Cytoskeleton, Nucleus and Integrin Recycling in Cell Migration

摘要 后生动物细胞迁移有助于发育和动态平衡过程，包括组织和 器官、伤口愈合、组织更新和免疫反应。细胞迁移的改变会导致发育缺陷， 炎症、持续性伤口和转移。因此，破译细胞迁移的机制具有广泛的意义 了解生理和病理过程。相当大的努力使人们对 一些有助于细胞迁移的关键过程，包括前沿的突出和形成 粘连。细胞迁移的其他重要方面很少受到关注，我们对细胞迁移的理解存在差距 它们对细胞迁移的机制和贡献。这项提议试图填补这两个这样的知识空白 过程：核定位和整合素回收。这两个过程对细胞迁移都是必不可少的，但都是机械性的 对于他们是如何做到这一点的，人们缺乏了解。这项提议将建立在先前两个项目中取得的成果的基础上 在NIGMS支持下的甘德森实验室。对于核定位，我们将研究链接器如何 核骨架和细胞骨架(LINC)复合体是机械加固的，以抵抗移动所需的巨大力 并将使用和开发新的张力传感器来直接测量原子核上的力。我们将决定 LINC复合体如何选择肌动蛋白细丝或微管进行核运动以及 这些相互作用导致了不同的细胞迁移模式。我们将检验新的假设，即原子核作为一个 肌动蛋白细丝的“张力电阻”，以及微管运输的一个极性因素。对于整合素回收，我们将测试 总的假设是循环使用的整合素以一种活跃的构象移动，这会导致新的黏附形成 在靠近前沿的地方以极化的方式。我们将使用新的整合素探针和新的整合素回收系统来 确定整合素胞吐的部位及其与新形成的粘连的关系。在细胞迁移过程中，我们将确定 回收的整合素是否来自细胞后部，并有助于靠近前沿的新生黏附形成。 我们将确定我们假设的负责粘连极化改造的微管机制。 从回收的整合素中提取整合素，并测试回收的整合素在整合素信号中发挥作用的可能性。我们的建议 对这两个过程的研究将促进对细胞迁移的基本机制和潜力的理解 确定在细胞迁移出错的情况下进行干预的新目标。拟议中的研究还将提供丰富的 为博士后研究员和研究生提供了推进培训和开发自己项目的场所。

SUMMARY Metazoan cell migration contributes to developmental and homeostatic processes, including formation of tissues and organs, wound healing, tissue renewal and immune responses. Alterations in cell migration lead to developmental defects, inflammation, persistent wounds and metastasis. Deciphering mechanisms of cell migration thus has broad significance for understanding both physiological and pathological processes. Considerable effort has led to deep understanding of some of the critical processes that contribute to cell migration, including protrusion of the leading edge and formation of adhesions. Other important aspects of cell migration have received scant attention and there is a gap in our understanding of their mechanism and contribution to cell migration. This proposal seeks to fill this knowledge gap for two such processes: nuclear positioning and integrin recycling. Both these processes are essential for cell migration, but mechanistic understanding of how they do so is lacking. This proposal will build on results previously obtained in two previous projects in the Gundersen laboratory supported by NIGMS. For nuclear positioning, we will examine how the linker of nucleoskeleton and cytoskeleton (LINC) complex is mechanically reinforced to resist the large forces necessary to move the nucleus and will use and develop new tension sensors to directly measure forces on the nucleus. We will determine how the LINC complex selects actin filament or microtubules for nuclear movement and the functional consequences of these interactions for different modes of cell migration. We will test new hypotheses that the nucleus functions as a “tension resistor” for actin filaments and as a polarity factor for microtubule trafficking. For integrin recycling, we will test the overall hypothesis that recycled integrins travel in an active conformation and that this seeds new adhesion formation in a polarized manner near the leading edge. We will use new integrin probes and a new integrin recycling system to identify sites of integrin exocytosis and their relationship to newly formed adhesions. In migrating cells, we will determine whether recycled integrin derives from the cell rear and contributes to nascent adhesion formation near the leading edge. We will identify the microtubule machinery that we hypothesize is responsible for the polarized reformation of adhesions from recycled integrins and test the possibility that the recycled integrin plays a role in integrin signaling. Our proposed studies on these two processes will advance understanding of the basic mechanisms of cell migration and potential identify new targets for intervening in cases when cell migration goes awry. The proposed studies will also provide fertile ground for postdoctoral fellows and graduate students to advance their training and develop their own projects.