Article

A DNA Repair Complex Functions as an Oct4/Sox2 Coactivator in Embryonic Stem Cells

Howard Hughes Medical Institute, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
Cell (Impact Factor: 32.24). 09/2011; 147(1):120-31. DOI: 10.1016/j.cell.2011.08.038
Source: PubMed

ABSTRACT

The transcriptional activators Oct4, Sox2, and Nanog cooperate with a wide array of cofactors to orchestrate an embryonic stem (ES) cell-specific gene expression program that forms the molecular basis of pluripotency. Here, we report using an unbiased in vitro transcription-biochemical complementation assay to discover a multisubunit stem cell coactivator complex (SCC) that is selectively required for the synergistic activation of the Nanog gene by Oct4 and Sox2. Purification, identification, and reconstitution of SCC revealed this coactivator to be the trimeric XPC-nucleotide excision repair complex. SCC interacts directly with Oct4 and Sox2 and is recruited to the Nanog and Oct4 promoters as well as a majority of genomic regions that are occupied by Oct4 and Sox2. Depletion of SCC/XPC compromised both pluripotency in ES cells and somatic cell reprogramming of fibroblasts to induced pluripotent stem (iPS) cells. This study identifies a transcriptional coactivator with diversified functions in maintaining ES cell pluripotency and safeguarding genome integrity.

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    • "While the XPC impact on OGG1 activity was previously described by D'Errico et al. [8]. the functional interaction between XPC and APE1 revealed in this work is a novel observation on the role of XPC in repairing oxidized lesions. Studies support additional XPC functions in different cellular mechanisms independent of the NER pathway[8,9,383940414243. In the transcription process without DNA damage, studies show that transcriptional pre-initiation (PIC) complex formation was preceded by sequential recruitment of certain NER factors, including XPC protein . "
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    ABSTRACT: Oxidative DNA damage is considered to be a major cause of neurodegeneration and internal tumors observed in syndromes that result from nucleotide excision repair (NER) deficiencies, such as Xeroderma Pigmentosum (XP) and Cockayne Syndrome (CS). Recent evidence has shown that NER aids in removing oxidized DNA damage and may interacts with base excision repair (BER) enzymes. Here, we investigated APE1 and OGG1 expression, localization and activity after oxidative stress in XPC-deficient cells. The endogenous APE1 and OGG1 mRNA levels were lower in XPC-deficient fibroblasts. However, XPC-deficient cells did not show hypersensitivity to oxidative stress compared with NER-proficient cells. To confirm the impact of an XPC deficiency in regulating APE1 and OGG1 expression and activity, we established an XPC-complemented cell line. Although the XPC complementation was only partial and transient, the transfected cells exhibited greater OGG1 expression and activity compared with XPC-deficient cells. However, the APE1 expression and activity did not significantly change. Furthermore, we observed a physical interaction between the XPC and APE1 proteins. Together, the results indicate that the responses of XPC-deficient cells under oxidative stress may not only be associated with NER deficiency per se but may also include new XPC functions in regulating BER proteins.
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    • "Transcriptional regulation can be measured, described, and understood at several levels. Because much of the transcription reaction can be reconstituted in vitro from purified components, some aspects of its regulation can be addressed at the molecular level (Lemon et al. 2001; Fong et al. 2011, 2014). The structures of RNA polymerase II (Cramer et al. 2000; Gnatt et al. 2001) and many components of the preinitiation complex have been resolved at atomic resolution. "
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    ABSTRACT: Transcription, the first step of gene expression, is exquisitely regulated in higher eukaryotes to ensure correct development and homeostasis. Traditional biochemical, genetic, and genomic approaches have proved successful at identifying factors, regulatory sequences, and potential pathways that modulate transcription. However, they typically only provide snapshots or population averages of the highly dynamic, stochastic biochemical processes involved in transcriptional regulation. Single-molecule live-cell imaging has, therefore, emerged as a complementary approach capable of circumventing these limitations. By observing sequences of molecular events in real time as they occur in their native context, imaging has the power to derive cause-and-effect relationships and quantitative kinetics to build predictive models of transcription. Ongoing progress in fluorescence imaging technology has brought new microscopes and labeling technologies that now make it possible to visualize and quantify the transcription process with single-molecule resolution in living cells and animals. Here we provide an overview of the evolution and current state of transcription imaging technologies. We discuss some of the important concepts they uncovered and present possible future developments that might solve long-standing questions in transcriptional regulation.
    Preview · Article · Jan 2016 · Cold Spring Harbor Symposia on Quantitative Biology
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    • "There are additional examples of DNA repair enzymes functioning as potential coactivators at distal enhancers. The XPC complex can serve both as a classical DNA repair factor and as a transcriptional coactivator in ES cells (Figure 2A) (Fong et al., 2011). Moreover, components of the nucleotide excision repair pathway have been implicated in transcriptional activation upon DNA demethylation and gene looping (Le May et al., 2012), whereas the base-excision repair enzyme TDG is emerging as a key player in regulating DNA methylation (Wyatt, 2013). "
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    ABSTRACT: Comparative genome analyses reveal that organismal complexity scales not with gene number but with gene regulation. Recent efforts indicate that the human genome likely contains hundreds of thousands of enhancers, with a typical gene embedded in a milieu of tens of enhancers. Proliferation of cis-regulatory DNAs is accompanied by increased complexity and functional diversification of transcriptional machineries recognizing distal enhancers and core promoters and by the high-order spatial organization of genetic elements. We review progress in unraveling one of the outstanding mysteries of modern biology: the dynamic communication of remote enhancers with target promoters in the specification of cellular identity.
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