Figure 3: Probing the magnitude and impact of OM stiffness.
A) In a microfluidic flow cell, exponentially growing E. coli cells are under mechanical stress due to turgor pressure, which is born predominantly by the PG rather than the OM. Cells shrink suddenly when exposed to a hyperosmotic shock due to water efflux, decreasing the overall stress on the cell envelope. In this shrunken state, the cell wall remains partially extended as the OM experiences compression. Removal of the OM by detergent or EDTA treatment allows the cell wall to fully relax to its rest length.
B) Growth of E. coli cells initially embedded within a narrow channel leads to exposure of part of the cell body to fluid flow, whose hydrodynamic force can be tuned. EDTA-treated cells deflect more than untreated cells for a given flow strength40, highlighting the strength of the OM.
C) AFM measurements directly confirm the loss of cell stiffness by EDTA treatment. EDTA-treated cells indented more than untreated cells for the same amount of applied force; several outer membrane mutants also exhibited more indentation40.
D) In addition to EDTA treatment, OM stiffness can be compromised by deletion of various OM proteins, intercalation of the OM-specific dye FM4-64, or LPS modification such as deletion of the O-antigen.
E) E. coli cells adopt a wall-less L-form or wall-deficient spheroplast state when exposed to phage or cell wall-targeting antibiotics, in which the OM bears stress to avoid envelope rupture.
F) Cells with compromised OM stiffness are more susceptible to death during osmotic-shock oscillations.