Precise regulation of nuclear size relative to cell size is a robust feature of cellular biology, yet mechanisms governing nuclear size control remain poorly understood. We recently proposed a physical model in which the nuclear-to-cell volume ratio (N/C ratio) at steady state is set by a balance between colloid osmotic pressures generated by numbers of osmotically active macromolecules within the nucleus and cytoplasm, and by nuclear envelope tension. In fission yeast, with minimal envelope tension, nuclear size acts as an osmometer. As a key test of this model, we increased osmotic pressure in a compartment-specific manner by massively expressing exogenous proteins targeted to either the nucleus or cytoplasm. Loading the nucleus with millions of copies of mCherry-GST-NLS proteins caused a striking, dose-dependent expansion of nuclear volume, elevating the N/C ratio up to 4-fold. Conversely, expression of mCherry-GST-NES proteins in the cytoplasm reduced the N/C ratio in a dose-dependent fashion. As a control, equivalent high expression of mCherry in both compartments did not affect growth or nuclear size. These data, which closely fit model predictions, provide quantitative support for this osmotic model for nuclear size and yield fundamental parameters for osmotic theory in living cells. Additionally, these perturbations altered the physical properties of the nucleoplasm. Expression of mCherry-GST-NLS in the nucleus significantly increased diffusion of 40 nm-GEM nanoparticles, consistent with nuclear expansion and dilution of endogenous nuclear components. Conversely, mCherry-GST-NES expression in the cytoplasm reduced nuclear diffusivity. To test whether macromolecular crowding affects nuclear condensate assembly, we observed consistent effects on Swi6/HP1 heterochromatin loci and the nucleolus. Together, this work establishes a tunable experimental strategy to manipulate the N/C ratio and physical properties of the nucleoplasm, providing a powerful tool to study how osmotic forces influence nuclear size control, molecular crowding, and nuclear organization.
Lemière et al. (Sun,) studied this question.