Niwa, H., Masui, S., Chambers, I., Smith, A. G. & Miyazaki, J. Phenotypic complementation establishes requirements for specific POU domain and generic transactivation function of Oct-3/4 in embryonic stem cells. Mol. Cell. Biol. 22, 1526-1536

Stem Cell Regulation Research, Area of Molecular Therapeutics, Course of Advanced Medicine, Osaka University Graduate School of Medicine, Suita C, Osaka 565-0871, Japan.
Molecular and Cellular Biology (Impact Factor: 5.04). 04/2002; 22(5):1526-36. DOI: 10.1128/MCB.22.5.1526-1536.2002
Source: PubMed

ABSTRACT Transcription factors of the POU family govern cell fate through combinatorial interactions with coactivators and corepressors. The POU factor Oct-3/4 can define differentiation, dedifferentation, or self-renewal of pluripotent embryonic stem (ES) cells in a sensitive, dose-dependent manner (H. Niwa, J.-I. Miyazali, and A. G. Smith, Nat. Genet. 24:372-376, 2000). Here we have developed a complementation assay based on the ability of Oct-3/4 transgenes to rescue self-renewal in conditionally null ES cells and used this to define which domains of Oct-3/4 are required to sustain the undifferentiated stem cell phenotype. Surprisingly, we found that molecules lacking either the N-terminal or C-terminal transactivation domain, though not both, can effectively replace full-length Oct-3/4. Furthermore, a fusion of the heterologous transactivation domain of Oct-2 to the Oct-3/4 POU domain can also sustain self-renewal. Thus, the unique function of Oct-3/4 in ES cell propagation resides in combination of the specific POU domain with a generic proline-rich transactivation domain. Interestingly, however, Oct-3/4 target gene expression elicited by the N- and C-terminal transactivation domains is not identical, indicating that at least one class of genes activated by Oct-3/4 is not required for ES cell propagation.

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Available from: Jun-ichi Miyazaki, Jan 10, 2014
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    • "Immunophenotypic characterization of both cell types demonstrated the presence of the common, well-defined human mesenchymal stem cell markers CD90, CD44, CD73, CD166, CD105, CD29, CD13, CD49e, CD54, as well as the embryonic stem-cell markers TRA-1-60, TRA-1-81, SSEA-3 and -4 and STRO-1 (Bilic et al. 2008; Díaz- Prado et al. 2010; Stadler et al. 2008). In addition, amniotic epithelial cells express Oct-4 and Nanog, two transcription factors known to be required for selfrenewal and pluripotency (Niwa et al. 2002; Chambers et al. 2003). Unlike human embryonic stem cells, amnion epithelial cells do not express telomerase and are nontumorigenic upon transplantation. "
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    • "Twenty-four to forty-eight hours after transfection, the cells were collected and lysed, and luciferase assay was performed with the Dual-Luciferase Reporter Assay System (Promega). Reporter plasmids were a kind gift from Professor Hitoshi Niwa (Riken Institute, Japan) (Niwa et al., 2002). The expression vector used was the pCAG-IP plasmid. "
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    • "Indeed, most of maternal mutant (M) spg embryos and MZ embryos rescued by Pou5f1 mRNA injection develop normally, despite the vastly different Pou5f1 activity levels in these embryos. This situation apparently differs from the mode of Pou5f1 action in ES cells, where the levels of Pou5f1/Oct4 need to be precisely tuned (Niwa et al., 2000, 2002). However, the situation in the later stages of mammalian development, that is, in the pluripotent epiblast cells after implantation, may be different and it would be interesting to test, whether the zebrafish model regulatory scenario fits to the expression of double Pou5f1 and SoxB1 targets in these systems. "
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