SUMOylation regulates p27<sup>Kip1</sup>stability and localization in response to TGFβ

Lovisa, Sara; Citro, Simona; Sonego, Maura; Dall'Acqua, Alessandra; Ranzuglia, Valentina; Berton, Stefania; Colombatti, Alfonso; Belletti, Barbara; Chiocca, Susanna; Schiappacassi, Mónica; Baldassarre, Gustavo

doi:10.1093/jmcb/mjv056

Cited by 10 publications

(14 citation statements)

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“…On the other side, accumulation of nuclear p27 seems to be essential for ensuring cell cycle arrest following antiproliferative stimuli, such as contact inhibition 9 or TGFb stimulation. 10 Overall, our recent findings support the notion that p27 plays a very complex role in the control of cell proliferation. Different layers of control, mostly based on post-translational modifications, dictate its subcellular localization, expression, and activity in order to allow fine integration of mitogenic and antimitogenic stimuli before the decision of whether the cell will proliferate is made.…”

Section: P27 Gets On the Ras Stagesupporting

confidence: 84%

p27kip1: An all-round tumor suppressor

Belletti

Fabris

Baldassarre

2016

Molecular & Cellular Oncology

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Our recent study has uncovered an additional mechanism by which the cell cycle inhibitor p27kip1 controls cell proliferation. Through its effect on the activity of the microtubule destabilizing protein Stathmin, p27kip1 modulates full H-Ras activation and, as a consequence, the MAPK signaling cascade. This regulatory mechanism influences the cell cycle in vitro and tissue and/or organ growth in mice, in vivo and, when unbalanced, may lead to uncontrolled proliferation and tumor onset.

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Section: P27 Gets On the Ras Stagesupporting

confidence: 84%

p27kip1: An all-round tumor suppressor

Belletti

Fabris

Baldassarre

2016

Molecular & Cellular Oncology

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“…Extraction of total proteins, Western blot, and immunoprecipitations analyses were performed as described (Sonego et al , ; Fabris et al , ; Lovisa et al , ). Primary antibodies used were as follows: vinculin (N‐19, 1:1,000), CDK6 (C‐21, sc‐177, 1:800), CDK4 (C‐22, sc‐260, 1:400), pRB1S780 (sc‐12901, 1:400), PSF (39‐1, sc‐101137, 1:800), CHK1 (G‐4, sc‐8408, 1:500), luciferase (sc‐32896), lamin A (C‐20 sc‐6214) (Santa Cruz Biotechnology); FOXO3 (75D8, #2497, 1:600), cyclin D3 (DCS22, #2936, 1:500), pCHK1S296 (#2349, 1:500), H2AX (D17A3, #7631, 1:500), ATR (#2790, 1:500), ATM (#2873, 1:500), and cleaved caspase‐3 Asp175 (#9661, 1:500) (Cell Signaling); cyclin D1 (DCS‐6, CC12, 1:1,000) (Millipore); γH2AXS139 (#2577, 1:500) (Upstate Biotechnology); RB1 (554136, 1:500) and GRB2 (610111, 1:300) (BD Biosciences); actin (A5060, 1:500), tubulin (T5168, 1:1,000), Flag (F3040), OP18 (O0138, 1:1,000), and V5 (A7345)‐ and HA (A2095)‐agarose conjugated (Sigma‐Aldrich Co); GFP (11 814 440 001, 1:500) (Roche); Ki67 (ab15580, 1:1,000) (Abcam).…”

Section: Methodsmentioning

confidence: 99%

CDK6 protects epithelial ovarian cancer from platinum‐induced death via FOXO3 regulation

et al. 2017

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Epithelial ovarian cancer (EOC) is an infrequent but highly lethal disease, almost invariably treated with platinum‐based therapies. Improving the response to platinum represents a great challenge, since it could significantly impact on patient survival. Here, we report that silencing or pharmacological inhibition of CDK6 increases EOC cell sensitivity to platinum. We observed that, upon platinum treatment, CDK6 phosphorylated and stabilized the transcription factor FOXO3, eventually inducing ATR transcription. Blockage of this pathway resulted in EOC cell death, due to altered DNA damage response accompanied by increased apoptosis. These observations were recapitulated in EOC cell lines in vitro, in xenografts in vivo, and in primary tumor cells derived from platinum‐treated patients. Consistently, high CDK6 and FOXO3 expression levels in primary EOC predict poor patient survival. Our data suggest that CDK6 represents an actionable target that can be exploited to improve platinum efficacy in EOC patients. As CDK4/6 inhibitors are successfully used in cancer patients, our findings can be immediately transferred to the clinic to improve the outcome of EOC patients.

