Ledl, A., Schmidt, D. & Muller, S. Viral oncoproteins E1A and E7 and cellular LxCxE proteins repress SUMO modification of the retinoblastoma tumor suppressor. Oncogene 24, 3810-3818

Department of Molecular Cell Biology, Max Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried, Germany.
Oncogene (Impact Factor: 8.46). 06/2005; 24(23):3810-8. DOI: 10.1038/sj.onc.1208539
Source: PubMed

ABSTRACT The retinoblastoma tumor suppressor protein (pRB) is a major regulator of cell-cycle progression and cellular differentiation. Central to pRB function is the pocket domain, which serves as the main binding region for cellular regulators. In tumors pRB is frequently inactivated by mutations in the pocket domain or by binding of viral oncoproteins to this region. A characteristic feature of these viral oncoproteins and many cellular pRB-binding partners is an LxCxE sequence motif, which interacts with pRB's pocket domain. Here, we show that the ubiquitin-like modifier SUMO is covalently attached to a distinct residue (K720) of pRB within the B-box of the pocket region that binds LxCxE-motif proteins. We provide evidence that SUMO preferentially targets the active, hypophosphorylated form of pRB and show that tumorigenic mutations of pRB in the pocket domain lead to a loss of SUMOylation. Notably, the level of pRB SUMOylation is controlled by the interaction of pRB with viral and cellular LxCxE-motif proteins. Inhibitors of pRB function, including the viral oncoproteins E1A and E7 and the cellular E1A-like inhibitor of differentiation EID-1, completely abolish SUMO modification of pRB. Conversely, pRB mutants deficient in binding of LxCxE-motif proteins exhibit a drastically enhanced modification by SUMO. Finally, we provide evidence that SUMOylation can influence pRB function, as the SUMO-deficient pRB(K720R) mutant exerts a slightly higher repressive potential on an E2F-responsive reporter gene than wild-type pRB. Taken together, these data identify SUMO modification as a novel post-translational modification of pRB that may control pRB activity by modulating LxCxE-pocket interactions.

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    • "The Leu-Tyr-Cys-Tyr-Glu (L-Y-C-Y-E) motif (aa 22–26) comprises the core for pRb pocket binding (Comerford et al., 1991). The LYCYE pRb-binding motif of high risk HPV E7 proteins has been implicated in the immortalization and transformation of the host cell (Ledl et al., 2005; Narechania et al., 2004). "
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    ABSTRACT: Cervical cancer is the second most common cancer among women worldwide and remains a clinical problem despite improvements in early detection and therapy. The human papillomavirus (HPV) type 16 (HPV16) E7 oncoprotein expressed in cervical carcinoma cells are considered as attractive tumor-specific antigen targets for immunotherapy. Since the transformation potential of the oncogenes, vaccination based of these oncogenes is not safe. In present study, DNA vaccine expressing the modified variant with mutation in pRb-binding motif of the HPV-16 E7 oncoprotein was generated. A novel modified E7 gene with mutation in LYCYE motif was designed and constructed and the immunogenicity and antitumor effect of therapeutic DNA vaccines encoding the mutant and wild type of E7 gene were investigated. The L-Y-C-Y-E pRb-binding motif of E7 proteins has been involved in the immortalization and transformation of the host cell. The results showed that the mutant and wild type HPV-16 E7 vectors expressed the desired protein. Furthermore, the immunological mechanism behind mutant E7 DNA vaccine can be attributed at least partially to increased cytotoxic T lymphocyte, accompanied by the up-regulation of Th1-cytokine IFN- γ and TNF- β and down-regulation of Th3-cytokine TGF-β. Immunized mice with mutant plasmid demonstrated significantly stronger cell immune responses and higher levels of tumor protection than wild-type E7 DNA vaccine. The results exhibit that modified E7 DNA vaccine may be a promising candidate for development of therapeutic vaccine against HPV-16 cancers.
    Journal of Virological Methods 05/2014; DOI:10.1016/j.jviromet.2014.05.013 · 1.78 Impact Factor
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    • "), oxidative stress (Kurepa et al. 2003; Manza et al. 2004; Saitoh and Hinchey 2000; Zhou et al. 2004), DNA damage (reviewed by Jackson and Durocher 2013) and viral infection (Ledl et al. 2005; Muller and Dejean 1999; Parkinson and Everett 2000). Each of these factors impinges upon proteins involved in sumoylation to affect the level of their transcription or their activity (summarised in Table 2). "
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    ABSTRACT: A large number of proteins are modified post-translationally by the ubiquitin-like protein (Ubl) SUMO. This process, known as sumoylation, regulates the function, localisation and activity of target proteins as part of normal cellular metabolism, e.g., during development, and through the cell cycle, as well as in response to a range of stresses. In order to be effective, the sumoylation pathway itself must also be regulated. This review describes how the SUMOylation process is regulated. In particular, regulation of the SUMO conjugation and deconjugation machinery at the level of transcription and by post-translational modifications is discussed.
    Chromosoma 06/2013; 122(6). DOI:10.1007/s00412-013-0422-0 · 4.60 Impact Factor
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    • "For bacterial expression of GST-fusion proteins, cDNA sequences were inserted into pGEX-4T-1 (GE Healthcare) or pCDF PylT-1 (kindly provided by Jason W. Chin, MRC Laboratory for Molecular Biology, Cambridge, UK). All other plasmids are described previously (Finkbeiner et al., 2011; Haindl et al., 2008; Klein et al., 2009; Ledl et al., 2005). Sitedirected mutagenesis was carried out using the QuikChange Mutagenesis Kit (Stratagene) according to the manufacturer's instructions. "
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    ABSTRACT: The attachment of the SUMO modifier to proteins controls cellular signaling pathways through noncovalent binding to SUMO-interaction motifs (SIMs). Canonical SIMs contain a core of hydrophobic residues that bind to a hydrophobic pocket on SUMO. Negatively charged residues of SIMs frequently contribute to binding by interacting with a basic surface on SUMO. Here we define acetylation within this basic interface as a central mechanism for the control of SUMO-mediated interactions. The acetyl-mediated neutralization of basic charges on SUMO prevents binding to SIMs in PML, Daxx, and PIAS family members but does not affect the interaction between RanBP2 and SUMO. Acetylation is controlled by HDACs and attenuates SUMO- and PIAS-mediated gene silencing. Moreover, it affects the assembly of PML nuclear bodies and restrains the recruitment of the corepressor Daxx to these structures. This acetyl-dependent switch thus expands the regulatory repertoire of SUMO signaling and determines the selectivity and dynamics of SUMO-SIM interactions.
    Molecular cell 05/2012; 46(6):759-70. DOI:10.1016/j.molcel.2012.04.006 · 14.02 Impact Factor
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