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Title: Interrogation of the molecular mechanisms underlying the neural specification of human embryonic stem (hES) cells
Author: FitzPatrick, Lorna
ISNI:       0000 0004 6057 6626
Awarding Body: St George's, University of London
Current Institution: St George's, University of London
Date of Award: 2016
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LIN28 is an RNA-binding protein and a key regulator of developmental timing. It has dual functionality; preventing the maturation of microRNAs such as the let-7 family and increasing the translation efficiency of targeted mRNAs. LIN28 is highly expressed in human embryonic stem (hES) cells and is rapidly down regulated during neural maturation. Nuclear factor kappa B (NFkB) is a family of transcription factors most notable its role in inflammation. In 2009, it was reported that NFkB mediated cell transformation by directly upregulating UN28 transcription. However, in recent years a number of studies have demonstrated that NFkB plays an integral part in neurogenesis, when UN28 is transcriptionally silent. Activation of the canonical NFkB signalling pathway has been shown to be necessary for early neural stem cell (NSC) differentiation and inhibition of NFkB activity in vitro blocks neural maturation, although the mechanism remains elusive. The aim of this project is to clarify the roles of LIN28 and NFkB throughout neural specification of human embryonic stem (hES) cells in vitro using a reproducible targeted differentiation protocol. I have utilised lentiviral technology to modulate and monitor LIN28 and NFkB activity, respectively. Transcriptomic microarray analysis at key stages of neural differentiation indicated an inverse relationship between NFkB signalling and UN28.1 found that as neural progenitors (NPCs) differentiate, they downregulate LIN28 and become dependent on oxidative phosphorylation (OXPHOS) as a means of generating ATP. Furthermore, over extended passage, NPCs acquired high levels of NFkB activity and accumulated reactive oxygen species (ROS). Furthermore, late passage NPCs exhibited slower cell cycling, increased levels of cell cycle inhibitor p21 and critically an increased propensity for terminal differentiation. Constitutive activation of NFkB in early passage NPCs resulted in increased OXPHOS and p21 expression, suggesting that NFkB regulates metabolic activity and cell cycle progression during neural specification of hES cells.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available