Nové molekulární mechanismy účastnící se kontroly buněčného cyklu

Aquino, Cecilia

Cecilia Aquino Perez, M. Sc. Doctoral thesis abstract In this doctoral, thesis we aimed to find and study novel mechanisms regulating cell cycle phase transitions in non-stressed conditions and in context of the cell response to various types of stress. First, we focused on studying Polo-like kinase 3 that has previously been implicated in activation of the cell cycle checkpoint after DNA damage. For this, we employed CRISPR/Cas9- mediated gene editing to knock-out PLK3 in RPE cells while in parallel performing RNA interference assays and submitting the cells to different types of stress. The main observation was that in both systems PLK3 was disposable for response to DNA damage, hypoxia and osmotic stress. Through mass spectrometry analysis of purified EGFP-PLK3 we identified PP6 and its regulatory subunits PPP6R1 and PPP6R3 as novel PLK3 interactors. We observed that PLK3 is phosphorylated in its conserved residue Thr-219 and that PP6 depletion boosted PLK3 phosphorylation status but did not affect its kinase activity. The possible regulation of PLK3 trough PP6 is interesting and its biological relevance will be addressed by future research. Next, we performed a transcriptomic analysis in human RPE-FUCCI cells aiming to identify new regulators of the cell cycle. We selected Family with sequence...

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