PhD Dissertation Defense: Jiawei Sun

Title: Physical structures and properties of the Gram-negative cell envelope
Abstract: The bacterial cell envelope is essential for maintaining cell integrity while supporting growth and division. In Gram-negative bacteria, it consists of three distinct layers: the cytoplasmic membrane (CM), a rigid peptidoglycan cell wall (CW), and an asymmetric outer membrane (OM). Although the cell relies on the envelope for viability, shape maintenance, and mechanical integrity, the physical structures and properties underlying these functions remain largely understudied, limiting the development of strategies to destabilize the bacterial envelope. This thesis employs a biophysical approach to investigate two important envelope structures, revealing a mechanical role for MreB filaments as potential linkers between the CM and the CW, and identifying molecular interactions that control OM fluidity.
The bacterial actin homolog MreB is crucial for cell shape determination in many rod-shaped bacteria. Using microfluidics to apply oscillatory hyperosmotic shocks, we found that MreB mechanically couples the CM to the CW, thereby combining the two layers for improved load-bearing capacity. The MreB-mediated linkage, especially at the cell poles, is sufficient to resist plasmolysis, indicating that MreB is the major mechanical connection between the two layers.
The OM is a versatile layer that protects the cell with its selective permeability and mechanical stiffness. Unlike most lipid bilayers such as the fluid CM, the OM exhibits extremely limited lateral diffusion, resulting in spatial heterogeneity. Using fluorescence recovery after photobleaching assay alongside genetic and chemical perturbations, we identified key molecular interactions affecting OM fluidity, including increased phospholipids in the outer leaflet that fluidize the OM. Increased OM fluidity was found to be associated with shorter cells through altered divisome dynamics, linking envelope properties to the regulation of bacterial cell body plan.
Please contact Madelyn Bernstein for the Zoom link