Dynamic Remodeling of the Plastid Envelope Membranes - A Tool for Chloroplast Envelope in vivo Localizations.
Dynamic Remodeling of the Plastid Envelope Membranes - A Tool for Chloroplast Envelope in vivo Localizations.
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DOI：
10.3389/fpls.2012.00007
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发表时间：
2012
影响因子：
5.6
通讯作者：
Weber AP
中科院分区：
文献类型：
--
作者：
Breuers FK;Bräutigam A;Geimer S;Welzel UY;Stefano G;Renna L;Brandizzi F;Weber AP
两个包膜界定了质体，质体是植物细胞的决定性细胞器。内膜和外膜的蛋白质和脂质成分是独特的。一些研究试图建立这两种膜的蛋白质组；然而，由于它们非常接近，因此很难区分它们。在这里，我们描述了一种新方法来区分两个膜之间蛋白质的定位，使用基于活细胞成像和瞬时表达的简单方法。我们的方法基于 GFP 融合分布的分析，旨在验证外膜蛋白质组学数据。为了区分外包膜和内包膜蛋白定位，我们使用 AtTOC64-GFP 和 AtTIC40-GFP 作为各自的对照。在我们的分析过程中，我们观察到在蛋白质过度表达的条件下膜增殖和叶绿体形状丧失。增殖的形态变化与过表达蛋白质的亚细胞器分布相关。特别是，当内包膜中形成多层膜时，外包膜形成长的延伸到细胞质中。使用电子显微镜，我们发现这些延伸是质体，是质体的动态特征。由于膜的行为不同并且与蛋白质定位有关，因此我们建议基于膜形态差异分析的体内研究可用于区分蛋白质的内膜和外膜定位。为了证明这种方法的适用性，我们证明了 AtLACS9 到外包膜的定位。我们还讨论了蛋白质对膜行为的影响以及蛋白质插入膜的调节，并提供了关于基质形成的新假设。
Two envelope membranes delimit plastids, the defining organelles of plant cells. The inner and outer envelope membranes are unique in their protein and lipid composition. Several studies have attempted to establish the proteome of these two membranes; however, differentiating between them is difficult due to their close proximity. Here, we describe a novel approach to distinguish the localization of proteins between the two membranes using a straightforward approach based on live cell imaging coupled with transient expression. We base our approach on analyses of the distribution of GFP-fusions, which were aimed to verify outer envelope membrane proteomics data. To distinguish between outer envelope and inner envelope protein localization, we used AtTOC64–GFP and AtTIC40–GFP, as respective controls. During our analyses, we observed membrane proliferations and loss of chloroplast shape in conditions of protein over-expression. The morphology of the proliferations varied in correlation with the suborganellar distribution of the over-expressed proteins. In particular, while layers of membranes built up in the inner envelope membrane, the outer envelope formed long extensions into the cytosol. Using electron microscopy, we showed that these extensions were stromules, a dynamic feature of plastids. Since the behavior of the membranes is different and is related to the protein localization, we propose that in vivo studies based on the analysis of morphological differences of the membranes can be used to distinguish between inner and outer envelope localizations of proteins. To demonstrate the applicability of this approach, we demonstrated the localization of AtLACS9 to the outer envelope membrane. We also discuss protein impact on membrane behavior and regulation of protein insertion into membranes, and provide new hypotheses on the formation of stromules.
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影响因子：
4.1
作者：
Braeutigam, Andrea;Shrestha, Roshan P.;Weber, Andreas P. M.
通讯作者：
Weber, Andreas P. M.
影响因子：
7
作者：
Ferro, M;Salvi, D;Rolland, N
通讯作者：
Rolland, N
影响因子：
7.4
作者：
Bleckmann, Andrea;Weidtkamp-Peters, Stefanie;Simon, Ruediger
通讯作者：
Simon, Ruediger
影响因子：
6.9
作者：
Hanson, MR;Köhler, RH
通讯作者：
Köhler, RH
DOI：
10.1111/j.1432-1033.1991.tb15714.x
发表时间：
1991-01-30
期刊：
EUROPEAN JOURNAL OF BIOCHEMISTRY
影响因子：
--
作者：
DRESESWERRINGLOER, U;FISCHER, K;FLUGGE, UI
通讯作者：
FLUGGE, UI