Investigating the roles of Arabidopsis STIC1 and STIC2 in chloroplast protein transport

叶绿体和线粒体是许多细胞的正常组成部分--它们是亚细胞结构，称为细胞器。有趣的是，这两个细胞器是从10亿多年前被其他细胞吞噬的细菌进化而来的，在许多方面它们仍然类似于自由生活的细菌。植物细胞中的叶绿体含有绿色色素叶绿素，并负责光合作用(捕捉阳光能量并利用其制造糖的过程)的反应。由于光合作用是向生物世界输入能量的唯一重要机制，叶绿体具有不可估量的重要性，不仅对植物，而且对地球上的所有生命都是如此。叶绿体在许多其他方面也很重要，因为它们在油脂、蛋白质和淀粉的合成中起着至关重要的作用。尽管叶绿体确实含有DNA(这是它们作为自由生活的光合细菌进化而来的遗迹)，因此可以制造自己的一些蛋白质，但构建一个功能齐全的叶绿体所需的3000种蛋白质中，90%是在细胞核的DNA上编码的。因此，大多数叶绿体蛋白是在细胞基质中的细胞器外形成的，称为细胞质。由于每个叶绿体都被双层膜或被膜包围，不受蛋白质被动运动的影响，这是一个重要的问题。为了克服这个问题，叶绿体进化出了一种复杂的蛋白质输入装置，它使用能量(以三磷酸腺苷的形式)驱动蛋白质从细胞质输入。这种输入装置由两个分子机器组成：一个在外被膜上称为TOC(叶绿体外被膜上转运子的缩写)，另一个在内被膜上被称为TIC。每台机器由几个蛋白质组成，这些蛋白质相互协作，确保进口的效率。TIC机器的一个特点是，它从叶绿体内部或基质中招募一类特殊的蛋白质，称为“伴侣”。这些间质伴侣蛋白的作用就像马达一样，因为它们使用来自三磷酸腺苷的能量来驱动蛋白质输入。我们正在研究一种名为拟南芥的模式植物，它在研究中具有许多优势，例如可以获得大量的突变体(每个突变体都有一个特定基因的突变)。一种这样的突变植物tic40在TIC基因中存在缺陷，以至于叶绿体蛋白输入不能有效地发挥作用。几年前，我们发现了其他称为tic1和tic2的突变，它们显著提高了tic40中的蛋白质进口效率。最近，我们发现了STIC突变会影响哪些基因(从而影响哪些蛋白质)：STIC1属于一个已知的蛋白质运输因子家族，以前没有人认为这些因子作用于叶绿体被膜，而STIC2与一组功能未知的细菌蛋白质有关。有趣的是，我们已经证明STIC2可以与STIC1结合，也可以与基质伴侣和TIC机器结合。因此，我们认为STIC2可能是上述进口马达的一个新元件。它还可能有助于将新进口的蛋白质从TIC装置引导到它们的最终目的地，这可能是STIC1发挥作用的地方。我们将做实验来检验这些理论。由于叶绿体执行基本功能，而且蛋白质输入对叶绿体发育至关重要，因此，没有叶绿体蛋白质输入机制的植物无法存活也就不足为奇了(事实上，它们以胚胎的形式死亡)。同样，由于我们最终都依赖植物产品生存，因此在全球范围内，叶绿体蛋白的进口是必不可少的。由于叶绿体在许多重要的经济产物(如油、淀粉)的合成中起着重要作用，更好地了解这些细胞器是如何发展的将使我们能够提高作物的生产力或以其他方式操纵它们的产物。

Chloroplasts and mitochondria are normal components of many cells - they are sub-cellular structures called organelles. Interestingly, these two organelles evolved from bacteria that were engulfed by other cells over a billion years ago, and in many ways they still resemble free-living bacteria. Chloroplasts are found in plant cells, contain the green pigment chlorophyll, and are responsible for the reactions of photosynthesis (the process that captures sunlight energy and uses it to make sugars). As photosynthesis is the only significant mechanism of energy-input into the living world, chloroplasts are of inestimable importance, not just to plants but to all life on Earth. Chloroplasts are also important in many other ways, as they play essential roles in the synthesis of oils, proteins and starch. Although chloroplasts do contain DNA (a relic from their evolutionary past as free-living photosynthetic bacteria), and so can make some of their own proteins, >90% of the 3000 proteins needed to build a fully-functional chloroplast are encoded on DNA in the cell nucleus. Thus, most chloroplast proteins are made outside of the organelle in the cellular matrix known as the cytosol. As chloroplasts are each surrounded by a double membrane, or envelope, that is impervious to the passive movement of proteins, this presents a significant problem. To overcome the problem, chloroplasts evolved a sophisticated protein import apparatus, which uses energy (in the form of ATP) to drive the import of proteins from the cytosol. This import apparatus comprises two molecular machines: one in the outer envelope membrane called TOC (an abbreviation of "Translocon at the outer envelope of chloroplasts"), and another in the inner envelope membrane called TIC. Each machine is made up of several proteins which cooperate to ensure the efficiency of import. One of the features of the TIC machine is that it recruits a special class of proteins from the chloroplast interior, or stroma, called "chaperones". These stromal chaperones act like a motor as they use the energy from ATP to drive protein import. We work on a model plant called Arabidopsis that has many advantages for research, such as an availability of numerous mutants (each one with a mutation in a specific gene). One such mutant plant, tic40, has a defect in a TIC gene such that chloroplast protein import does not work efficiently. Several years ago we identified other mutations called stic1 and stic2 (stic stands for "suppressor of tic40") which significantly improve protein import efficiency in tic40. Recently, we discovered which genes (and therefore which proteins) the stic mutations affect: STIC1 belongs to a family of well-known protein transport factors that were not previously thought to act in the chloroplast envelope, while STIC2 is related to a group of bacterial proteins of unknown function. Interestingly, we have shown that STIC2 can bind to STIC1, as well to stromal chaperones and the TIC machine. Thus, we believe that STIC2 may be a new component of the aforementioned import motor. It may also help to guide newly-imported proteins from the TIC apparatus to their final destination, which is perhaps where STIC1 plays its role. We will do experiments to test these theories. As chloroplasts carry out essential functions, and because protein import is essential for chloroplast development, it is not surprising that plants without a functional chloroplast protein import machinery are unable to survive (in fact, they die as embryos). Similarly, as we are all ultimately dependent upon plant products for survival, it follows that chloroplast protein import is essential on a global scale. As chloroplasts play major roles in the synthesis of many economically important products (e.g., oils, starch), a better understanding of how these organelles develop will enable us to enhance the productivity of crop plants or otherwise manipulate their products.