear how actin-binding proteins are involved in the RVD process. In the present study, an involvement of an actin-binding protein, a-actinin-4 (ACTN4), in RVD was examined in human epithelial HEK293T cells. Overexpression of ACTN4 significantly facilitated RVD, whereas siRNA-mediated downregulation of endogenous ACTN4 suppressed RVD. When the cells were subjected to hypotonic stress, the content of ACTN4 increased in a 100,000 x g pellet, which was sensitive to cytochalasin D pretreatment. Protein overlay assays revealed that ABCF2, a cytosolic member of the ABC transporter superfamily, is a binding partner of ACTN4. The ACTN4-ABCF2 interaction was markedly enhanced by hypotonic stimulation and required the NH2-terminal region of ABCF2. Overexpression of ABCF2 suppressed RVD, whereas downregulation of ABCF2 facilitated RVD. We then tested whether ABCF2 has a suppressive effect on the activity of volume-sensitive outwardly rectifying anion channel (VSOR), which is known to mediate Cl- efflux involved in RVD, because another ABC transporter member, CFTR, was shown to suppress VSOR activity. Whole-cell VSOR currents were largely reduced by overexpression of ABCF2 and markedly enhanced by siRNA-mediated depletion of ABCF2. Thus, the present study indicates that ACTN4 acts as an enhancer of RVD, whereas ABCF2 acts as a suppressor of VSOR and RVD, and suggests that a swelling-induced interaction between ACTN4 and ABCF2 prevents ABCF2 from suppressing VSOR activity in the human epithelial cells. J. Cell. Physiol. 227: 34983510, 2012. (C) 2012 Wiley Periodicals, Inc.