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“…Spagnuolo et al reported that reduction of sumoylation in S100A4 would decrease its nuclear import brought by low dose paclitaxel, leading to halts of tumor invasiveness and hematogenous metastatization [ 4 ]. Baldassarre and Schiappacassi et al reported that K134 of p27kip1 is the sumoylation site in tumor-derived cell in response to TGF-β [ 24 ]. There are no other study focusing on the post-translational regulation in cholangiocarcinoma.…”

Section: Discussionmentioning

confidence: 99%

“…This assay was undertaken as described by Lovista et al [ 24 ]. The whole reaction was undertaken in 30 μl reaction system with 50 mM Tris–HCl (pH 7.4), 150 mM NaCl, 5 mM MgCl 2 , 2 mM DTT, 2 mM ATP, 270 ng SAE1/SAE2 (Boston Biochem, USA), 300 ng UBC9 (Boston Biochem, USA), 4 µg SUMO1 (Boston Biochem, USA), and 3 µg His-p27 and incubated for 2 h at 30 °C.…”

Section: Methodsmentioning

confidence: 99%

“…However, it is still elusive how this protein is transported to the cytoplasm and the possible epigenetic modification processes as sumoylation contributing in this process. Indeed, only Baldassarre and Schiappacassi et al reported that K134 of p27kip1 is responsible for its sumoylation before it is transported through nuclear membrane in 293T/17 and Hela cells in response to TGF-β [ 24 ]. What’s happened of p27kip1 in the QBC939 and the role of sumoylation in this process is still unknown.…”

Section: Introductionmentioning

confidence: 99%

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Sumoylation in p27kip1 via RanBP2 promotes cancer cell growth in cholangiocarcinoma cell line QBC939

et al. 2017

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BackgroundCholangiocarcinoma is one of the deadly disease with poor 5-year survival and poor response to conventional therapies. Previously, we found that p27kip1 nuclear-cytoplasmic translocation confers proliferation potential to cholangiocarcinoma cell line QBC939 and this process is mediated by crm-1. However, no other post-transcriptional regulation was found in this process including sumoylation in cholangiocarcinoma.ResultsIn this study, we explored the role of sumoylation in the nuclear-cytoplasmic translocation of p27kip1 and its involvement of QBC939 cells’ proliferation. First, we identified K73 as the sumoylation site in p27kip1. By utilizing plasmid flag-p27kip1, HA-RanBP2, GST-RanBP2 and His-p27kip1 and immunoprecipitation assay, we validated that p27kip1 can serve as the sumoylation target of RanBP2 in QBC939. Furthermore, we confirmed crm-1’s role in promoting nuclear-cytoplasmic translocation of p27kip1 and found that RanBP2’s function relies on crm-1. However, K73R mutated p27kip1 can’t be identified by crm-1 or RanBP2 in p27kip1 translocation process, suggesting sumoylation of p27kip1 via K73 site is necessary in this process by RanBP2 and crm-1. Phenotypically, the overexpression of either RanBP2 or crm-1 can partially rescue the anti-proliferative effect brought by p27kip1 overexpression in both the MTS and EdU assay. For the first time, we identified and validated the K73 sumoylation site in p27kip1, which is critical to RanBP2 and crm-1 in p27kip1 nuclear-cytoplasmic translocation process.ConclusionTaken together, targeted inhibition of sumoylation of p27kip1 may serve as a potentially potent therapeutic target in the eradication of cholangiocarcinoma development and relapses.Electronic supplementary materialThe online version of this article (doi:10.1186/s12867-017-0100-5) contains supplementary material, which is available to authorized users.

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SUMOylation regulates p27^Kip1stability and localization in response to TGFβ

Cited by 10 publications

References 57 publications

p27kip1: An all-round tumor suppressor

p27kip1: An all-round tumor suppressor

CDK6 protects epithelial ovarian cancer from platinum‐induced death via FOXO3 regulation

Sumoylation in p27kip1 via RanBP2 promotes cancer cell growth in cholangiocarcinoma cell line QBC939

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SUMOylation regulates p27Kip1stability and localization in response to TGFβ

Cited by 10 publications

References 57 publications

p27kip1: An all-round tumor suppressor

p27kip1: An all-round tumor suppressor

CDK6 protects epithelial ovarian cancer from platinum‐induced death via FOXO3 regulation

Sumoylation in p27kip1 via RanBP2 promotes cancer cell growth in cholangiocarcinoma cell line QBC939

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SUMOylation regulates p27^Kip1stability and localization in response to TGFβ