Regulation of Pluripotency and Cellular Reprogramming by the Ubiquitin-Proteasome System

Howard Hughes Medical Institute and Department of Pathology, New York University School of Medicine, New York, NY 10016, USA
Cell stem cell (Impact Factor: 22.27). 10/2012; 11(6). DOI: 10.1016/j.stem.2012.09.011
Source: PubMed


Although transcriptional regulation of stem cell pluripotency and differentiation has been extensively studied, only a small number of studies have addressed the roles for posttranslational modifications in these processes. A key mechanism of posttranslational modification is ubiquitination by the ubiquitin-proteasome system (UPS). Here, using shotgun proteomics, we map the ubiquitinated protein landscape during embryonic stem cell (ESC) differentiation and induced pluripotency. Moreover, using UPS-targeted RNAi screens, we identify additional regulators of pluripotency and differentiation. We focus on two of these proteins, the deubiquitinating enzyme Psmd14 and the E3 ligase Fbxw7, and characterize their importance in ESC pluripotency and cellular reprogramming. This global characterization of the UPS as a key regulator of stem cell pluripotency opens the way for future studies that focus on specific UPS enzymes or ubiquitinated substrates.

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Available from: Charles L Farnsworth, Oct 08, 2015
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    • "Importantly, the somatic cell is not a tabula rasa and expresses genes that antagonize reprogramming, as has been shown for tumor suppressors (p53, INK4a/ARF, LATS2) (Kawamura et al., 2009; Qin et al., 2012; Zhao et al., 2008) and H3K9 methyltransferases (SETDB1, SUV39H, EHMT2) (Chen et al., 2013). In addition, focused RNAi screens have revealed other pathways that act as barriers to reprogramming, such as TGF-b signaling (Samavarchi-Tehrani et al., 2010), H3K79 methylation by DOT1L (Onder et al., 2012), or protein ubiquitination (Buckley et al., 2012). These findings suggest that other critical barriers to reprogramming are likely to exist, but no genome-wide functional screen has yet been carried out in mouse or human iPSC generation. "
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    ABSTRACT: Reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) holds enormous promise for regenerative medicine. To elucidate endogenous barriers limiting this process, we systematically dissected human cellular reprogramming by combining a genome-wide RNAi screen, innovative computational methods, extensive single-hit validation, and mechanistic investigation of relevant pathways and networks. We identify reprogramming barriers, including genes involved in transcription, chromatin regulation, ubiquitination, dephosphorylation, vesicular transport, and cell adhesion. Specific a disintegrin and metalloproteinase (ADAM) proteins inhibit reprogramming, and the disintegrin domain of ADAM29 is necessary and sufficient for this function. Clathrin-mediated endocytosis can be targeted with small molecules and opposes reprogramming by positively regulating TGF-β signaling. Genetic interaction studies of endocytosis or ubiquitination reveal that barrier pathways can act in linear, parallel, or feedforward loop architectures to antagonize reprogramming. These results provide a global view of barriers to human cellular reprogramming.
    Cell 07/2014; 158(2):449-61. DOI:10.1016/j.cell.2014.05.040 · 32.24 Impact Factor
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    • "Interestingly, the majority of the cells then became refractory to reprogramming, with few cells subsequently proceeding to the next steps of reprogramming. A possible explanation could be the " innate immunity " triggering protein degradation, which might explain why even in secondary systems not all cells reprogram, but can be rescued by additional overexpression of Yamanaka factors (Buckley et al., 2012; Polo et al., 2012). The observed changes in gene and protein expression correlate with a hierarchical sequence of events that reflect a deeper interaction between OKSM, co-factors and the chromatin, which together will ultimately dictate the epigenetic state. "
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    ABSTRACT: Despite advances in the field of somatic cell reprogramming, an understanding and exploration of the underlying mechanisms governing this process are only recently emerging. It is now increasingly apparent that key sequential events correlate with the reprogramming process; a process previously thought to be random and unpredictable is now looking, to a greater extent, defined and controlled. Herein, we will review the key cellular and molecular events associated with the reprogramming process, giving an integrative and conciliatory view of the different studies addressing the mechanism of nuclear reprogramming.
    Stem Cell Research 04/2014; 12(3):754-761. DOI:10.1016/j.scr.2014.03.007 · 3.69 Impact Factor
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    • "Thus, the long-lived stem cell may have mechanisms by which it rids itself of damaged proteins that are not present in its daughter cells, presumably working through the differential regulation of its protein degradation machinery. Consistent with this idea, human ESC cells have almost six times higher proteasomal activity (Buckley et al., 2012; Vilchez et al., 2012). "
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    ABSTRACT: Proteins are notorious for their unpleasant behavior-continually at risk of misfolding, collecting damage, aggregating, and causing toxicity and disease. To counter these challenges, cells have evolved elaborate chaperone and quality control networks that can resolve damage at the level of the protein, organelle, cell, or tissue. On the smallest scale, the integrity of individual proteins is monitored during their synthesis. On a larger scale, cells use compartmentalized defenses and networks of communication, capable sometimes of signaling between cells, to respond to changes in the proteome's health. Together, these layered defenses help protect cells from damaged proteins.
    Cell 03/2014; 157(1):52-64. DOI:10.1016/j.cell.2014.03.007 · 32.24 Impact Factor
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