Symmetry breaking and polarization of cell in 3D environments

 描述（由申请人提供）：细胞外基质（ECM）的力学、3D结构和生物化学组成显著影响多种生物学结果，包括细胞极化、分化、凋亡、迁移和增殖。例如，ECM对齐触发极化细胞形态并促进远离原发性肿瘤的迁移。然而，它仍然知之甚少，细胞如何时空整合3D ECM线索，以选择性地编排下游信号。本研究的目的是进一步了解这一过程。具体来说，我将研究间充质细胞如何自发地建立前后细胞极性在三维ECM环境。对于这些手段，我将评估时空演变的细胞胶原蛋白接口，整合素激活，Rho家族GTdR信号，和细胞骨架动力学，在间充质细胞进行三维极化。我推测，ECM纤维定向正常的细胞表面将触发整合素聚集和成熟的基质粘附的形成（目的1）。此外，我假设Rho家族GTdR的激活将分为局灶性粘附依赖性和独立性机制（目标2），这将导致ECM环境依赖性调节肌动蛋白聚合，捆绑，树突状分支，肌动球蛋白收缩和解聚（目标3）。这将通过嵌入重建的ECM样环境中的间充质细胞的光片成像、急性药理学扰动和ECM纤维取向、细胞形状、信号转导和细胞骨架动力学的计算机视觉分析来实现。这项研究将作为进一步研究的基础，可以评估生理和病理生理ECM状态下的3D时空细胞信号。

 DESCRIPTION (provided by applicant): The mechanics, 3D structure, and biochemical composition of the extracellular matrix (ECM) significantly influences diverse biological outcomes, including cellular polarization, differentiation, apoptosis, migration, and proliferation For example, ECM alignment triggers polarized cell morphologies and facilitates migration away from the primary tumor. However, it remains poorly understood how cells spatiotemporally integrate 3D ECM cues in order to selectively orchestrate downstream signaling. The objective of the proposed research here aims to further our understanding this process. Specifically, I will investigate how mesenchymal cells spontaneously establish anteroposterior cell polarity in 3D ECM environments. Towards these means, I will evaluate the spatiotemporal evolution of cell-collagen interfaces, integrin activation, Rho family GTPase signaling, and cytoskeletal dynamics, in mesenchymal cells undergoing 3D polarization. I hypothesize that ECM fibers oriented normal to the cell surface will trigger integrin clustering and the formation of mature matrix adhesions (Aim 1). Furthermore, I hypothesize that Rho family GTPase activation will be bifurcated into focal adhesion-dependent and independent regimes (Aim 2), and that this will result in ECM context-dependent modulation of actin polymerization, bundling, dendritic branching, actomyosin contraction, and depolymerization (Aim 3). This will be accomplished by light- sheet imaging of mesenchymal cells embedded within reconstituted ECM-like environments, acute pharmacological perturbations, and computer vision analysis of ECM fiber orientation, cell shape, signal transduction, and cytoskeletal dynamics. This research will serve as the foundation from which further studies can evaluate 3D spatiotemporal cell signaling in physiological and pathophysiological ECM states.