Mechanisms of assembly and function of the septin cytoskeleton in Drosophila

果蝇隔膜细胞骨架的组装机制和功能

• 批准号: RGPIN-2021-03324
• 负责人: Hickson, Gilles
• 金额: $ 2.33万
• 依托单位: Université de Montréal
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=761938
• 关键词: Mechanisms; assembly; function; septin; cytoskeleton

Mechanisms of assembly and function of the Drosophila septin cytoskeleton. Background and Preliminary Data Septins form a family of highly conserved, cytoskeletal proteins that act at the interface of membranes and the actin- and microtubule-based cytoskeletons. Septins form palindromic, hetero-oligomeric rods, or "complexes", that can polymerize end-on-end into filaments. These filaments can assemble into higher-order structures such as bundled filaments, rings, gauzes and as we have found, tubes. Septins can act as scaffolds and as membrane diffusion barriers in diverse cellular structures such as the base of the primary cilium, the sperm annulus, and the cleavage furrow during cell division. However, it is not clear how higher order assemblies form, are regulated or how they impact cellular homeostasis. Septins share a conserved organization including a central GTPase domain that is essential for complex formation and more variable N- and C-termini that contain polybasic regions (N-termini) and coiled-coil domains (C-termini) whose roles are poorly understood. We have identified three prominent structures comprising higher-order assemblies of septins in cultured Drosophila Schneider's S2 cells: cytoplasmic rings, cytoplasmic tubes and the plasma membrane-anchored cytokinetic ring. Preliminary experiments show that these structures have differential requirements for their formation. Hypothesis, Aims and Approach Our long-term aim is to define the molecular basis of the full complement of septin higher-order assemblies and to understand their specific functions in both cellular and organismal contexts. We hypothesize that specific interaction partners collaborate with specific septin domains to produce different higher-order assemblies and that these differentially regulate cellular architecture and function. Using a combination of biochemical and cell biological approaches we will perform a comprehensive analysis of Drosophila septins to define the molecular determinants of each type of higher-order structure. We propose the following specific aims: 1) Define the requirements for the assembly of septin-based structures in Drosophila S2 cells. 2) Define septin assembly mechanisms and interactions with actin in vitro. 3) Define the roles of the septin cytoskeleton in vivo in Drosophila. These studies will elucidate the molecular mechanisms by which septins can assemble into different higher-ordered structures in living cells, and will shed light on the undoubtedly complex interplay between septins, other cytoskeletal elements and the plasma membrane. This will greatly advance our understanding of the molecular events underlying the multitude of cellular processes in which the septins participate.

果蝇隔蛋白细胞骨架的组装和功能机制。Septins是一个高度保守的细胞骨架蛋白家族，作用于细胞膜与肌动蛋白和微管骨架的界面。隔蛋白形成回文的异源寡聚棒或“复合物”，其可以端对端地连接成细丝。这些纤维可以组装成更高级的结构，如成束的纤维、环、纱布和我们已经发现的管。隔膜蛋白可以作为支架和作为膜扩散屏障在不同的细胞结构，如基础的初级纤毛，精子环，和分裂沟在细胞分裂。然而，目前尚不清楚更高级的组装体如何形成，如何调节或它们如何影响细胞内稳态。Septins共享一个保守的组织，包括一个中心的GTdR结构域，这是必不可少的复杂的形成和更可变的N-和C-末端，含有多碱基区域（N-末端）和卷曲螺旋结构域（C-末端），其作用知之甚少。我们已经确定了三个突出的结构，包括在培养的果蝇Schneider的S2细胞中的septins的高阶组件：细胞质环，细胞质管和质膜锚定的细胞动力学环。初步实验表明，这些结构的形成有不同的要求。假设，目标和方法我们的长期目标是定义septin高阶组装的完整补充的分子基础，并了解它们在细胞和生物体环境中的特定功能。我们假设特定的相互作用伙伴与特定的septin域合作，产生不同的高阶组件，这些差异调节细胞的结构和功能。使用生物化学和细胞生物学的方法相结合，我们将进行一个全面的分析果蝇septins，以确定每种类型的高阶结构的分子决定因素。我们提出了以下具体目标：1）定义果蝇S2细胞中基于septin的结构组装的要求。 2)定义septin组装机制和与肌动蛋白在体外的相互作用。3)定义果蝇细胞骨架中septin的作用。这些研究将阐明septins在活细胞中组装成不同的高级结构的分子机制，并将阐明septins，其他细胞骨架元素和质膜之间无疑复杂的相互作用。这将极大地推进我们对septins参与的众多细胞过程的分子事件的理解。