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Src Family Tyrosine Kinase Signaling in Mouse and Human Embryonic Stem Cells

Zhang, Xiong (2013) Src Family Tyrosine Kinase Signaling in Mouse and Human Embryonic Stem Cells. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Embryonic stem (ES) cells are derived from the inner cell mass of the blastocyst stage embryo and are characterized by self-renewal and pluripotency. Previous work has implicated the Src family of protein-tyrosine kinases (SFKs) in the self-renewal and differentiation of mouse ES (mES) cells. These kinases display dynamic expression and activity changes during ES cell differentiation, suggesting distinct functions in the control of developmental fate. To test the hypothesis that c-Src and its closest phylogenetic relative, c-Yes, act in biological opposition to one another, I first showed that enforced expression of active c-Yes blocked ES cell differentiation to embryoid bodies by maintaining pluripotency gene expression. To determine the interplay of c-Src and c-Yes in mES cell fate determination, I employed a chemical genetics approach to generate c-Src and c-Yes mutants that are resistant to A-419259, a potent pyrrolopyrimidine inhibitor of the Src kinase family. This method allowed us to investigate individual kinase function in the presence of A-419259. I found that c-Src activity alone induces mES cell differentiation to the ectoderm and endoderm, while c-Yes inhibits this process. These studies show that even closely related kinases such as c-Src and c-Yes have unique and opposing functions in the same cell type.
While Src kinase signaling has been investigated in mES cells, the role of this kinase family in human ES (hES) cells is largely unknown. Using quantitative real-time RT-PCR, I determined the relative expression profile of individual SFK members in undifferentiated hES cells vs. embryoid bodies derived from them. Like mES cells, hES cells express multiple SFK members with dynamic transcription changes during EB differentiation, indicating that individual members may play non-redundant roles. To assess the role of SFK activity in hES cells, I treated hES cell cultures with SFK inhibitors. SFK inhibition maintained hES cell colony morphology and expression of the pluripotency marker Tra-1-60 in differentiation medium. These observations support a role for Src family kinase signaling in the regulation of hES fate, and suggest that some parallels may exist in mouse and human ES cells for this intracellular signaling network.


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Item Type: University of Pittsburgh ETD
Status: Unpublished
CreatorsEmailPitt UsernameORCID
Zhang, Xiongxiz51@pitt.eduXIZ51
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorSmithgall, Thomas Etsmithga@pitt.eduTSMITHGA
Committee ChairChaillet, J Richardchaillet@pitt.eduCHAILLET
Committee MemberHukriede, Neil Ahukriede@pitt.eduHUKRIEDE
Committee MemberSchatten,
Committee MemberWells, Alanwellsa@msx.upmc.eduAHW6
Date: 21 August 2013
Date Type: Publication
Defense Date: 11 June 2013
Approval Date: 21 August 2013
Submission Date: 20 August 2013
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 175
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Integrative Molecular Biology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: Embryonic Stem Cells, Src Family Kinases, Pluripotency, Differentiation, Cell Fate Determination
Date Deposited: 21 Aug 2013 14:26
Last Modified: 19 Dec 2016 14:41


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