80
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: not found

      An Oct4-Centered Protein Interaction Network in Embryonic Stem Cells

      Read this article at

      ScienceOpenPublisherPMC
      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Summary

          Transcription factors, such as Oct4, are critical for establishing and maintaining pluripotent cell identity. Whereas the genomic locations of several pluripotency transcription factors have been reported, the spectrum of their interaction partners is underexplored. Here, we use an improved affinity protocol to purify Oct4-interacting proteins from mouse embryonic stem cells (ESCs). Subsequent purification of Oct4 partners Sall4, Tcfcp2l1, Dax1, and Esrrb resulted in an Oct4 interactome of 166 proteins, including transcription factors and chromatin-modifying complexes with documented roles in self-renewal, but also many factors not previously associated with the ESC network. We find that Esrrb associated with the basal transcription machinery and also detect interactions between transcription factors and components of the TGF-β, Notch, and Wnt signaling pathways. Acute depletion of Oct4 reduced binding of Tcfcp2l1, Dax1, and Esrrb to several target genes. In conclusion, our purification protocol allowed us to bring greater definition to the circuitry controlling pluripotent cell identity.

          Highlights

          ► An improved affinity method identified an Oct4 interactome of >160 proteins ► Iterative purification with Oct4 interactors helped define the network ► Network shows links to the basal transcription machinery and signaling pathways ► Transcription factor interactions may facilitate recruitment to the genome

          Related collections

          Most cited references 38

          • Record: found
          • Abstract: found
          • Article: not found

          In vitro reprogramming of fibroblasts into a pluripotent ES-cell-like state.

          Nuclear transplantation can reprogramme a somatic genome back into an embryonic epigenetic state, and the reprogrammed nucleus can create a cloned animal or produce pluripotent embryonic stem cells. One potential use of the nuclear cloning approach is the derivation of 'customized' embryonic stem (ES) cells for patient-specific cell treatment, but technical and ethical considerations impede the therapeutic application of this technology. Reprogramming of fibroblasts to a pluripotent state can be induced in vitro through ectopic expression of the four transcription factors Oct4 (also called Oct3/4 or Pou5f1), Sox2, c-Myc and Klf4. Here we show that DNA methylation, gene expression and chromatin state of such induced reprogrammed stem cells are similar to those of ES cells. Notably, the cells-derived from mouse fibroblasts-can form viable chimaeras, can contribute to the germ line and can generate live late-term embryos when injected into tetraploid blastocysts. Our results show that the biological potency and epigenetic state of in-vitro-reprogrammed induced pluripotent stem cells are indistinguishable from those of ES cells.
            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            Conversion of embryonic stem cells into neuroectodermal precursors in adherent monoculture.

            Mouse embryonic stem (ES) cells are competent for production of all fetal and adult cell types. However, the utility of ES cells as a developmental model or as a source of defined cell populations for pharmaceutical screening or transplantation is compromised because their differentiation in vitro is poorly controlled. Specification of primary lineages is not understood and consequently differentiation protocols are empirical, yielding variable and heterogeneous outcomes. Here we report that neither multicellular aggregation nor coculture is necessary for ES cells to commit efficiently to a neural fate. In adherent monoculture, elimination of inductive signals for alternative fates is sufficient for ES cells to develop into neural precursors. This process is not a simple default pathway, however, but requires autocrine fibroblast growth factor (FGF). Using flow cytometry quantitation and recording of individual colonies, we establish that the bulk of ES cells undergo neural conversion. The neural precursors can be purified to homogeneity by fluorescence activated cell sorting (FACS) or drug selection. This system provides a platform for defining the molecular machinery of neural commitment and optimizing the efficiency of neuronal and glial cell production from pluripotent mammalian stem cells.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: not found

              A protein interaction network for pluripotency of embryonic stem cells.

              Embryonic stem (ES) cells are pluripotent and of therapeutic potential in regenerative medicine. Understanding pluripotency at the molecular level should illuminate fundamental properties of stem cells and the process of cellular reprogramming. Through cell fusion the embryonic cell phenotype can be imposed on somatic cells, a process promoted by the homeodomain protein Nanog, which is central to the maintenance of ES cell pluripotency. Nanog is thought to function in concert with other factors such as Oct4 (ref. 8) and Sox2 (ref. 9) to establish ES cell identity. Here we explore the protein network in which Nanog operates in mouse ES cells. Using affinity purification of Nanog under native conditions followed by mass spectrometry, we have identified physically associated proteins. In an iterative fashion we also identified partners of several Nanog-associated proteins (including Oct4), validated the functional relevance of selected newly identified components and constructed a protein interaction network. The network is highly enriched for nuclear factors that are individually critical for maintenance of the ES cell state and co-regulated on differentiation. The network is linked to multiple co-repressor pathways and is composed of numerous proteins whose encoding genes are putative direct transcriptional targets of its members. This tight protein network seems to function as a cellular module dedicated to pluripotency.
                Bookmark

                Author and article information

                Contributors
                Journal
                Cell Stem Cell
                Cell Stem Cell
                Cell Stem Cell
                Cell Press
                1934-5909
                1875-9777
                02 April 2010
                02 April 2010
                : 6
                : 4
                : 369-381
                Affiliations
                [1 ]Department of Cell Biology, Erasmus MC, Dr. Molewaterplein 50, 3015GE Rotterdam, The Netherlands
                [2 ]Proteomics Center, Erasmus MC, Dr. Molewaterplein 50, 3015GE Rotterdam, The Netherlands
                [3 ]MRC Centre for Regenerative Medicine, Institute for Stem Cell Research, School of Biological Sciences, University of Edinburgh, King's Buildings, West Mains Road, Edinburgh EH9 3JQ, UK
                Author notes
                []Corresponding author r.poot@ 123456erasmusmc.nl
                Article
                STEM610
                10.1016/j.stem.2010.02.014
                2860243
                20362541
                © 2010 ELL & Excerpta Medica.

                This document may be redistributed and reused, subject to certain conditions.

                Categories
                Resource

                Molecular medicine

                stemcell

                Comments

                Comment on this article