Remodeling the microtubule cytoskeleton during epithelial cell division and differentiation

项目摘要/摘要 分裂和分化细胞需要不同排列的微管才能发挥作用，而目标是 这项建议是为了了解分化细胞如何重组它们的微管以进行分裂。有丝分裂 细胞使用中心体作为它们的微管组织中心(MTOC)来形成放射状微管阵列，该阵列 帮助将细胞分裂成两个女儿。分化细胞通常将非中心体位置指定为其MTOC。 在极化的上皮细胞中，顶膜是MTOC，它形成平行的微管阵列，这些微管阵列 对细胞极性和细胞内运输很重要。然而，在发育过程中，上皮细胞经常分裂，组织 维护和癌症，构成了一个障碍：他们必须暂时失去他们的平行微管和 重建放射状微管阵列。人们对上皮细胞如何完成这种重组知之甚少， 这对成功的细胞分裂至关重要。 这项建议将使用两个互补的模型，发育中的线虫肠道和原代人类 肠道“类器官”细胞，以揭示上皮细胞用于重塑其微管的机制 分裂的细胞骨架，以及在发育和疾病中破坏这种重塑的后果。这个 线虫胚胎肠道是一种简单的活体上皮管，易于可视化和操作，具有 经过微管重塑以进行分裂的细胞的固定数量。此外，C. 细胞分裂和微管组织的优雅是保守的，使它成为发现的理想环境 调节其他系统中微管组织的新基因和新机制。通过将费尔德曼 实验室最近开发的与经典技术相结合的技术，拟议的实验将确定 在非中心体MTOC处物理地保持和释放微管，以及导致 这一定位变化与细胞分裂相一致。这些新发现的基因和途径将是 测试原代人类肠道细胞的保守作用，以及由 微管组织紊乱。 这一建议解决了极化细胞如何重组为细胞的基本生物学问题 组织。建议的实验将覆盖整个获奖期，并在指导期间进行技术培训 这一阶段将促进独立阶段的实验。一个由专家导师和合作者组成的团队将进行培训 萨利博士提出了对这项研究的成功至关重要的新方法。此外，萨利博士还将参加 在当地会议和科学会议上，参加职业规划课程，并定期与她的导师会面 和咨询委员会讨论她的科学进步，并为工作申请和面试做准备。 萨利博士的两位导师都全力支持她成功制定她的研究计划，她将 带上她一起创办一个独立的学术研究实验室。

Project Summary/Abstract Dividing and differentiating cells require different arrangements of microtubules to function, and the goal of this proposal is to understand how differentiating cells reorganize their microtubules in order to divide. Mitotic cells use centrosomes as their microtubule organizing centers (MTOCs) to form radial microtubule arrays that help split the cell into two daughters. Differentiating cells often designate a non-centrosomal sites as their MTOC. In polarized epithelial cells, the apical membrane is the MTOC, and it forms parallel microtubules arrays that are important for cell polarity and intracellular transport. However, epithelial cells often divide in development, tissue maintenance, and cancer, presenting an obstacle: they must temporarily lose their parallel microtubules and reestablish radial microtubule arrays. Little is known about how epithelial cells accomplish this reorganization, which is critical for successful cell division. This proposal will use two complementary models, the developing C. elegans intestine and primary human intestinal “organoid” cells, to uncover the mechanisms that epithelial cells use to remodel their microtubule cytoskeleton for division, and the consequence of disrupting this remodeling in development and disease. The C. elegans embryonic intestine is a simple in vivo epithelial tube that is easy to visualize and manipulate, with a fixed number of cells that undergo microtubule remodeling to divide. In addition, many of the proteins used in C. elegans for cell division and microtubule organization are conserved, making it an ideal context for discovering new genes and mechanisms that regulate microtubule organization in other systems. By combining the Feldman lab’s recently developed techniques with classic ones, the proposed experiments will identify the factors that physically hold and release microtubules at the non-centrosomal MTOC, and the molecular signals that cause this localization to change concordant with cell division. These newly discovered genes and pathways will be tested for a conserved role in primary human intestinal cells, and for cancer-related defects resulting from disrupted microtubule organization. This proposal addresses the fundamental biological question of how polarized cells reorganize for cell division. The proposed experiments will cover the entire award period, and technical training during the mentored phase will facilitate experiments in the independent phase. A team of expert mentors and collaborators will train Dr. Sallee in new methods that are critical to the success of this research. In addition, Dr. Sallee will participate in local meetings and scientific conferences, attend career planning courses, and meet regularly with her mentors and advisory committee to discuss her scientific progress and to prepare for job applications and interviews. Both of Dr. Sallee’s mentors are fully committed to her success in establishing her research plan that she will take with her to start an independent academic research lab.