Nuclear translocation of Skp2 facilitates its destruction in response to TGFβ signaling

University of Pittsburgh, Pittsburgh, Pennsylvania, United States
Cell cycle (Georgetown, Tex.) (Impact Factor: 4.57). 01/2011; 10(2):285-92. DOI: 10.4161/cc.10.2.14517
Source: PubMed

ABSTRACT Skp2, a F-box protein that determines the substrate specificity for SCF ubiquitin ligase, has recently been demonstrated to be degraded by Cdh1/APC in response to TGFβ signaling. The TGFβ-induced Skp2 proteolysis results in the stabilization of p27 that is necessary to facilitate TGFβ cytostatic effect. Previous observation from immunocytochemistry indicates that Cdh1 principally localizes in the nucleus while Skp2 mainly localizes in the cytosol, which leaves us a puzzle on how Skp2 is recognized and then ubiquitylated by Cdh1/APC in response to TGFβ stimulation. Here, we report that Skp2 is rapidly translocated from the cytosol to the nucleus upon the cellular stimulation with TGFβ. Using a combinatorial approach of immunocytochemistry, biochemical-fraction-coupled immunoprecipitation, mutagenesis as well as protein degradation assay, we have demonstrated that the TGFβ-induced Skp2 nucleus translocation is critical for TGFβ cytostatic effect that allows physical interaction between Cdh1 and Skp2 and in turn facilitates the Skp2 ubquitylation by Cdh1/APC. Disruption of nuclear localization motifs on Skp2 stabilizes Skp2 in the presence of TGF-β signaling, which attenuates TGFβ-induced p27 accumulation and antagonizes TGFβ-induced growth inhibition. Our finding reveals a cellular mechanism that facilitates Skp2 ubiquitylation by Cdh1/APC in response to TGFβ.

22 Reads
  • Source
    • "Thus, in addition to the known cell cycle–controlled activation of APC/C-Cdh1 (Morgan, 1999; Peters, 2006), which generates nuclear Cdh1 activity, temporal control of this proteolytic pathway may also occur at the level of cytoplasmic substrates . This spatiotemporal mechanism for the control of Cdh1- mediated proteolysis may be conserved among eukaryotes, since the APC/C and Cdh1 were reported to localize to the nucleus in mammalian cells (Jörgensen et al., 1998; Gieffers et al., 1999; Topper et al., 2002; Zhou et al., 2003), and degradation of human Skp2 by the APC/C-Cdh1 pathway required Skp2 nuclear import, which was controlled by transforming growth factor β signaling (Hu, Liu, et al., 2011). In fact, substrate-level control of Cdh1-de- pendent proteolysis may even be more widespread in metazoan organisms, in which the APC/C-Cdh1 ubiquitin ligase functions in nonmitotic processes such as metabolism, cell differentiation, and survival (Gieffers et al., 1999; Eguren et al., 2011; Almeida, 2012). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The anaphase-promoting complex/cyclosome (APC/C) controls a variety of cellular processes through its ability to target numerous protein substrates for timely degradation. Substrate selection by this ubiquitin ligase depends on related activator proteins, Cdc20 and Cdh1, which bind and activate the APC/C at distinct cell cycle stages. Biochemical and structural studies revealed that Cdc20 and Cdh1 carry conserved receptor domains to recognize specific sequence motifs in substrates, such as d- and KEN-boxes. The mechanisms for ordered degradation of APC/C substrates, however, remain incompletely understood. Here, we describe minimal degradation sequences (degrons) sufficient for rapid APC/C-Cdh1-specific in vivo degradation. The polo kinase Cdc5-derived degron contained an essential KEN motif, while a single RxxL-type d-box was the relevant signal in the Cdc20-derived degradation domain, indicating that either motif may support specific recognition by Cdh1. In both degrons, the APC/C recognition motif was flanked by a nuclear localization sequence. Forced localization of the degron constructs revealed that proteolysis mediated by APC/C-Cdh1 is restricted to the nucleus and maximally active in the nucleoplasm. Levels of Iqg1, a cytoplasmic Cdh1 substrate, decreased detectably later than the nuclear-localized Cdh1 substrate Ase1, indicating that confinement to the nucleus may allow for temporal control of APC/C-Cdh1-mediated proteolysis. © 2014 by The American Society for Cell Biology.
    Molecular Biology of the Cell 12/2014; 26(5). DOI:10.1091/mbc.E14-09-1342 · 4.47 Impact Factor
  • Source
    • "It was also suggested that TGF-β promotes translocation of Skp2 into the nucleus, where it is degraded by the anaphase-promoting complex/cyclosome (APC/C)-Cdh1 E3 ligase. In addition, TGF-β decreases Cks1 mRNA expression, which allows p27 to accumulate following G1 arrest [50–52]. Taken together, these findings indicate that TGF-β is an important upstream signal that regulates the Skp2/p27 axis. "
    [Show abstract] [Hide abstract]
    ABSTRACT: S-phase kinase-associated protein 2 (Skp2) is an F-box protein component of the Skp/Cullin/F-box-type E3 ubiquitin ligase that targets several cell cycle regulatory proteins for degradation through the ubiquitin-dependent pathway. Skp2-mediated degradation of p27, a cyclin-dependent kinase inhibitor, is involved in cell cycle regulation. Tubular epithelial cell proliferation is a characteristic feature of renal damage that is apparent in the early stages of nephropathy. The p27 level is associated with the progression of renal injury, and increased Skp2 expression in progressive nephropathy is implicated in decreases of p27 expression. In Skp2(-/-) mice, renal damage caused by unilateral ureteral obstruction (UUO) was ameliorated by p27 accumulation, mainly in tubular epithelial cells. However, the amelioration of UUO-induced renal injury in Skp2(-/-) mice was prevented by p27 deficiency in Skp2(-/-)/p27(-/-) mice. These results suggest that the Skp2-mediated reduction in p27 is a pathogenic activity that occurs during the progression of nephropathy. Here, we discuss the roles of the Skp2/p27 axis and/or related signaling pathways/components in the progression of chronic nephropathy.
    Cellular and Molecular Life Sciences CMLS 12/2012; 70(18). DOI:10.1007/s00018-012-1232-x · 5.81 Impact Factor
  • Source
    • "In the vast majority of cases, SKP2 overexpression inversely correlates with p27 KIP1 expression (Nakayama and Nakayama, 2006; Frescas and Pagano, 2008), which is consistent p27 KIP1 being a key target of SKP2 and to be rarely mutated in cancer (Chu et al., 2008). Finally, in recent years, SKP2 overexpression has been shown to mediate resistance to TRAIL induced apoptosis (Schuler et al., 2011), and radio-and chemoresistance of human cancer cells (Ishii et al., 2004; Davidovich et al., 2008; Chan et al., 2012; Totary-Jain et al., 2012; Wang et al., 2012b). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Neuroblastoma is the most common extra-cranial solid tumor of childhood. Despite significant advances, it currently still remains one of the most difficult childhood cancers to cure, with less than 40% of patients with high-risk disease being long-term survivors. MYCN is a proto-oncogene implicated to be directly involved in neuroblastoma development. Amplification of MYCN is associated with rapid tumor progression and poor prognosis. Novel therapeutic strategies which can improve the survival rates whilst reducing the toxicity in these patients are therefore required. Here we discuss genes regulated by MYCN in neuroblastoma, with particular reference to p53, SKP2, and DKK3 and strategies that may be employed to target them.
    Frontiers in Oncology 11/2012; 2:173. DOI:10.3389/fonc.2012.00173
Show more