Dynamics of Escherichia coli ’s passive response to a sudden decrease in external osmolarity

Abstract

For most cells, a sudden decrease in external osmolarity results in fast water influx that can burst the cell. To survive, cells rely on the passive response of mechanosensitive channels, which open under increased membrane tension and allow the release of cytoplasmic solutes and water. Although the gating and the molecular structure of mechanosensitive channels found in Escherichia coli have been extensively studied, the overall dynamics of the whole cellular response remain poorly understood. Here, we characterize E. coli's passive response to a sudden hypoosmotic shock (downshock) on a single-cell level. We show that initial fast volume expansion is followed by a slow volume recovery that can end below the initial value. Similar response patterns were observed at downshocks of a wide range of magnitudes. Although wild-type cells adapted to osmotic downshocks and resumed growing, cells of a double-mutant (ΔmscL, ΔmscS) strain expanded, but failed to fully recover, often lysing or not resuming growth at high osmotic downshocks. We propose a theoretical model to explain our observations by simulating mechanosensitive channels opening, and subsequent solute efflux and water flux. The model illustrates how solute efflux, driven by mechanical pressure and solute chemical potential, competes with water influx to reduce cellular osmotic pressure and allow volume recovery. Our work highlights the vital role of mechanosensation in bacterial survival.osmotic downshock | bacterial mechanosensing | single-cell imaging B iology offers an array of intriguing mechanical solutions, both active and passive, often exceeding what is currently possible with man-made methods. Understanding how biological systems achieve different functionalities under mechanical stimuli can inform new, thus-far-unexplored design principles. One such passive control system is the bacterial response to sudden decreases in external osmolarities.A Gram-negative cell's fluid cytoplasm is separated from the external environment by the inner membrane, the periplasmic space, and the outer membrane. Ordinarily, the total solute concentration within the cytoplasm is higher than that of the environment, resulting in a positive osmotic pressure on the cell wall (termed turgor pressure) (1). Escherichia coli is able to respond to both increases and decreases in external concentrations. An increase in external osmolarity (hyperosmotic shock or upshock) results in water efflux from the cell interior, causing cellular volume to shrink and osmotic pressure to drop to zero (2). E. coli responds by actively accumulating specific solutes (osmolytes), such as potassium, proline, and glycine-betaine (2). Accumulation of osmolytes in the cell's cytoplasm causes reentry of water, cell volume increase, and recovery of osmotic pressure (3, 4). A downward shift in external osmolarity (termed hypoosmotic shock or downshock) causes fast water influx into the cell's cytoplasm. As a result, the osmotic pressure increases and the cell expands in a nonlinear fashion (5,6

Osmotaxis in Escherichia coli through changes in motor speed

“…At t = 5min, the extracellular osmolarity is elevated by delivering 400 mM sucrose, at a local flow rate of 0.68 µl/min [Buda et al, 2016], causing an osmotic upshock of 488 mOsmol/kg. Immediately upon upshock, the motor stops switching rotational direction and speed drops.…”

“…Osmotic shocks were performed while the slide was in the microscope by adding 24 µl of the shocking solution to one end and immediately bringing a piece of tissue paper to the other, resulting in flow and exchange of media. The flow duration was approximately 10-15 s and the local flow rate that the attached bacterium experienced was 0.68 µl/min [Buda et al, 2016]. Tunnel was sealed after the shock to prevent evaporation and thus potential further increase in osmolarity throughout the course of the experiment.…”

Cooperative response and clustering: Consequences of membrane-mediated interactions among mechanosensitive channels

“…3 shows that a cluster could take a time of the order of 100 ms to open. Another recent work [36] also highlights the large cell-to-cell variability of the downshock responses. Furthermore this work shows a very slow cell volume recovery, which may also indicate channels cooperative activity.…”

Dynamic clustering regulates activity of mechanosensitive membrane channels

“…To prevent it, a portfolio of MSC in E. coli's inner membrane act as "pressure release valves" that open * Electronic address: a.saric@ucl.ac.uk and create a nano-sized pore at the centre of the protein. This in turn enables solute and water efflux, reestablishing desired osmotic pressure inside the cell [10][11][12]. This response is fast and solely regulated by the membrane tension and chemical potential of water and solutes [12].…”

“…This in turn enables solute and water efflux, reestablishing desired osmotic pressure inside the cell [10][11][12]. This response is fast and solely regulated by the membrane tension and chemical potential of water and solutes [12].…”