Gap junction turnover, intracellular trafficking, and phosphorylation of connexin43 in brefeldin A-treated rat mammary tumor cells.
Gap junction turnover, intracellular trafficking, and phosphorylation of connexin43 in brefeldin A-treated rat mammary tumor cells.
复制标题
DOI：
10.1083/jcb.131.5.1193
复制
发表时间：
1995-12
期刊：
影响因子：
--
通讯作者：
Kasprzak L
中科院分区：
文献类型：
--
作者：
Laird DW;Castillo M;Kasprzak L
当一个细胞的连接蛋白寡聚体(连接蛋白)注册并与相邻细胞的连接蛋白配对形成紧密堆积的阵列(斑块)时，细胞间缝隙连接通道被认为是形成的。在目前的研究中，我们使用大鼠乳腺BICR-M1Rk肿瘤细胞系来检测连接蛋白43(Cx43)的运输、成熟和动力学。结果表明，Cx43除了存在于高尔基体细胞外，还存在于这些细胞和正常大鼠肾细胞中。Brefeldin A(BFA)阻止Cx43转运到乳腺细胞表面，并阻止Cx43的42-kD形式磷酸化为44-kD和46-kD物种。然而，Cx43的磷酸化在BFA存在的情况下发生，而它仍然是ER或高尔基体的驻留，产生43kD形式的Cx43。此外，在去除BFA后，捕获在内质网/高尔基体内的42-和43-kD形式的Cx43可用于缝隙连接组装。用BFA处理6h的乳腺细胞失去了先前存在的缝隙连接“斑块”，以及44-和46-kD形式的Cx43和功能偶联。这些事件在去除BFA后1h是可逆的，不依赖于蛋白质的合成。综上所述，我们提供了强有力的证据表明，在BICR-M1Rk肿瘤细胞中：(A)Cx43在ER/高尔基体中瞬时磷酸化，(B)被困在ER/高尔基体中的Cx43不会快速降解，并可在BFA被移除后用于组装新的缝隙连接通道，(C)缝隙连接斑块的快速周转与Cx43 44-和46-kD形式的丢失有关。
Intercellular gap junction channels are thought to form when oligomers of connexins from one cell (connexons) register and pair with connexons from a neighboring cell en route to forming tightly packed arrays (plaques). In the current study we used the rat mammary BICR-M1Rk tumor cell line to examine the trafficking, maturation, and kinetics of connexin43 (Cx43). Cx43 was conclusively shown to reside in the Golgi apparatus in addition to sites of cell-cell apposition in these cells and in normal rat kidney cells. Brefeldin A (BFA) blocked Cx43 trafficking to the surface of the mammary cells and also prevented phosphorylation of the 42-kD form of Cx43 to 44- and 46-kD species. However, phosphorylation of Cx43 occurred in the presence of BFA while it was still a resident of the ER or Golgi apparatus yielding a 43-kD form of Cx43. Moreover, the 42- and 43-kD forms of Cx43 trapped in the ER/Golgi compartment were available for gap junction assembly upon the removal of BFA. Mammary cells treated with BFA for 6 h lost preexisting gap junction "plaques," as well as the 44- and 46-kD forms of Cx43 and functional coupling. These events were reversible 1 h after the removal of BFA and not dependent on protein synthesis. In summary, we provide strong evidence that in BICR-M1Rk tumor cells: (a) Cx43 is transiently phosphorylated in the ER/Golgi apparatus, (b) Cx43 trapped in the ER/Golgi compartment is not subject to rapid degradation and is available for the assembly of new gap junction channels upon the removal of BFA, (c) the rapid turnover of gap junction plaques is correlated with the loss of the 44- and 46-kD forms of Cx43.