Geometry-dependent assembly of the septin cytoskeleton

细胞形态与功能密切相关，可以用血浆来描述 膜曲率细胞曲率的许多变化发生在微米尺度上， 蛋白质是纳米级的，这就提出了细胞如何感知、控制和 使用微米级的几何形状。septins是一个保守的，形成细胞毒性的家族， 这些蛋白质优先聚集在微米级膜曲率的位点。Septins 聚集在许多弯曲的表面上，包括在细胞动力学沟，树突棘， 膜泡，围绕细胞内细菌和纤毛和鞭毛的基部。鉴于这些 不同的细胞环境，septins的功能障碍与多种人类疾病有关，包括 许多癌症、神经病和不孕症。在微米级膜弯曲的部位， septins可以影响蛋白质在膜中的扩散，作为支架， 信号蛋白，并影响细胞皮层的硬度。如何弯曲隔垫 组装形成并将信号蛋白募集到局部膜上对于理解 分隔符如何将单元格几何形状与响应关联起来。隔蛋白丝组装发生在 脂质双层或其他细胞骨架网络上的短（~24- 32 nm）寡聚体棒的退火。 我们假设细胞调节膜亲和力、长度、密度和几何形状， 以曲率依赖的方式排列隔片。本提案的目的是 确定引导隔片在曲面上组装的机制并测量如何 弯曲的组件调节信号网络。我们将联合收割机 包括高分辨率荧光、扫描电镜和高速原子力的方法 显微镜（HS-AFM），建模，蛋白质组学和分子遗传学。根据初步 数据，我们假设，曲率依赖性septin组装涉及的机制， 在几个长度尺度上。本文的工作有三个目的：（1）分析分离素 曲率传感中的膜相互作用;（2）确定Septin的生物物理性质 （3）确定弯曲的分隔蛋白组件如何招募 特异性信号蛋白从拟议的实验，我们将了解如何纳米 长度尺度机制有助于感应微米- 尺度曲率这些研究还将揭示septin支架如何作为一种细胞因子而改变。 局部曲率函数这项拟议研究的长期目标是确定如何 septins识别微米级曲率，然后使用形状信息进行调制， 细胞功能。

Summary/abstract Cell shape is integral to function and can be described in terms of plasma membrane curvature. Many changes in cell curvature occur on the micrometer scale but proteins are nanometers in size, raising the question as to how cells can perceive, control and use micrometer-scale geometry. The septins are a conserved, filament-forming family of proteins that preferentially assemble at sites of micrometer-scale membrane curvature. Septins assemble on many curved surfaces including at the cytokinetic furrow, dendritic spines, membrane blebs, around intracellular bacteria and bases of cilia and flagella. Given these diverse cell contexts, malfunction of septins is linked to diverse human diseases including many cancers, neuropathies and infertility. At sites of micrometer-scale membrane curvature, septins can influence the diffusion of proteins in the membrane, act as scaffolds to bring together signaling proteins, and impact the rigidity of the cell cortex. How curved septin assemblies form and recruit signaling proteins to the local membrane is critical to understand how septins link cell geometry to responses. Septin filament assembly occurs through annealing of short (~24-32nm) oligomeric rods on lipid bilayers or other cytoskeletal networks. We hypothesize that cells modulate the membrane affinity, length, density, and geometrical arrangement of septins in a curvature-dependent manner. The goal of this proposal is to identify the mechanisms directing assembly of septins on curved surfaces and to measure how curved assemblies regulate signaling networks. We will combine a variety of imaging approaches including high-resolution fluorescence, SEM and high-speed atomic force microscopy (HS-AFM), modeling, proteomics, and molecular genetics. Based on preliminary data, we hypothesize that curvature-dependent septin assembly involves mechanisms at work on several length scales. This work will be directed by three aims: (1) Analyze septin membrane interaction in curvature sensing; (2) Determine the biophysical properties of septin filaments that enable curvature sensing; (3) Identify how curved septin assemblies recruit specific signaling proteins. From the proposed experiments, we will learn how nanometer length scale mechanisms contribute to the emergent mesoscale process of sensing micron- scale curvature. These studies will also reveal how septin scaffolding may change as a function of local curvature. The long-term goal of this proposed study is to identify how septins recognize micrometer-scale curvature and then use shape information to modulate cellular functions.