Date of Graduation
12-2014
Document Type
Dissertation (PhD)
Program Affiliation
Genes and Development
Degree Name
Doctor of Philosophy (PhD)
Advisor/Committee Chair
Pierre D. McCrea
Committee Member
Jae-il Park
Committee Member
Xiaobing Shi
Committee Member
Zhimin Lu
Committee Member
Amy K. Sater
Abstract
The canonical-Wnt pathway and beta-catenin have been extensively studied to determine their contributions to stem cell biology, but less is known about p120-catenin in the nuclear compartment. P120 is developmentally required as a consequence of its biochemical and functional interactions with cadherins, small-GTPases and transcriptional regulators. We report here that p120-catenin binds to and negatively regulates REST and CoREST, that others have indicated form a repressive complex having diverse key roles in developmental and pathologic gene regulation. We thus provide the first evidence for a direct upstream modulator of REST/CoREST function. Using mouse embryonic stem cells (mESCs), mammalian cell lines, Xenopus embryos, and in vitro systems, we show that p120 directly binds to the zinc finger/DNA-binding region of REST, as well as to CoREST. Chromatin immuno-precipitation and other approaches indicate that p120 protein levels negatively determine the extent of REST/CoREST bound to RE1 consensus binding sites and negatively influence REST/CoREST protein stability. As would be predicted, p120 overexpression and depletion have complementary effects upon REST/CoREST gene-target activity. Thus, p120 depletion in mESCs reduces REST/CoREST gene-target expression, while p120 overexpression has a converse effect. Importantly, p120 levels modulate the mRNA and protein levels of Oct4, Nanog, and Sox2, and have an impact upon the differentiation of mESCs towards neural fates. In assessing potential upstream inputs of this novel p120-REST/ CoREST pathway, REST gene targets were found to respond to the level of E-cadherin, with cadherin effects being dependent on p120-catenin as predicted. In summary, at both biochemical and functional levels, our findings reveal a central role of p120-catenin in the derepression (activation) of genes directly controlled by REST/CoREST, and in the modulation of stem cells.
Keywords
p120-catenin, REST, CoREST, mouse embryonic stem cell, differentiation, pluripotency