The role of microtubule plus-end capture at cortical sites in the assembly of apico-basal microtubules in polarised epithelial cells.

微管是管状结构，它对许多细胞功能都很重要，包括细胞内囊泡和分子的运输。因此，在细胞内正确组装和定位微管是非常重要的。微管在细胞内形成的特定模式因细胞类型和功能而异。大多数动物细胞都有一个微管的放射状排列，这些微管固定在一个叫做中心体的中心结构上。在这些细胞中，中心体的作用是开始微管的组装，并使微管紧紧地附着在中心体上。然而，许多特殊的细胞，如在肠道和肾脏中发现的极化上皮细胞，以不同的方式排列它们的微管。在这里，大多数微管被锚定在远离中心体的细胞顶端，并向基部运行，形成所谓的顶基阵列。我们最近的研究结果表明，顶基微管在中心体组装，释放并移动到顶端位置，在那里它们被捕获和锚定。这个项目的目的是发现微管是否首先从中心体中生长出来，与细胞皮层接触，从中心体中释放出来，然后在位于皮层的动力蛋白的拉动作用下向下移动。最终的结果将是微管的一端（正端）被拉到细胞基部，而另一端（负端）被锚定在细胞顶端。我们想知道粘附在微管生长末端的EB1和CLIP-170之类的蛋白质是否对动力蛋白或皮层的其他蛋白质的捕获很重要。例如，了解CLIP-170在现实生活中是否对微管的正常组装至关重要是非常重要的。因此，我们将分析从小鼠分离的内耳（耳蜗）中提取的不产生CLIP-170的组织。最后，我们还想知道被称为肌动蛋白的细丝是否有助于微管到达目的地。我们将使用一种叫做gfp的特殊荧光（发光）分子与蛋白质相连，这样微管或它们的正端就会发光，我们将拍摄它们随时间移动的视频，并使用荧光染料和专用显微镜观察这些重要蛋白质的定位。显然，微管对细胞的正常功能至关重要，我们需要确定它们是如何组织和控制的，然后才能揭开许多疾病的原因和后果。

Microtubules are tubular structures, which are important for many cellular functions including the transport of vesicles and molecules within cells. It is therefore very important that the microtubules are assembled and positioned correctly within cells. The specific pattern that microtubules form within cells vary depending on cell type and function. Most animal cells have a radial array of microtubules anchored at a centrally located structure called the centrosome. In these cells the centrosome's job is to start off the assembly of the microtubules and to keep them tightly bound to it. However, many specialised cells like polarised epithelial cells found in the gut and kidneys arrange their microtubules in a different way. Here most of the microtubules are anchored at the cell apex away from the centrosome and run towards the base forming a so called apico-basal array. Our recent findings suggest that the apico-basal microtubules are assembled at the centrosome, released and moved to apical sites where they are captured and anchored. The aim of this project is to find out whether the microtubules first grow out from the centrosome, make contact with the cell cortex, are released from the centrosome and then move downward by the pulling action of dynein motor proteins located at the cortex. The end result would be that one end (plus-end) of the microtubules is pulled down to the cell base while the other end (minus-end) becomes anchored at the cell apex. We want to find out whether proteins such as EB1 and CLIP-170, which stick to the growing end of microtubules, are important for the capture by dynein or other proteins at the cortex. It is very important to know if for example CLIP-170 is vital for the normal assembly of the microtubule in real life. We will therefore analyse tissue from the inner ear (cochlea) isolated from mice, which do not produce CLIP-170. Finally, we would also like to know whether thin filaments known as actin help the microtubules to get to their destination. We will use special fluorescent (glowing) molecules called GFPs linked to proteins so that the microtubules or their plus-ends glow and we will make movies of their movements over time and look at the localisation of these important proteins using fluorescent dyes and specialised microscopes. Microtubules are clearly vital for the normal function of a cell and we need to establish how they are organised and controlled before we can unravel the causes and consequences of many diseases.