Cells depend on a highly ordered organisation of their content and must develop strategies to maintain the anisotropic distribution of organelles during periods of nutrient shortage. One of these strategies is to solidify the cytoplasm, which was observed in bacteria and yeast cells with acutely interrupted energy production. Here, we describe a different type of cytoplasm solidification fission yeast cells switch to, after having run out of nutrients during multiple days in culture. It provides the most profound reversible cytoplasmic solidification of yeast cells described to date. Our data exclude the previously proposed mechanisms for cytoplasm solidification in yeasts and suggest a mechanism that immobilises cellular components in a size-dependent manner. We provide experimental evidence that, in addition to time, cells use intrinsic nutrients and energy sources to reach this state. Such cytoplasmic solidification may provide a robust means to protect cellular architecture in dormant cells.

Using correlative light and electron microscopy (CLEM), we studied the intracellular organization by of glucose-starved fission yeast cells (Schizosaccharomyces pombe) with regards to the localization of septin proteins throughout the cytoplasm. Thereby, we found that for cells carrying a deletion of the gene encoding septin-2 (spn2Δ), starvation causes a GFP-tagged version of septin-3 (spn3-GFP) and family members, to assemble into a single, prominent filamentous structure. It was previously shown that during exponential growth, spn2Δ cells form septin-3 polymers. However, the polymers we observed during exponential growth are different from the spn3p-GFP structure we observed in starved cells. Using CLEM, in combination with anti-GFP immunolabeling on plastic-sections, we could assign spn3p-GFP to the filaments we have found in EM pictures. Besides septin-3, these filamentous assemblies most likely also contain septin-1 as an RFP-tagged version of this protein forms a very similar structure in starved spn2Δ cells. Our data correlate phase-contrast and fluorescence microscopy with electron micrographs of plastic-embedded cells, and further on with detailed views of tomographic 3D reconstructions. Cryo-electron microscopy of spn2Δ cells in vitrified sections revealed a very distinct overall morphology of the spn3p-GFP assembly. The fine-structured, regular density pattern suggests the presence of assembled septin-3 filaments that are clearly different from F-actin bundles. Furthermore, we found that starvation causes substantial mitochondria fission, together with massive decoration of their outer membrane by ribosomes.

Cells depend on a highly ordered organization of their content and they must develop strategies to maintain the anisotropic distribution of organelles during periods of nutrient shortage. One of these strategies, observed in bacteria and in yeast cells with acutely interrupted energy production, is to solidify the cytoplasm. Here, we describe a different type of cytoplasm solidification that occurs in fission yeast cells having slowly run out of nutrients after multiple days of culturing. It provides the most profound reversible cytoplasmic solidification of yeast cells described to date. Our data suggest the involvement of a matrix with a certain mesh size that immobilizes cellular components in a size-dependent manner. We provide experimental evidence that cells need time, intrinsic nutrients and intrinsic energy sources to enter this state in the absence of external sources. Such cytoplasmic solidification may provide a robust means to protect cellular architecture in dormant cells.