Intrinsic clustering of flagellar basal body proteins in E. coli: A self-organization mechanism for assembly and regulation

Abstract

The assembly and spatial organization of flagellar basal bodies in Escherichia coli are crucial for motility and chemotaxis. Using fluorescence and single-molecule microscopy, we demonstrate that key basal body proteins, FliF and FlhA, self-organize into clusters from low to high expression conditions. Rather than forming new basal bodies, excess proteins accumulate around pre-existing structures, suggesting an autocatalytic mechanism. It is confirmed that clustering occurs even at low protein levels, indicating an intrinsic organizational principle rather than an artifact of overexpression. Fluorescence recovery after photobleaching (FRAP) revealed dynamic protein exchange within clusters, supporting a diffusion-capture model. Single-molecule analysis showed that FlhA actively remodels clusters, while FliF stabilizes them. 3D imaging suggested that basal body positioning optimizes flagellar distribution for efficient motility. These findings highlight a robust mechanism that regulates basal body positioning and flagellar assembly, ensuring adaptability to varying cellular conditions.

Keywords

• Flagellar assembly
• Protein self-organization
• Spatial regulation
• Fluorescence super-resolution imaging
• Single-molecule methods