Role of Nucleo-cytoskeleton Interactions in Cell Migration

 描述（由申请人提供）：几乎所有细胞都将其细胞核定位在反映细胞和组织功能的特定位置。参与核定位途径的蛋白质的破坏导致细胞生理学改变和人类疾病，包括肌营养不良症、心肌病、无脑畸形、小脑共济失调和肌张力障碍。核定位如何影响细胞功能尚不清楚。我们的目标是探讨迁移细胞中细胞核位置的机制和功能，其中细胞核在细胞区域中的位置是一致的。我们已经建立了一个模型受伤的单层系统，其中核运动建立了这个后方的位置，可以通过外部因素刺激和定量测量。我们还将开发一种新的方法，人工取代细胞核在粘附 细胞的离心力，并将我们的研究扩展到三维迁移系统，其中核运动可能是迁移的速率限制。我们早期的研究表明，核后部的位置依赖于肌动蛋白和肌球蛋白，需要在肌动蛋白索和核之间建立连接。这种连接是由LINC复合物组分nesprin-2G在外核膜和SUN 2在内核膜中的聚集介导的。这种聚集导致形成类似于“核粘连”的结构，我们将其称为TAN系，用于跨膜肌动蛋白相关的核系。我们将进一步探索TAN线的组装和功能，通过追求令人兴奋的初步研究表明，除了机械耦合细胞核的肌动蛋白细胞骨架，TAN线表现出影响核运动的机械化学信号。通过我们人工置换细胞核的新方法，我们发现了令人惊讶的证据，即细胞通过肌动蛋白或微管机制定位其细胞核，这取决于细胞核是否被置换到细胞的前部或后部。我们将使用这个系统来定义一种新的LINC复合物如何与微管和微管马达相关联，并测试细胞在细胞迁移的极化和活跃阶段是否在LINC复合物之间切换。为了了解核位置如何影响迁移，我们将测试核与局灶性粘连耦合并通过基于内部张力的机制调节其动力学的假设。最后，我们将探索不同LINC复合物对3D基质中细胞迁移的贡献，并测试特定假设，即当细胞核变形以挤压基质中的小孔时，可能需要不同的LINC复合物。这些研究将揭示LINC复合物定位细胞核的新机制，并为细胞核定位如何影响细胞迁移提供新的见解。这些信息将有助于了解与核定位途径相关的疾病是如何起源的，以便设计有效的治疗方法。

 DESCRIPTION (provided by applicant): Virtually all cells position their nucleus in specific locations that reflect cell and tissue function. Disruption of proteins involved in nuclear positioning pathways leads to altered cell physiology and human diseases including muscular dystrophies, cardiomyopathy, lissencephaly, cerebellar ataxia and dystonia. How nuclear positioning contributes to cell function is unclear. Our goal is to explore the mechanisms and function of nuclear position in migrating cells where nuclei are consistently positioned in the rea of cells. We have developed a model wounded monolayer system where nuclear movement establish this rearward position and can stimulated by external factors and measured quantitatively. We will also developed a novel method of artificially displacing nuclei in adherent cells by centrifugal force and will extend our studies to 3D migration systems where nuclear movements may be rate limiting for migration. Our earlier studies established that rearward nuclear position is actin- and myosin-dependent and requires the establishment of a connection between actin cables and the nucleus. This connection is mediated by the clustering of LINC complex components nesprin-2G in the outer nuclear membrane and SUN2 in the inner nuclear membrane. This clustering results in the formation of structures that resemble "nuclear adhesions" that we termed TAN lines for transmembrane actin-associated nuclear lines. We will further explore the assemble and function of TAN lines by pursuing exciting preliminary studies that suggest that in addition to mechanically coupling nuclei to the actin cytoskeleton, TAN lines exhibit mechanochemical signaling that affects nuclear movement. With our new method for artificially displacing nuclei, we have found surprising evidence that cells position their nuclei y actin or microtubule mechanisms depending on whether nuclei are displaced to the front of rear of the cells. We will use this system to define how a novel LINC complex associate with microtubules and microtubule motors and test whether cells switch between LINC complexes during polarization and active phases of cell migration. To understand how nuclear position influences migration, we will test the hypothesis that nuclei are coupled to focal adhesions and modulate their dynamics by an internal tension based mechanism. Lastly, we will explore the contribution of different LINC complexes to cell migration in 3D matrices and test the specific hypothesis that different LINC complexes may be required when nuclei deform to squeeze through small pores in the matrix. These studies will uncover novel mechanisms of LINC complex positioning of nuclei and provide new insights into how nuclear positioning influences cell migration. This information will contribute to understanding of how diseases associated with nuclear positioning pathways originate so that effective treatments can be designed.