EAGER: Topographically Induced Lateral Organization in Biomimetic Lipid Membranes

活细胞不断地经历形状变化，以促进对细胞生存至关重要的生物功能。这些形状的变化主要是由被称为细胞骨架的细胞内细丝和小管网络驱动的。在细胞重塑过程中，细胞骨架施加的力会导致细胞膜变形，细胞膜是一层薄薄的脂类和蛋白质，定义了细胞边界。重要的是，这种变形与细胞膜内分子排列和紧急功能的变化有关。然而，目前尚不清楚膜形状的变化在多大程度上可以诱导和调节分子重排，特别是在各种细胞功能中隐含的脂分子簇(或结构域)。这个早期概念探索性研究资助(AGER)项目旨在通过对膜曲率和新出现的类脂结构域进行系统的实验研究来探索这一重要现象。这将通过一种新颖的纳米柔性底物设计来实现，该底物上覆盖着一层柔软的水凝胶膜，模仿细胞细胞骨架。在所制备的底物上沉积的模型类脂膜将说明膜曲率和细胞骨架相互作用如何影响类脂域的形成、生长和稳定性。这些实验将与计算机模拟协同整合，以揭示所观察到的脂域行为背后的分子机制。在整个项目过程中，首席研究员将对研究生和本科生进行实验和计算方法方面的培训，为他们在STEM工作做好准备。该项目的调查结果将通过会议报告、同行评议出版物、开放源码数据分析软件和录像教学教程向科学界和公众传播。细胞膜具有不同的形状和曲率，其中很大一部分是由细胞骨架的动态变形驱动的，对细胞功能至关重要。细胞被认为利用膜重塑作为一种机制，将机械信号转化为成分重排和随后的生化过程。这种重排包括功能脂域的形成和稳定，这些功能域充当定位信号蛋白和维持细胞活力的平台。然而，对于膜形态如何驱动脂类结构域的结构和定位仍然缺乏清楚的了解。这个项目的主要目标是阐明在脂质膜中曲率诱导结构域图案形成的关键因素，包括局部曲率和能量惩罚。这将使用一套互补的实验、理论和计算方法，以及一种新的设计的地形结构水凝胶支架作为细胞骨架的替代品来实现。该项目的一个主要目标是优化膜的形貌和膜-支架的相互作用，以便对新出现的脂类结构域进行系统研究。其目的是确定地形和相互作用在控制脂质结构域的大小、定位和扩散方面的作用。该项目的一个显著特点是使用非镜面中子散射，辅之以理论发展和计算机模拟，以确定纳米尺度上的区域组织，即超出光学显微镜方法的分辨率。这一领域的发展将为研究纳米尺度的膜结构开辟新的途径，这些结构很少被探索，但对生物学应用至关重要，包括人工细胞和基于膜的生物传感器。该项目由分子和细胞生物科学(MCB)部门(分子生物物理和细胞动力学和功能集群)和土木、机械和制造创新部门的生物力学和机械生物学计划联合资助。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Living cells constantly undergo shape changes to facilitate biological functions that are essential to the cell survival. These shape changes are primarily driven by an intracellular network of filaments and tubules, known as the cytoskeleton. During cell reshaping, forces applied by the cytoskeleton cause deformations in the cell membrane, a thin layer of lipids and proteins defining the cell boundary. Importantly, such deformations have been linked to changes in molecular arrangements, and emergent functions, within cell membranes. However, it is unclear to what extent variations in membrane shapes can induce and regulate molecular rearrangements, particularly clusters (or domains) of lipid molecules implied in various cell functions. This EArly-concept Grant for Exploratory Research (EAGER) project aims to explore this important phenomenon through systematic experimental studies of membrane curvature and emergent lipid domains. This will be accomplished through a novel design of nanofabricated substrates, coated with a soft hydrogel film, that mimic the cell cytoskeleton. Model lipid membranes, deposited on the fabricated substrates, will illustrate how membrane curvature and cytoskeletal interactions influence the formation, growth, and stability of lipid domains. The experiments will be synergistically integrated with computer simulations to uncover the molecular mechanisms underlying the observed lipid domain behavior. Throughout the course of the project, the principal investigator will train graduate and undergraduate students in experimental and computational methods which will prepare them for careers in STEM. Findings from this project will be communicated to the scientific community and the general public through conference presentations, peer-reviewed publications, open-source data analysis software, and video-recorded instructional tutorials. Cell membranes adopt various shapes and curvatures, much of which are driven by dynamic cytoskeletal deformations and are critical to the cell function. Cells are thought to use membrane reshaping as a mechanism to translate mechanical signals into compositional rearrangements and subsequent biochemical processes. Such rearrangements include the formation and stabilization of functional lipid domains that act as platforms for localizing signaling proteins and maintaining the cell viability. However, a clear understanding of how membrane topography drives the structuring and localization of lipid domains is still lacking. The main goal of this project is to elucidate the key factors underlying curvature-induced domain patterning in lipid membranes, including local curvature and energetic penalties. This will be achieved using a suite of complementary experimental, theoretical, and computational methods along with a novel design of topographically structured hydrogel scaffolds as a proxy to the cell cytoskeleton. A primary objective of the project is to optimize membrane topography and membrane-scaffold interactions to allow systematic studies of emergent lipid domains. The aim is to identify the role of topography and interactions in controlling the size, localization, and diffusion of the lipid domains. A distinctive feature of this project is the use of off-specular neutron scattering, aided with theoretical developments and computer simulations, to determine domain organization on the nanoscale, i.e. beyond the resolution of optical microscopy methods. Developments in this area will open new avenues for investigations of nanoscale membrane structures that are seldom explored but are critical to biological applications, including artificial cells and membrane-based biosensors.This project is jointly funded by Molecular and Cellular Biosciences (MCB) Division (Molecular Biophysics and Cellular Dynamics and Function clusters) and the Biomechanics and Mechanobiology Program at Civil, Mechanical, and Manufacturing Innovation Division.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.