Conjugation of the ubiquitin-like (Ubl) protein NEDD8 (neural precursor cell expressed developmentally downregulated protein 8) to target proteins, or neddylation, is a fundamental biological process that controls many key cellular functions, including cell cycle progression, the DNA damage response, and apoptosis. Like ubiquitylation, neddylation is catalyzed through an enzymatic cascade consisting of an E1-activating enzyme complex (NAE1/APPBP1-UBA3), two E2 enzymes (Ubc12/Ube2M and Ube2F) and several E3 enzymes such as Really Interesting New Gene (RING)-box protein 1 (Rbx1) and its close homologue Rbx2 (reviewed in [Soucy et al., 2010; Enchev et al., 2014; Brown and Jackson, 2015]). Additional proteins called DCUN1D1-5 (defective in cullin neddylation, domain containing 1–5) have been described in human cells as cofactors for RBX1 and RBX2, which potentiate neddylation activity (Kim et al., 2008; Kurz et al., 2008; Monda et al., 2013). The most well-studied target for NEDD8 conjugation are the cullins (CUL1,−2,–3,−4A,−4B,−5, and −7), which are the scaffold subunits of the largest family of ubiquitin E3 ligases, the cullin-RING ligases (CRLs). NEDD8 transfer from the E2 to a lysine (Lys) residue within the cullin promotes conformational changes that increase the catalytic activity of the CRL, while also blocking the binding of the CRL exchange factor CAND1 (Duda et al., 2008; Saha and Deshaies, 2008; Pierce et al., 2013). Structural changes induced by NEDD8 conjugation to cullin subunits ultimately contribute to the efficient ubiquitylation of downstream CRL substrates and their degradation by the 26S proteasome.

By inducing CRL activation and assembly in this way, NEDD8 ligation to cullins controls a high proportion of ubiquitylation events in cells (~10–20%) (Soucy et al., 2009), making this pathway an attractive target for pharmacological manipulation of protein turnover or proteostasis. An inhibitor of the NEDD8 E1 enzyme, MLN4924, has been recently developed and shown to potently and rapidly suppress neddylation in cells, thereby inhibiting CRL activity. Significantly, preclinical studies using MLN4924 have demonstrated its antitumor activity in several human tumor xenografts and hematological malignancies (Soucy et al., 2009, 2010; Brownell et al., 2010). While the antitumorigenic activity of MLN4924 has been largely attributed to CRL substrates, it is unknown whether non-cullin targets of neddylation could also play important roles in tumorigenesis.

Although cullins are the primary substrates for NEDD8 conjugation, more recently several non-cullin neddylation targets have been described. Among this list of proteins are p53 (Xirodimas et al., 2004), E2F1 (Loftus et al., 2012; Aoki et al., 2013), ribosomal protein L11 (Sundqvist et al., 2009), Smurf1 (Xie et al., 2014) and Histone H4 (Ma et al., 2013b). In addition, large-scale proteomic surveys have uncovered several other neddylation substrates both within and outside the CRL pathway (Jones et al., 2008; Xirodimas et al., 2008). However, some of these non-cullin substrates remain to be fully validated as true NEDD8 targets, as these studies relied on overexpression of epitope-tagged NEDD8 constructs as bait. Since NEDD8 conjugation can also occur through the ubiquitylation machinery under these conditions (Hjerpe et al., 2012), it can be difficult to assign bona fide neddylation substrates in such experiments. Another challenge in the identification of non-cullin neddylation targets is the relatively low abundance and transient nature of NEDD8 modification events in cells, limiting neddylation detection at an endogenous level by proteomic approaches.

