Unravelling the cytoskeletal network that governs mobility and positioning of magnetic organelles in bacteria: novel players and functions.

趋磁细菌合成独特的细胞器（磁小体），这些细胞器由膜结合的磁铁矿晶体组成，它们将它们连接起来，以对抗它们主动聚集成规则链的固有物理趋势。排列的磁小体的总磁矩与明确定义的亚细胞位置相结合，最有效地作为地球磁场传感器。趋磁细菌中的磁小体形成、链组装和定位是遗传控制的，以实现在原核细胞中发现的最高结构水平之一。MTB如何将磁小体组装成规则的链，以及这种组装如何定位、分裂和均分到子细胞是MTB细胞生物学的主要有趣问题。我们在前期工作中发现了一种新的参与磁小体组装的细胞骨架结构--肌动蛋白样磁小体丝，这为深入理解这一过程提供了一把钥匙。然而，在前一个资助期获得的最新结果表明，磁小体丝可能只是一个更复杂的细胞骨架网络的一个组成部分。在这个后续的建议，我们计划扩大我们的深入了解磁细菌细胞生物学和分子机制的基础上形成和精确定位高度有序的磁小体链在完善的模型Magnetocellum gryphiswaldense。通过遗传学和生物化学的方法，我们将分析已知的和新的磁小体相关的链组装在体内和体外的关键球员的相互作用。此外，我们将详细分析磁小体迁移的机制及其与细胞周期的协调，例如蛋白质相互作用测定，以揭示与保守的细胞骨架和细胞分裂蛋白的联系。这些蛋白质在不同突变背景中的动态定位将在细胞周期期间通过先进的荧光显微镜包括超分辨率技术进行跟踪。此外，细胞骨架蛋白形成的结构将通过电子显微镜进行分析，包括先进的电子冷冻断层扫描。这些研究将通过体外分析进行补充，旨在了解细胞骨架蛋白在分子水平上的相互作用。我们的研究将揭示细菌细胞骨架结构的作用和进化以及原核细胞器的组织，分离和亚细胞定位的基本见解。

Magnetotactic bacteria synthesize unique organelles (magnetosomes) that consist of membrane-bounded magnetite crystals, and they concatenate them against their immanent physical tendency to agglomerate actively into regular chains. The summarized magnetic moments of aligned magnetosomes combined with a well defined subcellular position function most efficiently as an earth magnetic field sensor. Magnetosome formation, chain assembly and positioning in magnetotactic bacteria are genetically controlled to accomplish one of the highest structural levels found in prokaryotic cells. How MTB assemble their magnetosomes into regular chains and how this assemblage becomes positioned, split and equipartitioned to daughter cells are major intriguing questions of MTB cell biology. The discovery of a novel cytoskeletal structure involved in magnetosome assembly, the actin-like magnetosome filament, by our previous work provided a key for fundamental understanding of this process. However, most recent results obtained in the previous funding period suggest that the magnetosome filament may represent just one component of a much more complex cytoskeletal network. In this follow-up proposal, we plan to extend our in-depth understanding of magnetobacterial cell biology and the molecular mechanisms underlying formation and precise localization of highly ordered magnetosome chains in the well-established model Magnetospirillum gryphiswaldense. By genetic and biochemical approaches, we will analyze the interplay of known and new magnetosome-related key players for chain assembly in vivo and in vitro. In addition, we will analyze the mechanism of magnetosome mobility and its coordination with the cell cycle in detail by, e.g. protein interaction assays to reveal connections to conserved cytoskeletal and cell division proteins. The dynamic localization of these proteins in different mutant backgrounds will be followed during cell cycle by advanced fluorescence microscopy including superresolution techniques. In addition, structures formed by cytoskeletal proteins will be analyzed by electron microscopy including advanced electron cryo-tomography. These studies will be complemented by in vitro analyses which aim to understand the interactions of cytoskeletal proteins on molecular level. Our studies will reveal fundamental insights into the roles and evolution of cytoskeletal structures in bacteria and the organization, segregation and subcellular positioning of prokaryotic organelles.