Robust surface-to-mass coupling and turgor-dependent cell width determine bacterial dry-mass density

<p><span>During growth, cells must expand their cell volumes in coordination with biomass to control the level of cytoplasmic macromolecular crowding. Dry-mass density, the average ratio of dry mass to volume, is roughly constant between different nutrient conditions in bacteria, but it remains unknown whether cells maintain dry-mass density constant at the single-cell level and during non-steady conditions. Furthermore, the regulation of dry-mass density is fundamentally not understood in any organism. Using quantitative phase microscopy and a new image-analysis pipeline, we measured absolute single-cell mass and shape of the model organisms </span><i><span>Escherichia coli</span></i><span> and </span><i><span>Caulobacter crescentus</span></i><span> with improved precision and accuracy. We found that cells control dry-mass density indirectly, by expanding their surface, rather than volume, in direct proportion to biomass growth – according to a new surface growth law. At the same time, cell width is controlled independently. Therefore, cellular dry-mass density varies systematically with cell shape, both during the cell cycle or after nutrient shifts, while the surface-to-mass ratio remains nearly constant on the generation time scale. </span><span><span>Transient deviations from constancy during nutrient shifts can be reconciled with turgor-pressure variations and the resulting elastic changes in surface area.</span></span><span> Finally, we find that plastic changes of cell width after nutrient shifts are likely driven by turgor variations, demonstrating an important regulatory role of mechanical forces for width regulation. In conclusion, turgor-dependent cell width and a new, slowly varying surface-to-mass coupling constant are the independent variables that determine dry-mass density.</span></p>