Date of Award

2009

Degree Type

Thesis

Degree Name

Master of Science

Program

Biology

Supervisor

Dr. Gregory M. Kelly

Abstract

Mouse F9 teratocarcinoma cells in culture differentiate into primitive endoderm when treated with retinoic acid and into parietal endoderm with subsequent treatment with dibutyryl cAMP. This in vitro model has been studied extensively as these chemically induced events mimic one o f the earliest epithelial-to-mesenchymal transitions in mouse embryogenesis. During differentiation to primitive endoderm Wnt6 expression is up- regulated by RA, and the result is the activation o f the canonical W nt/p-catenin signaling pathway. The factor(s) responsible for the activation o f the Wnt6 gene is not known, but in silico analysis reveals that its promoter region contains a putative binding site for the transcription factor GATA6. In this study, the expression of Gata6 following retinoic acid treatment was examined and found to be up-regulated during primitive and parietal endoderm differentiation. Overexpression of Gata6 alone induced biochemical,

molecular, and morphological markers o f primitive endoderm and was sufficient in up- regulating the expression of Wnt6. Furthermore, this up-regulation was accompanied by the activation of the canonical Wnt/p-catenin signaling pathway, as evident by the increase in phospho-GSK3p levels. Gata6 expressing cells were also capable of com pleting the epithelial-to-m esenchym al transition and differentiating into parietal endoderm when treated with dibutyryl cAMP. Together, these results show that Gata6 overexpression is sufficient to up-regulate the expression of Wnt6, a signaling molecule previously reported by our lab to activate the canonical Wnt/p-catenin pathway, and provides new insight to the signaling mechanisms involved in the specification of primitive endoderm.

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