Cells dynamically adapt their nuclear volumes and proliferation rates during single to multicellular transitions

Tumour development and progression are associated with biophysical alterations that manifest across multiple spatial scales, from the subcellular to multicellular tissue scale. While cells dynamically regulate their biophysical properties like volumes and mechanics in dependence of cell state and function, it is unclear how these properties are controlled in the dense multicellular environment of a developing tumour. Here, we quantified cell and nuclear volumes of single cancer cells, while they grew into multicellular tumour spheroids within well-defined, tuneable biohybrid polymer hydrogels. We quantitatively showed that the formation of multicellular structures is associated with marked reductions of cellular and nuclear volumes, cell cycle delays as well as cell mechanical alterations, and that these changes are coupled. Single-to-multicellular transitions coincided with a drastic decrease in median nuclear volumes by up to 60%, as well as overall cell volume decrease. Nuclear volume decrease could not be explained by compression due to confining microenvironments. Instead, cell cycle adaptions were one significant contributor, with smaller-sized G1 cells accumulating in growing clusters, an effect that was reversed by CDK1 inhibition. Another contributor was nuclear volume decrease in cells within clusters that was associated with higher mass density and stiffness and could be abrogated upon cell release from clusters. In turn, multicellular-to-single cell transitions that happened in cells that invaded from a tumour spheroid into the surrounding matrix, were accompanied by nuclear volume increases and cell softening. Taken together, our study provides insights into how cells dynamically adapt their cellular/nuclear volumes, cell cycle progression and mechanics in dependence of the multicellular state.