Like other protein post-translational modifications (PTMs), neddylation is reversible. COP9 signalosome complex subunit 5 (CSN5), a metallo-protease and component of the eight-subunit COP9 signalosome complex (CSN), is the major cullin deneddylase in human cells (Lyapina et al., 2001; Cope, 2002). CSN is specific for neddylated cullins (Lingaraju et al., 2014; Cavadini et al., 2016); however, deneddylase(s) controlling non-cullin neddylated substrates have been poorly defined (Figure 1A). Recently, a cysteine protease called SENP8 (also known as DEN1 or NEDP1) has been characterized that functions distinctly from CSN in deneddylating primarily non-cullin substrates (Chan et al., 2008; Mergner et al., 2015) as well as hyper-neddylated cullins (Mendoza et al., 2003; Wu et al., 2003). SENP8 selectively interacts with NEDD8 and not ubiquitin (Gan-Erdene et al., 2003; Shen et al., 2005), and also plays a redundant role in proteolytic processing of the precursor form of NEDD8 in conjunction with ubiquitin C-terminal hydrolase isozyme 3 (UCHL3) (Wada et al., 1998; Wu et al., 2003). Criteria defining the unique substrate preferences of CSN and SENP8 are still not clear; however, a previous study showed distinct neddylation defects in DEN1^null versus CSN5^null Drosophila larvae, suggesting that the two enzymes have non-overlapping functions (Chan et al., 2008). Moreover, the specific substrates for NEDD8 deconjugation by SENP8, as well as the phenotypic consequences of long-term SENP8 depletion, have not been thoroughly profiled in mammalian cells.

Figure 1

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Figure 1—source data 1

NEDD8- modified peptides identified by MS analysis of FLAG-NEDD8 IP samples.

https://doi.org/10.7554/eLife.24325.003

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To capture and enrich for potentially low-level non-cullin neddylated substrates, we employed a strategy to stabilize and trap the ‘neddylated’ state of proteins by using an ectopically expressed, deconjugation-resistant mutant of NEDD8, as previously performed using uncleavable ubiquitin and SUMO modifiers (Békés et al., 2011, 2013). The deconjugation-resistant form of NEDD8 was expressed in cells using a doxycycline (Dox)-inducible expression system to minimize the potential of overexpression artifacts, followed by immuno-affinity purification and identification of neddylated substrates by mass spectrometry (MS). Strikingly, we observed enrichment of not only neddylated cullins using this technique but also several novel non-cullin targets, including the NEDD8 E2-conjugating enzyme, Ubc12. In a series of experiments, we confirmed that Ubc12 becomes auto-neddylated both in vitro and in cells, and that Ubc12 and other NEDD8 pathway components form aberrant NEDD8-conjugates in the absence of deneddylating activity. Moreover, we identified SENP8, but not CSN, as the deconjugating enzyme responsible for reversing aberrant neddylation of non-cullin substrates. Importantly, SENP8 loss-of-function studies showed that SENP8 is required for proper CRL activation and G1/S phase cell cycle progression. Finally, to identify candidate substrates with altered ubiquitylation status that could contribute to this defective G1/S regulation, we performed an unbiased proteomics screen to compare the relative abundance of ubiquitylated peptides in parental versus SENP8 knockout cells. Using this approach, we uncovered several potentially relevant substrates with reduced ubiquitylation and proteasome-mediated degradation in SENP8-deficient cells that have important roles in cell cycle regulation. Therefore, our findings suggest a novel role for SENP8 in the quality control of the NEDD8 conjugation pathway through prevention of aberrant neddylation of pathway components and demonstrate the functional consequences of perturbing this regulation on CRL activation and cell growth control.

In this study, we developed a novel strategy to identify new non-cullin neddylation targets, adding to our current understanding of cellular roles for NEDD8 conjugation. Using a deconjugation-resistant FLAG-NEDD8-L73P mutant, we were able to isolate several NEDD8-conjugated proteins in cells for identification by MS (Figure 1), several of which were also found in previous analyses using ectopically-expressed NEDD8 (Jones et al., 2008; Xirodimas et al., 2008). Our system provides an advantage over previous proteomic approaches in that expression of even relatively low levels of the FLAG-NEDD8-L73P is sufficient to enable stabilization and IP of neddylated proteins for MS analysis, thus avoiding potential artificial effects of NEDD8 overexpression (Xirodimas et al., 2008; Kim et al., 2011; Hjerpe et al., 2012). This feature aided our detection of several NEDD8-conjugates unique to this study, including MCM7, DDB1, Rab14, FEM1C, RNF187 and others (see Figure 1—source data 1 for complete list). Future work will be needed to further validate these as true neddylation substrates and functionally characterize these newly identified NEDD-conjugates in cells. Nevertheless, our MS dataset provides a valuable resource to expand upon previous studies of the NEDD8-modified proteome. Recently, several criteria for validation of bona fide neddylation substrates have been delineated (Enchev et al., 2014). In light of our current findings, we propose that identifying the physiological deneddylase could be added to these criteria to show potential dynamic and reversible neddylation of target substrates.

