Electronic Thesis and Dissertation Repository

Degree

Doctor of Philosophy

Program

Physiology and Pharmacology

Collaborative Specialization

Developmental Biology

Supervisor

Betts, Dean H.

2nd Supervisor

Watson, Andrew J.

Co-Supervisor

Abstract

The earliest cell fate specification events during mammalian development occur in the blastocyst-stage preimplantation embryo, during which a pluripotent cell population is established. These cells form the basis of the developing fetus and must be correctly specified in order for successful development to occur. Cell signalling in response to environmental cues has a critical role in cell differentiation. The signalling adaptor protein p66Shc is expressed in mammalian embryos and promotes apoptosis and permanent embryo arrest in response to stress-inducing conditions. However, loss-of-function studies suggest that p66Shc may be important for embryonic development to the blastocyst stage. In this thesis, I aimed to determine the role of p66Shc in mouse blastocyst development and mouse embryonic stem cell function. Through a combination of environmental modulation of p66Shc expression, experimental knockdown, and genetic knockout of p66Shc in mouse preimplantation embryos and mouse embryonic stem cells, I demonstrated that p66Shc is required for normal embryo physiology, and correct cell lineage-associated marker expression in the blastocyst inner cell mass and mouse embryonic stem cells. First, I observed that p66Shc is normally upregulated at the blastocyst stage in vivo, and oxygen-induced increases in p66Shc expression are associated with altered embryo metabolism in vitro. Secondly, I demonstrated that knockdown of p66Shc transcript abundance significantly alters the timing and proportion of cells expressing lineage-associated transcription factors in the blastocyst inner cell mass. Lastly, I observed that knockout of p66Shc in mouse embryonic stem cells alters the expression of the core pluripotency marker NANOG and causes an upregulation of mesoderm-associated markers during stem cell differentiation. Collectively, my work provides insight into a novel role for p66Shc during preimplantation embryo development, expanding the diversity of cellular functions attributed to p66Shc in mammalian development.

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