Browsing by Subject "TFAP2C"

The proper differentiation of cells in early human development is essential for the success of a pregnancy. The first cell-fate decision occurs when totipotent cells differentiate into inner cell mass and trophectoderm, forming a blastocyst. Trophoblast cells differentiate from the trophectoderm and form the placenta. Defects in trophoblast development can lead to several pathologies, such as preeclampsia, miscarriage and intrauterine growth restriction. TFAP2C is a gene that is known to have an important role in the differentiation of trophoblast, although its function is not completely understood. Recent studies have suggested that TFAP2C has a promoter that has not been previously annotated, and it is not yet known what functions different TFAP2C promoters have. The aim of this thesis is to characterise how the activation of this novel TFAP2C promoter or the consensus promoter affects TFAP2C expression and whether the activation of different promoters changes how human pluripotent stem cells differentiate. In addition, this thesis aims to detect whether different TFAP2C protein variants, which are produced by these promoters, explain the possible variation in differentiation. TFAP2C promoters were activated with CRISPR activation, and different TFAP2C variants were expressed as transgenes in pluripotent stem cells. Gene expression was studied with immunocytochemistry and quantitative reverse transcription PCR. The activation of the consensus promoter increased TFAP2C expression more than the activation of the novel promoter. However, activation of both and expression of the protein variants produced from them made cells differentiate into trophoblast-like cells and express trophoblast markers. Nonetheless, the novel promoter and protein variant seemed to differentiate cells into trophoblast more efficiently. Because the activation of the promoters and the expression of their corresponding protein variants led to similar results, it seems likely that the alteration in differentiation is caused by the different TFAP2C protein variants. The functional difference between variants might be affected by the presence of a SUMOylation motif in the wild-type TFAP2C. These results may help to better understand the role of TFAP2C in human embryonic development and create applications for assisted reproduction and infertility treatments, as well as facilitate the development of cell models that can be used in research and therapy.