Re-organisation of microtubule minus-ends during apico-basal epithelial polarisation and differentiation

细长上皮细胞（如肠和肾上皮细胞）的发育过程中，被称为微管的管状结构发生了戏剧性的重排，形成了从细胞顶端到底部的顶基束。微管的重排很可能依赖于分子分布的变化，如固定微管一端的九蛋白，并将它们组织成特定的模式，如顶基束，这可能对这些细胞的正常功能至关重要。上皮伸长和微管束形成缺陷导致功能丧失、细胞异常迁移和癌症。微管是如何在细长的上皮细胞中组织成束的还不完全清楚。我们所知道的大部分都来自于对培养的柱状扁平细胞片的研究，这些细胞片没有自然的组织结构。重要的是，我们现在已经建立了一种新的活体三维肠道类器官培养系统，该系统模拟正常肠道发育，产生具有细长上皮细胞的迷你肠道样结构。该系统为研究上皮延伸和组织发育过程中的微管重组提供了一种强大的新方法，使用活体显微镜成像和研究抑制潜在关键分子的功能。我们已经证明，九蛋白沿着微管移动，并重新定位到微管束末端被锚定的根尖外周附着物。我们还表明，伸长的上皮细胞首先形成微管的径向模式，从细胞中心向外延伸到细胞周围，微管相关分子（如CLIP-170）对于介导微管和细胞周围之间的接触很重要。这种接触可能对九蛋白的重新定位很重要，因此对微管重组也很重要。因此，Ninein和CLIP170可能在微管重组中发挥关键作用，从而也在上皮发育中发挥关键作用。其他人的研究表明，九肽对新血管、神经元、皮肤的发育和干细胞的测定都很重要。总而言之，这表明9 - in在细胞和组织发育中起着重要作用。该项目的目的是确定负责九蛋白重新定位及其在微管重组和上皮发育中的作用的机制。这个项目的发现可能会对我们理解细胞发育、干细胞命运和疾病状态产生深远的影响。

Development of elongated epithelial cells such as those of the gut and kidney involves a dramatic rearrangement of tubular structures called microtubules into apico-basal bundles that run from the apex to the base of the cells. Microtubule rearrangement is likely to be dependent on a change in the distribution of molecules such ninein that anchor one end of the microtubules and organises them into specific patterns like the apico-basal bundles and this is likely to be vital for the normal function of these cells. Defects in epithelial elongation and microtubule bundle formation lead to loss of function, abnormal cell migration and cancer. How the microtubules become organised into bundles in elongating epithelial cells is not fully understood. Most of what we do know has come from investigations of columnar shaped flat sheets of cells in culture that do not have the natural tissue architecture. Importantly, we have now established a novel live three dimensional gut organoid culture system that mimics normal gut development producing mini gut-like structures with elongated epithelial cells. This system provides a powerful new way to study microtubule reorganisation during epithelial elongation and tissue development using live microscope imaging and studies that inhibit the function of potential key molecules. We have shown that ninein travels along microtubules and relocates to apical peripheral attachments where the ends of the microtubule bundle become anchored. We have also shown that elongating epithelial cells first form a radial pattern of microtubules that run from the cell centre and out to the cell periphery and that microtubule associated molecules such as CLIP-170 are important for mediating contact between the microtubules and the cell periphery. This contact is likely to be important for the relocation of ninein and therefore also for microtubule reorganisation. Ninein and CLIP170 are therefore likely to play critical roles in microtubule reorganisations and thus also in epithelial development. Studies by others have shown that ninein is important for the development of new blood vessels, neurons, skin and for stem cell determination. All in all, this points to ninein having a major role in cell and tissue development. The aims of this project are to determine the mechanisms responsible for ninein relocation and its role in microtubule reorganisation and epithelial development. Findings from this project are likely to have far-reaching implications for our understanding of cell development, stem cell fate and diseased states.