Limiting transport steps and novel interactions of Connexin-43 along the secretory pathway
Limiting transport steps and novel interactions of Connexin-43 along the secretory pathway
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DOI：
10.1007/s00418-009-0617-x
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发表时间：
2009-09-01
影响因子：
2.3
通讯作者：
Duden, Rainer
中科院分区：
文献类型：
--
作者：
Majoul, Irina V.;Onichtchouk, Daria;Duden, Rainer
连接蛋白是在所有脊椎动物中表达的四跨膜结构域蛋白，形成连接细胞的可渗透间隙连接通道。在这里，我们分析了 Connexin-43 (Cx43) 向质膜的转运，并研究了小 GTP 酶沿分泌途径的作用。我们发现，GTP 和 GDP 限制性 Sar1 都能阻止 Cx43 从内质网 (ER) 退出，但只有 GTP 限制性 Sar1 将 Cx43 阻滞在 COP II 包被的 ER 退出位点中，并在 ER 组分中积累 14-3-3 蛋白。 FRET-FLIM 数据证实，Cx43 已经以寡聚形式存在于 ER 出口位点，表明 14-3-3 在 Cx43 寡聚化中发挥体内作用。 Cx43 从 ER 的退出可以被其他因素阻断，例如 COP I 外壳的 β 亚基的表达或作用于基于微管的动力蛋白运动复合体的 p50/动力蛋白。 GTP 限制性 Arf1 阻断高尔基体中的 Cx43。最后，我们发现 GTP 限制性 Arf6 可从细胞-细胞界面上去除 Cx43 间隙连接斑块，并将其靶向降解。这些数据提供了小 GTP 酶如何通过分泌途径调节 Cx43 运输、促进或消除通过间隙连接的细胞间通讯的分子解释。
Connexins are four-transmembrane-domain proteins expressed in all vertebrates which form permeable gap junction channels that connect cells. Here, we analysed Connexin-43 (Cx43) transport to the plasma membrane and studied the effects of small GTPases acting along the secretory pathway. We show that both GTP- and GDP-restricted Sar1 prevents exit of Cx43 from the endoplasmic reticulum (ER), but only GTP-restricted Sar1 arrests Cx43 in COP II-coated ER exit sites and accumulates 14-3-3 proteins in the ER fraction. FRET-FLIM data confirm that already in ER exit sites Cx43 exists in oligomeric form, suggesting an in vivo role for 14-3-3 in Cx43 oligomerization. Exit of Cx43 from the ER can be blocked by other factors-such as expression of the beta subunit of the COP I coat or p50/dynamitin that acts on the microtubule-based dynein motor complex. GTP-restricted Arf1 blocks Cx43 in the Golgi. Lastly, we show that GTP-restricted Arf6 removes Cx43 gap junction plaques from the cell-cell interface and targets them to degradation. These data provide a molecular explanation of how small GTPases act to regulate Cx43 transport through the secretory pathway, facilitating or abolishing cell-cell communication through gap junctions.