Fusion and fission of cellular membranes involve dramatic, protein-mediated changes in membrane curvature. Many of the experimental methods useful for investigating curvature sensing or generation require specialized equipment. We have developed a system based on supported lipid bilayers (SLBs) in which lipid tubules are simple to produce and several types of membrane remodeling events can be readily imaged using widely available instrumentation (e.g., tubule fission and/or membrane budding). Briefly, high ionic strength during lipid bilayer deposition results in incorporation of excess lipids in the SLB. After sequentially washing with water and physiological ionic strength buffer solutions, lipid tubules form spontaneously. We find that tubule formation results from solution-dependent spreading of the SLB; washing from water into physiological ionic strength buffer solution leads to expansion of the bilayer and formation of tubules. Conversely, washing from physiological buffer into water results in contraction of the membrane and loss of tubules. We demonstrate the utility of these supported tubulated bilayers, termed "STuBs," with an investigation of Sar1B, a small Ras family G-protein known to influence membrane curvature. The addition of Sar1B to STuBs results in dramatic changes in tubule topology and eventual tubule fission. Overall, STuBs are a simple experimental system, useful for monitoring protein-mediated effects on membrane topology in real time, under physiologically relevant conditions.

中文翻译：

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一个简单的支持管式双层系统，用于评估蛋白质介导的膜重塑。

细胞膜的融合和裂变涉及膜曲率的急剧的，蛋白质介导的变化。许多用于研究曲率感测或生成的实验方法都需要专用设备。我们已经开发了基于支持的脂质双层（SLB）的系统，其中脂质小管易于生产，并且可以使用广泛使用的仪器（例如，小管裂变和/或膜芽接）轻松地对几种类型的膜重塑事件进行成像。简而言之，脂质双层沉积过程中的高离子强度导致SLB中掺入了过多的脂质。依次用水和生理离子强度缓冲溶液洗涤后，脂质小管即刻形成。我们发现，小管形成是由于SLB的溶液依赖性扩散而引起的。从水中清洗到生理离子强度缓冲溶液中会导致双层膨胀并形成小管。相反，从生理缓冲液清洗到水中会导致膜的收缩和小管的丢失。我们通过对Sar1B（一种已知影响膜曲率的小Ras家族G蛋白）的研究，证明了这些支持的管状双分子层（称为“ STuBs”）的实用性。在STuB中添加Sar1B会导致小管拓扑结构发生巨大变化，并最终导致小管裂变。总体而言，STuB是一个简单的实验系统，可用于在生理相关条件下实时监测蛋白质介导的对膜拓扑的影响。从生理缓冲液清洗到水中会导致膜收缩和小管丢失。我们通过对Sar1B（一种已知影响膜曲率的小Ras家族G蛋白）的研究，证明了这些支持的管状双分子层（称为“ STuBs”）的实用性。在STuB中添加Sar1B会导致肾小管拓扑结构发生巨大变化，并最终导致肾小管裂变。总体而言，STuB是一个简单的实验系统，可用于在生理相关条件下实时监测蛋白质介导的对膜拓扑的影响。从生理缓冲液冲洗到水中会导致膜收缩和小管丢失。我们通过对Sar1B（一种已知影响膜曲率的小Ras家族G蛋白）的研究，证明了这些支持的管状双分子层（称为“ STuBs”）的实用性。在STuB中添加Sar1B会导致肾小管拓扑结构发生巨大变化，并最终导致肾小管裂变。总体而言，STuB是一个简单的实验系统，可用于在生理相关条件下实时监测蛋白质介导的对膜拓扑的影响。在STuB中添加Sar1B会导致肾小管拓扑结构发生巨大变化，并最终导致肾小管裂变。总体而言，STuB是一个简单的实验系统，可用于在生理相关条件下实时监测蛋白质介导的对膜拓扑的影响。在STuB中添加Sar1B会导致肾小管拓扑结构发生巨大变化，并最终导致肾小管裂变。总体而言，STuB是一个简单的实验系统，可用于在生理相关条件下实时监测蛋白质介导的对膜拓扑的影响。