Differentiation Induced Changes in Centrosomes and Microtubule Organization

描述(申请人提供)：分化导致不同细胞类型的微管组织发生变化，包括上皮细胞、肌肉和神经元。这需要丧失中心体的微管组织活性，并形成非中心体微管阵列。对于这种重组是如何发生的，以及由此产生的微管的功能，我们的了解非常有限。在一定程度上，这是由于缺乏工具来观察和具体扰乱组织中的微管组织。我们工作的长期目标是了解不同的微管组织是如何在分化细胞中产生的，并揭示它们的功能。这项提议的目的是确定中心体组织活性的丧失和微管减去末端锚定蛋白如何协同诱导特定的微管阵列。根据我们的初步数据，我们假设分化导致中心体微管的成核和锚定的变化。与细胞质负端锚定蛋白结合，这允许在分化的细胞中形成特定的细胞骨架结构。我们已经开发了工具/技术，使我们能够充分表征中心体核活动、超微结构和蛋白质组成的变化，这些变化发生在表皮分化时。此外，我们还将阐明导致中心体组成和活性变化的潜在分子机制。最后，我们开发了新的小鼠品系，将用于通过特定破坏微管负端锚定蛋白来研究非中心体微管阵列在皮肤和肠道中的作用。这些研究将阐明在分化的细胞中调节中心体活动和微管重组的机制，并确定这些阵列在两个不同组织中的生理功能。

DESCRIPTION (provided by applicant): Differentiation induces a change in microtubule organization in cell types as diverse as epithelia, muscles and neurons. This requires loss of the centrosome's microtubule organizing activity and formation of non- centrosomal microtubule arrays. We have a very limited understanding of how this reorganization occurs and the functions of the resulting microtubules. In part, this is due to lack of tools to observe and specifically perturb microtubule organization in tissues. The long-term goal of our work is to understand how diverse microtubule organizations are generated in differentiated cells and to expose their functions. The objectives of this proposal are to determine how loss of centrosomal organizing activity and microtubule minus end anchoring proteins collaborate to induce specific microtubule arrays. Based on our preliminary data, we hypothesize that differentiation causes changes in both nucleation and anchoring of microtubules at centrosomes. In combination with cytoplasmic minus end anchoring proteins, this allows for specific cytoskeletal structures to form in differentiated cells. We have developed tools/technologies that will allow us to fully characterize the changes in centrosome nucleation activity, ultrastructure and protein composition that occur upon differentiation in the epidermis. In addition, we will elucidate the underlying molecular mechanisms that lead to changes in centrosome composition and activity. Finally, we have developed novel mouse lines that will be used to examine the roles of noncentrosomal microtubule arrays in both the skin and the intestine through specific disruption of microtubule minus end anchoring proteins. These studies will elucidate the mechanisms regulating centrosome activity and microtubule reorganization in differentiated cells and determine the physiologic functions of these arrays in two different tissues.