P2X4 Forms Functional ATP-activated Cation Channels on Lysosomal Membranes Regulated by Luminal pH
P2X4 Forms Functional ATP-activated Cation Channels on Lysosomal Membranes Regulated by Luminal pH
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DOI：
10.1074/jbc.m114.552158
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发表时间：
2014-06-20
影响因子：
4.8
通讯作者：
Dong, Xian-Ping
中科院分区：
文献类型：
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作者：
Huang, Peng;Zou, Yuanjie;Dong, Xian-Ping
P2X 受体通常被称为质膜阳离子通道，参与多种细胞功能。这些通道的特性已在质膜上得到广泛研究。然而，对阿米巴原虫的研究表明，P2X 受体也存在于细胞内，并参与囊泡与质膜的融合。最近，研究表明，除了质膜表达之外，哺乳动物 P2X4 也定位于细胞内的溶酶体中。然而，尚不清楚溶酶体 P2X4 受体是否作为通道发挥作用以及它们如何被激活和调节。在本文中，我们证明 P2X4 及其天然配体 ATP 在 COS1 和 HEK293 细胞的溶酶体中富集。通过直接记录扩大的溶酶体液泡的膜电流，我们证明溶酶体 P2X4 形成由管腔侧 ATP 以 pH 依赖性方式激活的通道。虽然管腔侧的酸性 pH 值抑制 P2X4 活性，但在 ATP 存在的情况下增加管腔 pH 值会导致 P2X4 激活。我们进一步表明，对于质膜 P2X4，溶酶体 P2X4 被伊维菌素增强，但对苏拉明和 PPADS 不敏感，并且在激活后渗透大阳离子 N-甲基-D-葡萄糖胺。我们的数据表明，P2X4 在受管腔 pH 调节的溶酶体膜上形成功能性 ATP 激活的阳离子通道。结合已报道的低等生物体细胞内 P2X 的融合效应，我们推测溶酶体定位的 P2X4 可能在哺乳动物细胞中酸性细胞器的膜运输中发挥特定作用。
P2X receptors are commonly known as plasma membrane cation channels involved in a wide variety of cell functions. The properties of these channels have been extensively studied on the plasma membrane. However, studies in amoeba suggest that P2X receptors are also present intracellularly and involved in vesicle fusion with the plasma membrane. Recently, it was shown that in addition to plasma membrane expression, mammalian P2X4 was also localized intracellularly in lysosomes. However, it was not clear whether the lysosomal P2X4 receptors function as channels and how they are activated and regulated. In this paper, we show that both P2X4 and its natural ligand, ATP, are enriched in lysosomes of COS1 and HEK293 cells. By directly recording membrane currents from enlarged lysosomal vacuoles, we demonstrated that lysosomal P2X4 formed channels activated by ATP from the luminal side in a pH-dependent manner. While the acidic pH at the luminal side inhibited P2X4 activity, increasing the luminal pH in the presence of ATP caused P2X4 activation. We further showed that, as for the plasma membrane P2X4, the lysosomal P2X4 was potentiated by ivermectin but insensitive to suramin and PPADS, and it permeated the large cation N-methyl-D-glucamine upon activation. Our data suggest that P2X4 forms functional ATP-activated cation channels on lysosomal membranes regulated by luminal pH. Together with the reported fusion effect of intracellular P2X in lower organisms, we speculate that the lysosome-localized P2X4 may play specific roles in membrane trafficking of acidic organelles in mammalian cells.