Physical and Chemical Properties of Lipid Rafts

描述（由申请人提供）：脂筏是生物膜中一个重要的新概念，但仍存在争议，因为此时细胞生物学家对筏的定义-抗洗涤剂膜和胆固醇提取诱导的筏功能废除-没有明确的细胞相关性，如通过显微镜方法确定的。在具体目标I中，我们将比较应用于活细胞的复杂和高度适当的光显微镜方法的结果与细胞膜中脂质微区的三种极端模型的预测。这些方法包括荧光相关光谱、单粒子跟踪、单染料跟踪和单域成像。模型和实验之间的比较的结果应该导致相当大的改进，我们的观点的脂质微区细胞表面。具体目标II旨在探索可以提供功能的筏的物理和化学性质。跨双层通讯以及为什么某些分子组分存在于筏中是当今该领域尚未回答的关键问题。使用不对称的平面支持的双层，我们将确定如何液体有序域的外单层与内单层沟通，以促进筏介导的信号转导过程。确定各种筏优先和交联膜组件到液体有序域和筏居民组件之间的竞争，在这些域中的网站的分区的因素也将被定义。由于筏洗涤剂抗性的概念是如此关键的概念，我们将图像的筏域存在于平面支撑膜使用荧光显微镜的各种洗涤剂的行动。

DESCRIPTION (provided by applicant): Lipid rafts are an important new concept in biomembranes but one that remains controversial because at this juncture the cell biologist's definitions of rafts--detergent resistant membranes and cholesterol extraction-induced abrogation of raft function--have no unambiguous cellular correlate as determined by microscopic methods. In Specific Aim I, we will compare the results of sophisticated and highly appropriate light microscopic methods applied to living cells to the predictions of three extreme models for lipid microdomains in cell membranes. The methods include fluorescence correlation spectroscopy, single particle tracking, single dye tracing, and single domain imaging. The results of the comparison between models and experiments should lead to a considerable refinement in our views of lipid microdomains on cell surfaces. Specific Aim II is directed towards exploring the physical and chemical properties of rafts that could provide functionality. Transbilayer communication and why certain molecular components reside in rafts are key unanswered questions in the field today. Using asymmetric planar supported bilayers, we will determine how liquid-ordered domains on the outer monolayer communicate with the inner monolayer to facilitate raft-mediated signal transduction processes. The factors that determine partitioning of various raft-preferring and cross-linked membrane components into liquid-ordered domains and the competition between raft-resident components for sites in these domains will also be defined. Because the concept of raft detergent resistance is so crucial to the concept, we will image the action of various detergents on raft domains existing in planar supported membranes using fluorescence microscopy.