Mitotic cell mechanics in a tissue context

对称和非对称细胞分裂都需要一套复杂的分子过程来确保基于微管的纺锤体相对于皮质信号的正确方向。在不对称分裂中，细胞首先根据内部或外部信号建立极性轴。然后星体微管读取这一信息，定位纺锤体，在两个子细胞中不对称地分离细胞命运决定因素。在对称分裂中，纺锤体通常读取顶基极性线索和有丝分裂细胞形状，以便以这样的方式排列以分裂释放组织张力。在这两种情况下，纺锤体取向可能对高保真染色体分离很重要。因此，了解有丝分裂过程中的力学作用是细胞生物学的一个基本问题。此外，目前认为，对称与非对称细胞分裂的控制比例在干细胞稳态中起着关键作用，这一现象被认为在癌症中被错误调节。虽然在确定调节细胞分裂的生化信号通路方面已经取得了很大进展，但对正常上皮细胞和干细胞感知、响应和抵抗外力的机制知之甚少。直到最近，力在单个贴壁细胞纺锤体取向中的作用才被研究，并且已知机械应力是参与上皮细胞调节的主要外部信号。我们的初步数据现在表明，力在组织的纺锤体取向中也起着重要作用。在这项工作的基础上，我们计划测试纺锤体取向在对称和非对称分裂期间如何响应外力，并确定所涉及的分子机制。为此，我们将探索有丝分裂细胞感知、响应和抵抗机械力的分子和细胞机制，使用Baum和Charras实验室开发的两个互补实验系统：i)一种能够对MDCK上皮单层进行机械扰动的装置，ii)果蝇，一种可以很容易地结合遗传学、机械扰动和实时成像的组织。通过这种方式，我们期望确定保守的分子机制，确保细胞分裂在力学变化的上皮环境中高保真地发生。我们将进一步测试力是否也在不对称分裂中发挥作用（作为干细胞分裂的简单模型）。我们期望这项工作对我们对细胞分裂、组织稳态、干细胞生物学和再生医学领域的基本问题的理解产生重大影响。有几条线索的证据清楚地表明，弥合我们对细胞分裂中力学作用的理解中的这一差距的重要性。首先，有丝分裂过程涉及细胞形状和皮质硬度的剧烈活跃变化，当这些变化受到干扰时可能导致细胞分裂失败和染色体错误分离；可能会导致癌症的发展其次，细胞在机械软介质中分裂和形成集落的能力是细胞转化和恶性的关键测试，这意味着力学在细胞分裂中发挥了作用。第三，在细胞培养中，有丝分裂纺锤体通常沿最大拉伸力的轴平行于基底平面。第四，拉伸应力可能会引发增加干细胞数量的对称分裂。通过这种方式，组织可以通过张力控制增殖和定向细胞分裂来增加细胞数量，从而直接响应对更多细胞的需求，从而确保整个组织的张力得到缓解。

Both symmetric and asymmetric cell divisions require a complex set of molecular processes to ensure the proper orientation of the microtubule-based spindle with respect to cortical cues. In asymmetric divisions, cells first establish an axis of polarity in response to internal or external cues. This is then read by astral microtubules to orient the spindle to segregate cell fate determinants asymmetrically in the two daughter cells. In symmetric divisions, the spindle typically reads apical-basal polarity cues and mitotic cell shape in order to align in such a way as to divide to release tissue strain. In both cases, spindle orientation is likely to be important for high fidelity chromosome segregation. Understanding the role of mechanics in mitosis is therefore a fundamental problem in cell biology. In addition, it is currently thought that controlled ratio of symmetric versus asymmetric cell division plays a critical role in stem cell homeostasis, a phenomenon thought to be misregulated in cancer.While much progress has been made in identifying biochemical signalling pathways that regulate cell division in general, little is known about the mechanisms by which normal epithelial cells and stem cells sense, respond to and resist forces. Only recently the role of force has been studied in spindle orientation of individual adherent cells, and mechanical stress is known to be a major external signal involved in epithelial cell regulation. Our preliminary data now show that forces also play a significant role in spindle orientation in tissues. Building on this work we plan to test how spindle orientation responds to external force during symmetric and asymmetric divisions and to identify the molecular machinery involved.To do so we will explore the molecular and cellular mechanisms by which mitotic cells sense, respond to and resist mechanical forces using two complementary experimental systems developed in the Baum and Charras labs: i) a device that enables the mechanical perturbation of MDCK epithelial monolayers, and ii) the Drosophila notum, a tissue where genetics, mechanical perturbations and live imaging can be readily combined. In this way, we expect to identify conserved molecular mechanisms that ensure that cell division occurs with high fidelity in the context of an epithelium subject tochanges in mechanics. We will further test whether force also a play a role in asymmetric divisions (as a simple model of stem cell divisions).We expect this work to have a significant impact on our understanding of fundamental questions in the fields of cell division, tissue homeostasis, stem cell biology and regenerative medicine. There are several lines of evidence that make clear the importance of bridging this gap in our understanding of the role of mechanics in cell division. First, passage through mitosis involves dramatic active changes in cell shape and cortical rigidity, which when perturbed may lead to cell division failure and chromosome mis-segregation; potentially contributing to cancer development. Second, the ability of a cell to divide and form a colony in a mechanically soft medium is a key test of cellular transformation and malignancy, implying a role for mechanics in cell division. Third, in cell culture the mitotic spindle typically aligns parallel to a substrate plane along the axis of greatest tensile force. Fourth, it is possible that symmetrical divisions which tend to increase the number of stem cells will be triggered by tensile stress. In this way tissues may respond directly to the need for more cells by increasing the number of cells via tensile-controlled proliferation combined to oriented cell division ensuring relieve of the tension across the tissue.