Study towards understanding of oscillatory mechanisms of cell-fate change

Prof. Dr. Reet Kurg

The process how cancer cells can escape lethality after cancer therapy and cause cancer relapse is not well understood. Treated cancer cells obtain many damages in the DNA that the system and its packing (i.e. the chromatin in the cell nucleus) loses its regulatory stability. However, since the human cells have evolved through the long evolution of Life, survival programs are saved in the genome memory. By activating the reprogramming capacity coupled to cellular senescence, cancer cells can explore and find these hidden evolutionary attractors to escape their situation. In this way the system reaches a bifurcation point with the possibilities of two cell fates, the senescence arresting a non-proliferating state, or stemness reprogramming cell development into a proliferating, clonogenic, survival state. The final outcome of this process is a stochastic event depending on co-factors to which a cell might be sensitive. In previous investigations by the Baltic groups (Tartu group in collaboration with Dr. Erenpreisa, Riga) using a cell model of embryonal carcinoma treated with etoposide, it was shown that before this exit, the cells fluctuate during several days between senescence and self-renewal. These fluctuations were associated with collaborative p53-dependent up- and down-regulation of two opposing cell fate regulators p21Cip1 (senescence/death) and OCT4A (stemness survival), reciprocally moderating each other (Huna et al., 2015). This bi-potential state provides additional time for the DNA repair while the cells switch between (pre-) senescence and self-renewal, and reverses to the initial definite state at recovery. In a previous work on the same model, it was found that the bi-potential state is characterized by DNA de-methylation in the heterochromatin and methylation in euchromatin, likely erasing phase separation between them (Salmina et al., 2017). The epigenetic histone state in hetero-and euchromatin in this model was not studied and is one of the tasks of the current project.

Using single molecule localization microscopy (SMLM) and mathematical evaluation methods (Ripley point distance statistics, Persistent Homology, Persistent Imaging, Principal Component Analysis) for point pattern analysis of specific fluorescence labelling, hetero- or euchromatin revealed characteristic network-like changes of the whole chromatin in cell nuclei after radiation treatment during DNA repair (Hausmann group). It was shown that successful DNA repair results in a loop in the 2D- topological space of two major components for the chromatin network, reaching approximately the state of untreated cells. Remaining damages or un-successful repair reasons topological fluctuations or in the worst cases escapes from this loop in the two major components spaces, i.e. the loop is not closed towards the beginning situation. To which extent characteristics exist in the chromatin network-pattern that are related to either (pre-)senescence or pluripotency and thus to the processes described above are questions of research.

The collaborative project brings together the expertise of both groups the Estonian and German one. The project will be strongly supported by consultations of Dr. Erenpreisa (Emeritus Scientist) and Dr. T. Freivalds in physics (Latvian University). The aim is to drive cells of  the model cell system into the fluctuating state after treatment with etoposide, defining the state of the cells before, during fluctuations and in the final outcome, and correlating these states with chromatin organization parameterized by image analysis of phase separation using confocal microscopy and topological data calculations after  SMLM data acquisition.

Direct: senior scientists, PhD students, master students, bachelor students involved in the research and measurements

Indirect: scientists informed by publication of results, students and participants of the symposium