Supracellular synchronization of actomyosin contractility during collective neural crest cell durotaxis

细胞集体迁移是一个在发育、再生和转移形成过程中高度协调的过程。神经嵴细胞是一种侵袭性间充质细胞群，它们不仅会根据化学因素集体迁移，还会沿着下层基质的刚度梯度迁移，这一过程被称为硬性迁移。集体趋动需要非凡水平的机械感应、对称性破坏、细胞间通讯和同步肌动球蛋白收缩，以协调定向的细胞外运动。然而，集体迁移的神经嵴细胞如何沟通，以便在硬移过程中跨几个细胞边界同步它们的突起和产生力的机器，仍然不清楚。利用非洲爪蟾胚胎作为模型系统，我们试图通过研究在体外和体内产生力的非肌肉肌球蛋白II的协同作用、通过间隙连接的细胞间通信和钙增殖来破译这些过程。在我的博士研究中，我在体外证明了在单细胞的力产生和形态发生过程中，非肌球蛋白II亲缘物A和B的内在特征是互补的。虽然A线提供快速的牵引力，这是启动整个细胞体收缩机制所必需的，但B线逐渐稳定亚细胞区域中预先启动的收缩，从而自主地在整个细胞体中产生极化的肌动球蛋白网络。虽然这个概念可以解释单个迁移细胞的形态发生，但在本项目中，我假设类似的原理可以应用于解释多细胞神经嵴细胞簇在体内的集体迁移性。为了研究在多个相邻细胞间介导同步肌动球蛋白收缩的途径，我们将重点关注通过间隙连接的细胞间通信，连接相邻细胞的细胞质的跨膜通道，允许信号分子和离子的交换。来自宿主实验室的初步数据表明，间隙连接参与了集体神经嵴迁移。考虑到不同的非肌肉肌球蛋白II类似物，我们将揭示间隙连接细胞间通信是否介导收缩机制的同步，以及它如何影响极化肌动球蛋白网络的产生。为了确定可能通过间隙连接传播并影响肌动球蛋白细胞骨架的信号，我们将重点关注钙离子和ip3介导的钙从内部储存中释放。钙是一种众所周知的肌动球蛋白刺激剂，来自宿主实验室的初步数据表明，IP3参与了集体神经嵴迁移过程中的钙信号传导。我们希望这个多学科项目将为一个尚未解决的核心问题提供答案：在集体细胞迁移过程中，单个细胞是如何协调的。

Collective cell migration is a highly coordinated process during development, regeneration and metastasis formation. Neural crest cells, an invasive mesenchymal cell population, collectively migrate not only in response to chemical factors but also along stiffness gradients in the underlying substratum, a process called durotaxis. Collective durotaxis requires extraordinary levels of mechanosensing, symmetry breaking, intercellular communication and synchronized actomyosin contraction, to coordinate a directional supracellular movement. However, how collectively migrating neural crest cells communicate, in order to synchronize their protrusive and force-generating machineries across several cell borders during durotaxis, is still not understood. Using Xenopus embryos as a model system, we seek to decipher these processes by investigating the synergy of force-generating nonmuscle myosin II paralogs, intercellular communication via gap junctions, and calcium propagation, in vitro and in vivo. In my PhD research, I showed that in vitro, the intrinsic features of the nonmuscle myosin II paralogs A and B complement each other during force generation and morphogenesis of single cells. While paralog A provides rapid tugging forces, necessary to initiate the contractile machinery throughout the cell body, paralog B gradually stabilizes pre-initiated contractions in subcellular regions to autonomously generate a polarized actomyosin network across the cell body. Although this concept can explain the morphogenesis of individual migrating cells, in this project I hypothesize that similar principles can be applied to explain collective durotaxis in vivo of multicellular neural crest cell cluster. To investigate the pathway that mediates synchronized actomyosin contraction across multiple adjacent cells, we will focus on intercellular communication via gap junctions, transmembrane channels that connect the cytoplasm of adjacent cells, allowing exchange of signalling molecules and ions. Preliminary data from the host lab suggest that gap junctions are involved in collective neural crest migration. We will unravel if gap junctional intercellular communication mediates the synchronization of the contractile machinery and how it influences the generation of a polarized actomyosin network, considering the distinct nonmuscle myosin II paralogs. In order to identify possible signals that could be propagated via gap junctions and influence the actomyosin cytoskeleton, we will focus on calcium ions and IP3-mediated calcium release from internal stores. Calcium is a well-known actomyosin stimulator and preliminary data from the host lab suggest that IP3 is involved in calcium signalling during collective neural crest migration. We expect that this multidisciplinary project will provide answers to a central yet unresolved question: how individual cells are coordinated during collective cell migration.