The Effect of SETD3 on Beta-Catenin and Canonical Wnt Signaling Pathway Activity in Mouse Embryonic Stem Cells

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2023-1-16

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Alganatay, Ceren

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Mouse embryonic stem cells (mESCs) are self-renewing, pluripotent cells that can differentiate into endoderm, mesoderm and neuroectoderm. As differentiation requires deactivation of pluripotency factors and activation of germ layer specific genes, many epigenetic factors play an essential role. A previously performed shRNA screen showed that SETD3 which is a SET-domain containing epigenetic factor, is an essential factor for mesendoderm differentiation of mESCs. Our preliminary data demonstrated that when mESCs were grown in standard serum-containing mESC medium prior to directed differentiation, wild type mESCs can successfully differentiate into mesendoderm, while SETD3 knockout (setd3∆) mESCs cannot. According to our preliminary data, we aimed to identify the effect of SETD3 on the levels and localization of β-catenin as well as on the canonical Wnt signaling pathway activity in mESCs. Proximity ligation assay revealed that SETD3 and β-catenin are interacting in mESCs, further supporting the role of canonical Wnt signaling pathway. Even though immunocytochemistry assay performed wild type and setd3∆ mESCs grown in the same medium did not show significant localization differences, western blot experiments performed with nuclear fractions showed that setd3∆ mESCs grown in standard serum-containing mESC medium had lower levels of β-catenin than wild type mESCs. Additionally, luciferase assay results demonstrated that, regardless of the growth medium, absence of SETD3 results with significantly lower β-catenin transcriptional activity. Observed lower nuclear βcatenin levels and lower canonical Wnt signaling pathway activity in the absence of SETD3 might explain the mesendoderm differentiation defect of setd3∆ mESCs grown in standard serum-containing mESC medium.

Subject Keywords

SETD3, Mouse Embryonic Stem Cells, Canonical Wnt Signaling Pathway, β-catenin

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https://hdl.handle.net/11511/102077

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Graduate School of Natural and Applied Sciences, Thesis

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Citation Formats

C. Alganatay, “The Effect of SETD3 on Beta-Catenin and Canonical Wnt Signaling Pathway Activity in Mouse Embryonic Stem Cells,” M.S. - Master of Science, Middle East Technical University, 2023.