Date with Destiny: Genetic and Epigenetic Factors in Cell Fate Decisions in Populations of Multipotent Stem Cells
University of Cambridge
Doctor of Philosophy (PhD)
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Edri, S. (2019). Date with Destiny: Genetic and Epigenetic Factors in Cell Fate Decisions in Populations of Multipotent Stem Cells (Doctoral thesis). https://doi.org/10.17863/CAM.35678
The governance of cell fate decisions during development is a fundamental biological problem. An important aspect of this is how cells exit a multipotent state and choose their fates in a correct manner and proportion. To tackle an aspect of this problem, I have focused on 2 multipotent models: one infinite self-renewal pluripotency in an artificial environment, and the other, bipotent progenitors in the context of the mouse embryo. The first model aimed to explore the effects of chromatin-associated factors on the ability of pluripotent mouse Embryonic Stem Cells (ESCs) to self-renew, via monitoring gene expression heterogeneity of key genes. The second model focused on Neural Mesodermal Progenitors (NMPs), a bipotent cell population found in the Caudal Lateral Epiblast (CLE) of mammalian embryos, which contributes to the spinal cord and paraxial mesoderm. The aim here was to derive NMPs in vitro which exhibit similar gene expression patterns and function like their mouse embryo counterpart and study their renewal and differentiation in detail. The first multipotent model explores the effects of chromatin remodelling on cell fate decisions, specifically investigating the consequences of inhibiting the histone acetyltransferase Kat2a on the ESCs fate. I found first, that the effect of Kat2a inhibition depends on the pluripotent state of the cells; cells in a ground state exhibit a resistance to Kat2a inhibition and maintain their pluripotency, whereas cells in a naïve state experience destabilization of their pluripotency gene regulatory network and shift towards differentiation. Second, that Kat2a inhibition in the naïve state results in a decline in the gene expression noise strength contributed by the promoter activation operation, which suggests that when ESCs become lineage-primed their transcriptional noise is constrained. In the bipotent model, the NMPs are identified as cells coexpressing Sox2 and T/Brachyury, a criterion used to derive NMP-like cells from ESCs in vitro. Comparison between the different NMPs protocols stresses that Epiblast Stem Cells (EpiSCs) are an effective source for deriving a multipotent population resembling the embryo Caudal Epiblast (CE), that generates NMPs. Furthermore, self-organization of this CE-like population, resulted in axially organized aggregates. Exploiting the mouse embryo CLE as a reference shows that EpiSCs derived NMPs, monolayers and aggregates, consist of a high proportion of cells with the embryo’s NMP signature. Importantly, studying this system in vitro sheds light on the sequence of events which lead to NMP emergence in vivo. On this basis, I conclude that understanding the initial state of cells at a crossroads is important to reveal the limitations it imposes on the cells fate exploration, hence makes it possible to mimic more precisely the fate decision process in vitro.
Pluripotency, In vitro culture, Neural Mesodermal Progenitors, Epiblast Stem Cells, mouse Embryonic Stem Cells
This work was supported by Cambridge Trust, Cambridge Philosophical Society, AJA Karten trust and Fitzwilliam college.
This record's DOI: https://doi.org/10.17863/CAM.35678
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