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

肠和肾等细长上皮细胞的发育涉及称为微管的管状结构戏剧性地重新排列成从细胞顶端到基部的顶端-基部束。微管重排很可能依赖于分子分布的变化，例如锚在微管一端并将它们组织成特定的模式，如顶端-基底束，这可能对这些细胞的正常功能至关重要。上皮延长和微管束形成的缺陷导致功能丧失、异常细胞迁移和癌症。微管是如何在伸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对培养的柱状扁平细胞的研究，这些细胞不具有天然的组织结构。重要的是，我们现在已经建立了一种新的活的三维肠道类器官培养系统，其模拟正常的肠道发育，产生具有伸长的上皮细胞的迷你肠道样结构。该系统提供了一种强大的新方法来研究上皮细胞伸长和组织发育过程中的微管重组，使用实时显微镜成像和抑制潜在关键分子功能的研究。我们已经表明，ninein旅行沿着微管和重新定位到顶端周边附件的微管束的两端成为锚定。我们还表明，伸长的上皮细胞首先形成从细胞中心向外延伸到细胞外周的微管的放射状图案，并且微管相关分子如CLIP-170对于介导微管和细胞外周之间的接触是重要的。这种接触可能是重要的搬迁ninein，因此也为微管重组。因此，Ninein和CLIP 170可能在微管重组中发挥关键作用，因此也在上皮发育中发挥关键作用。其他人的研究表明，ninein对新血管、神经元、皮肤的发育和干细胞的确定都很重要。总而言之，这表明ninein在细胞和组织发育中起着重要作用。该项目的目的是确定ninein重新定位的机制及其在微管重组和上皮发育中的作用。该项目的发现可能对我们理解细胞发育、干细胞命运和疾病状态产生深远影响。

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.