The keratin-filament cycle of assembly and disassembly

角蛋白丝的组装和拆卸循环

• 批准号: 196052332
• 负责人: Professor Dr. Rudolf E. Leube
• 金额: --
• 依托单位: Institut für Molekulare und Zelluläre Anatomie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2011
• 资助国家: 德国
• 起止时间: 2010-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/196052332?language=en
• 关键词: keratin; filament; cycle; assembly; disassembly

Keratin intermediate filaments are major constituents of the epithelial cytoskeleton. Their organizing function of cellular space is important for diverse epithelial functions requiring a high degree of keratin network plasticity. We have recently proposed that a continuous cycle of keratin filament assembly and disassembly affords rapid and precise adaptation to the different functional requirements and situations. The goal of the first project phase was to establish and apply tools for proving/disproving our concept of the keratin cycle and to examine its proposed aspects. Our experiments observing fluorescently-tagged keratin polypeptides fully supported the main features of the keratin cycle. Furthermore, they allowed us to quantitatively measure keratin network properties, speed of keratin movement and local turnover. We were further able to demonstrate that the new tools are capable detecting slight differences in keratin cycling taking place after seeding of cells and upon growth factor addition, migration induction, keratin phosphorylation, and downregulation of cytoskeletal cross linkers as well as in response to altered substrate stiffness. The second project phase intends to move from a descriptive stage of the keratin cycle to a molecular understanding. To achieve this, we will (i) further refine methods and tools for monitoring and manipulating the keratin cycle and its modulators, (ii) focus on identifying and characterizing the molecular motors involved in keratin movement and tension generation within the keratin network, and (iii) examine modulators of the keratin cycle that are identified in genetic screens. We will apply the newly developed image recording and analysis routines on high resolution microscopic image sets of improved fluorescent reporter cell lines to systematically study the effect of downregulating all of the relevant motor proteins and other modulators of the keratin cycle on keratin network organization, dynamics and function. Consequences on epithelial function, i.e. cell migration, cellular viscoelasticity and stiffness, proliferation and stress response will be correlated to these findings.Our long-term goal is to understand molecular mechanisms of keratin network plasticity in the context of epithelial pathophysiology.

角蛋白中间丝是上皮细胞骨架的主要组成部分。它们对细胞空间的组织功能对于需要高度角蛋白网络可塑性的多种上皮功能至关重要。我们最近提出，角蛋白丝的组装和拆卸的连续循环提供了快速和精确的适应不同的功能需求和情况。第一个项目阶段的目标是建立和应用工具来证明/反驳我们的角蛋白周期概念，并检查其提出的方面。我们观察荧光标记的角蛋白多肽的实验完全支持角蛋白循环的主要特征。此外，它们使我们能够定量测量角蛋白网络特性、角蛋白运动速度和局部周转。我们进一步证明，新工具能够检测细胞播种后、生长因子添加、迁移诱导、角蛋白磷酸化、细胞骨架交联剂下调以及底物硬度改变后发生的角蛋白循环的细微差异。第二个项目阶段打算从角蛋白循环的描述阶段转向分子理解。为了实现这一目标，我们将(i)进一步完善监测和操纵角蛋白周期及其调节剂的方法和工具，（ii）专注于识别和表征参与角蛋白运动和角蛋白网络中张力产生的分子马达，以及（iii）检查在遗传筛选中确定的角蛋白周期调节剂。我们将应用新开发的图像记录和分析程序，对改进的荧光报告细胞系的高分辨率显微图像集进行系统研究，下调所有相关的运动蛋白和角蛋白周期的其他调节剂对角蛋白网络组织、动力学和功能的影响。对上皮功能的影响，即细胞迁移、细胞粘弹性和刚度、增殖和应激反应将与这些发现相关。我们的长期目标是在上皮病理生理学的背景下了解角蛋白网络可塑性的分子机制。

项目成果

Flux-based 3D segmentation of keratin intermediate filaments in confocal laser scanning microscopy

共焦激光扫描显微镜中基于通量的角蛋白中间丝 3D 分割

• DOI: 10.1109/isbi.2012.6235510
• 发表时间: 2012
• 期刊: 2012 9th IEEE International Symposium on Biomedical Imaging (ISBI)
• 作者: Gerlind Herberich;Andreas Friedrich;Til Aach;Reinhard Windoffer;Rudolf E. Leube
• 通讯作者: Rudolf E. Leube

Signal and Noise Modeling in Confocal Laser Scanning Fluorescence Microscopy

共焦激光扫描荧光显微镜中的信号和噪声建模

• DOI: 10.1007/978-3-642-33415-3_47
• 发表时间: 2012
• 期刊: Medical image computing and computer-assisted intervention : MICCAI ... International Conference on Medical Image Computing and Computer-Assisted Intervention
• 作者: Gerlind Herberich;Reinhard Windoffer;Rudolf E. Leube;Til Aach
• 通讯作者: Til Aach

Measuring the regulation of keratin filament network dynamics

• DOI: 10.1073/pnas.1306020110
• 发表时间: 2013-06-25
• 期刊: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• 影响因子: 11.1
• 作者: Moch, Marcin;Herberich, Gerlind;Windoffer, Reinhard
• 通讯作者: Windoffer, Reinhard