In bacteria, a host of enzymes regulates the reproducible
and robust construction of the cell wall, whose mechanical integrity is crucial
for viability under osmotic stress.
Antibiotics that target these enzymes disrupt cell wall construction,
ultimately leading to mechanical failure of the cell. Our work explores the
physical mechanisms of cell growth and death, as a guide to understanding
antibiotic mechanisms that disrupt mechanical properties of the cell. We use a combination of cell wall
fluorescent labeling, high resolution time-lapse microscopy, and computational
image processing to characterize where, and with what dynamics, cell wall and
outer membrane growth occurs. Analysis
of cell-surface marker fluorescence indicates that the cytoskeleton is present
at sites of active growth and that chemical depolymerization of the
cytoskeleton causes homogeneous, unstructured growth and eventual cell death by
rupture. When combined with cell-shape
analysis, our data strongly suggest that dynamic localization of the bacterial
cytoskeleton is part of a curvature sensing and growth feedback mechanism that
orchestrates heterogeneous growth to maintain cell shape and regulate
mechanical stress. Using
surface-specific protein labeling, we show that outer membrane growth also
occurs in a similar heterogeneous manner.
Quantitative tracking of growth is an effective method for
characterizing cell wall mechanical failure resulting from antibiotic
treatment. These techniques pave the way
for studying the detailed dynamics of growth-associated proteins and their
disturbance by antibiotics.
NOTE: IT'S ON WEDNESDAY NOT THURSDAY!