Deciphering the role of actomyosin remodeling and mechanotransduction in sea urchin skeletogenesis

后生动物中的生物吸收矿物质，并建造复杂的结构来支持和保护它们；这些生物矿物质构成了地球上生命的大部分化石记录。生物矿物通常具有超出人造材料技术水平的结构特性，这激发了仿生矿物结构的制造。由于这些原因，细胞如何控制矿物的形状和性质对发育、细胞和进化生物学家以及材料科学家来说都很有趣，然而，调控生物矿化的分子机制还远远不清楚。多个迹象表明，肌动球蛋白网络参与了真核生物的生物矿化，从钙化和硅化的单细胞生物到脊椎动物的骨骼和牙齿发育。肌动球蛋白网络是机械传感和机械转导的重要组成部分，是细胞感知和响应细胞外环境机械特性的能力。然而，目前还不清楚生物矿化的哪些方面需要机械传感和机械转导，以及哪些效应蛋白参与了这一过程。这项建议的总体目标是以海胆胚胎和骨骼细胞培养为模型，破译肌动蛋白重塑和黏附蛋白在控制矿物生长和塑造矿物形态方面的作用。我们已经确定了肌动球蛋白重塑和黏附蛋白，它们是海胆骨骼形成的关键，并参与了骨形成和动态平衡，例如RhoA相关的线圈-线圈激酶(ROCK)、CDC42和粘着斑激酶(FAK)。我们将首先利用蛋白质组学和磷酸化蛋白质组学分析确定ROCK、CDC42、FAK调控的蛋白质，并研究它们在活体海胆胚胎中的功能。然后，我们将利用海胆骨骼细胞培养进行高级时空研究，使用先进的光学和电子显微镜。我们将研究控制和上述蛋白质扰动下的骨骼生长、形态和分子标记，以深入了解生物矿物沉积和生长的分子控制。这三种PI的互补方法和专业知识将使我们能够揭示肌动蛋白重塑网络和黏附蛋白之间的相互作用，以及这些相互作用如何控制形成的生物矿物的生长和形状。

Organisms across metazoans uptake minerals and build intricate structures that support and protect them; these biominerals constitute most of the fossil record of life on earth. Biominerals often have structural properties that are beyond the state of the art of manmade materials, which inspires the fabrication of biomimetic mineral structures. For all these reasons, how cells control mineral shape and properties is fascinating for developmental, cell and evolutionary biologists as well as for material scientists, yet, the molecular mechanisms that regulate biomineralization are far from clear. Multiple indications points to the actomyosin network's involvement in biomineralization in Eukaryotes, from calcifying and silicifying unicellular organisms to vertebrate bone and teeth development. The actomyosin network is an essential part of mechanosensing and mechanotransduction, the ability of cells to sense and respond to the mechanical properties of the extracellular environment. However, it is yet unknown which aspects of biomineralization require mechanosensing and mechanotransduction and which effector proteins participate in this process. The overall goal of this proposal is to decipher the roles of actomyosin remodeling and adhesion proteins in controlling mineral growth and shaping mineral morphology using sea urchin embryos and skeletogenic cell cultures as a model. We had already identified actomyosin remodeling and adhesion proteins that are key to sea urchin skeletogenesis and participate in bone formation and homeostasis, e.g. RhoA associated coil-coil Kinade (ROCK), CDC42 and Focal adhesion kinase (FAK). We will first identify the proteins regulated by ROCK, CDC42, FAK using proteomic and phospho-proteomic analyses and we will investigate their function in live sea urchin embryos. We will then utilize sea urchin skeletogenic cell cultures to conduct high spatio-temporal studies using advanced light and electron microscopy. We will study skeletal growth, morphology and molecular markers in control and under the perturbations of the abovementioned proteins to gain insight into the molecular control of biomineral deposition and growth. The complementary approaches and expertise of the three PIs will allow us to reveal the interactions between the actomyosin remodeling network and adhesion proteins, and how these interactions control the growth and the shape of the forming biomineral.