Invited Talk

Deformation of metabolically intact isolated nuclei

Fitzroy J. Byfield¹, Alison E. Patteson², Paul A. Janmey¹

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The cell nucleus is generally considered to be the stiffest organelle in the cell, with a Young’s modulus on the order of 1-10 kPa. But cell motility through 3D matrices with elastic moduli less than 1 kPa deform the nucleus to large strains capable of breaking the nuclear lamina, and lipid droplets in a hepatocyte exerting 100 Pa stresses on them also deform the nucleus. Metabolically active nuclei were produced from live cells by a centrifugation process that enucleates the cell. This process leaves behind a cytoplast and produces a nucleus that is wrapped by a plasma membrane and a thin layer of cytosol (a karyoplast), but no discernible cytoskeleton, endoplasmic reticulum, ribosomes or other large organelles1,2. Enucleation is more efficient from vimentin null fibroblasts compared to normal fibroblasts with a perinuclear vimentin cage, but the isolated nuclei are indistinguishable. Force-indentation curves are consistent with an elastic object with an apparent Young's modulus of 5 to 8 kilopascal, but retraction curves show that most of the work of compression is dissipated. Disrupting ATP production with glycolysis inhibitors stiffness the nucleus and eliminates dissipation. These results suggest that in metabolically intact nuclei, deformation is largely an active process that can be triggered by relatively modest forces that depend on internal and external mechanical stresses. Nuclear stiffness depends strongly on the elastic modulus of the substrate on which the cells are cultured, and nuclei are sensitive to changes in osmotic pressure of the surrounding medium, softening when they swell and stiffening when compressed. This method of isolating metabolically active karyoplasts can be done within 20 minutes and enables a range of studies of how cell perturbations, such as loss of intermediate filaments or linkers of the nuclear membrane to the cytoskeleton, impact the mechanical properties of the nucleus.