Breaking out of silence- epigenetics of cell cycle re-entry during fission yeast spore germination

Quiescent cells are all around us. We know surprisingly little about the behavior of cells during quiescence, and the epigenetic requirements of their state when they return to active cycling.

To look into these questions, we study chromatin structure and epigenetic requirements during spore germination in fission yeast Schizosaccharomyces pombe. Spores are dormant G0/G1 cells, and their germinaton serves as a model for returning to active cycling.

We have compared germination dynamics of quiescent spores from wild-type strains and mutants in various chromatin proteins. Spores deleted for the SuVar39h homolog Clr4, a H3K9 methyltransferase, germinate at a much slower rate than wild-type spores. This effect is specific to germination rather than mitotic growth, and does not originate from a meiotic defect. Single cell live imaging revealed that part of the spores does not enter the germination process, while another part dies immediately upon cell cycle entry. The rest of the spores delay in germination compared to wild-type dynamics. We set out to examine what is the role of Clr4 during spore germination. For this purpose, we performed ChIP of methylated H3K9 in wild-type spores during germination. We observe a rise in H3K9me signal on peri-centromeric regions during the process, suggesting that heterochromatin in this region is modified during germination, and that the absence of this modification in clr4 mutants could have caused the germination defect.

The same genetic requirements are true also for the RNAi Dicer gene, Dcr1. Upon deletion, delay in germination is observed, as well as additional phenotypes seen for clr4. Double mutants appear to be severely defective in germination.

We further show the importance of these pathways by showing that wild-type spores barely age in the lab, while 3 months old clr4 spores are unable to germinate. This experiment successfully models Werner`s syndrome phenotypes through spore germination.