Stochastic aspects of chromatin and nuclear envelope dynamics in mouse embryonic stem cells (Doctoral thesis)
Differentiation of embryonic stem cells is associated with progressive immobilization of chromatin proteins, increase in the relative proportions of heterochromatin and decline of global transcriptional activity. Although these events signify the gradual transition from “open” to “closed” chromatin states, hundreds of genes are de-repressed or activated during lineage commitment, suggesting differential remodeling of the chromatin landscape. Using a FRAP/FLIP-based approach in combination to live cell imaging, we have shown here that the mobility of chromatin proteins is not bijectively related to exit from pluripotency or activation of lineage-specific genes at the level of single cells. Comparison of dynamic ensembles further reveals the occurrence of random spatio-temporal fluctuations, which continuously scramble the chromatin landscape and create a “shifting mosaic steady-state” across and within heterochromatic domains. Apparently, less dynamic chromatin forms become increasingly more stable and more abundant as cells transit from one attractor state to the next, allowing subtle –yet, fate-determining- changes in gene expression mode, while maintaining a significant degree of developmental plasticity.New evidence implicates the nuclear envelope in the control of gene expression and the orderly progress of differentiation. Elaborating on this theme, we have overexpressed and knocked-out the polytopic membrane protein lamin B receptor (LBR) in mouse embryonic stem cells and studied the functional consequences at the cellular and the molecular level. The results show that increased LBR dosage limits the dynamic range of Nanog fluctuations and shifts naïve ESCs towards a primed metastate. This shift is paralleled by dramatic changes in expression profile and a mesendodermal differentiation impairment. Although the inappropriate expression of LBR does not lead to exit from the undifferentiated state, increased LBR dosage causes defects in embryoid body formation and inappropriate expression of lineage-specific markers upon induction of differentiation, suggesting involvement of this protein in critical cell fate decisions.
|Institution and School/Department of submitter:||Πανεπιστήμιο Ιωαννίνων. Σχολή Επιστημών Υγείας. Τμήμα Ιατρικής|
|Subject classification:||Embryonic stem cells|
|Keywords:||Εμβρυονικά βλαστικά κύτταρα,Χρωματίνη,Πυρηνικός φάκελος,Διαφοροποίηση|
|Appears in Collections:||Διδακτορικές Διατριβές|
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