We identified and characterized the NEDD8 E2 conjugating enzyme Ubc12 as a novel substrate for auto-neddylation both in vitro and in cells (Figure 2). Through a combination of both MS-based analysis and Ubc12 Lys site mutagenesis, we identified the site of Ubc12 NEDD8 conjugation at its extreme N-terminus (K3). This modification is distinct from the site of Ubc12~NEDD8 thioester intermediate formation, occurring when NEDD8 is transferred from the active site of the E1 to the E2 active site Cys residue during the NEDD8 conjugation reaction (Huang et al., 2007). We have also shown that Ubc12 neddylation is sensitive to MLN4924 treatment and is dependent on the catalytic Cys residue of Ubc12. NEDD8 modification at the N-terminus of Ubc12 contributes to reduced DCN1 binding and consequently altered CRL activities, although the mechanism by which this occurs is still unclear. One possibility is that the NEDD8 modification could directly impede the initial interaction between the N-terminally acetylated Ubc12 and the hydrophobic binding pocket of DCN1 (Scott et al., 2011). Alternatively, NEDD8 modification could occur on pre-formed Dcn1-Ubc12 complex to induce a conformational change, leading to dissolution of the complex. Further in vitro analyses using purified components will be required to discriminate between these possibilities. Unexpectedly, from our SILAC-MS analysis (Figure 4), we identified additional components within the NEDD8 pathway that can form NEDD8-conjugates, including the E1 subunits UBA3 (UBE1C) and NAE1 (APPBP1), as well as the DCNLs DCUN1D1 and DCUN1D5. Future structural and biochemical work will be necessary to better understand the functional roles of these individual NEDD8 modifications in regulating the efficiency of NEDD8 conjugation to cullin and non-cullin substrates.

Furthermore, we identified the NEDD8-specific cysteine protease, SENP8, as the deneddylase that deconjugates NEDD8 from Ubc12 and other NEDD8 pathway components in human cells. Generation of CRISPR-derived knockout cell lines allowed us to profile effects of SENP8 depletion on global neddylation changes through MS-based proteomics (Figure 4) and immunoblot analysis of NEDD8 conjugates (Figure 5). These analyses revealed that SENP8 deneddylating activity is important to inhibit the aberrant neddylation of Ubc12 and other NEDD8 pathway components in cells (Figures 4 and 5). We also uncovered several previously uncharacterized non-cullin neddylation substrates in the SENP8 knockout cells (see Figure 4—source data 1 for complete list), complementing our MS analyses using L73P-NEDD8 in Figure 1F.

Several adverse phenotypic consequences occur in SENP8 knockout cells, including decreased efficiency of NEDD8 conjugation to cullins (primarily CUL1 and CUL5), decreased CRL activity, and increased stabilization of CRL substrates, summarized in Figure 8. Accompanying these deficiencies, SENP8 knockout cells also showed aberrant G1/S cell cycle progression and accelerated cell growth (Figure 6). Interestingly, these cell cycle alterations partially phenocopy previously reported defects associated with the loss of either Cul1 (Dealy et al., 1999; Wang et al., 1999; Chen and Li, 2010) or Cul5 function (Burnatowska-Hledin et al., 2001; Buchwalter et al., 2008; Bradley et al., 2010; Ma et al., 2013a; Willis et al., 2017). Through an unbiased proteomic screen in which we enriched for K-ε-GG remnant-containing peptides in MG132-treated cells, we were able to identify several candidate substrates with deregulated ubiquitylation status and stability in the SENP8 knockout cells that could potentially contribute to G1/S progression defects. One prominent example is the SCF^βTrCP substrate Cdc25A. Cdc25A acts as a major regulator of both the G1/S and G2/M cell cycle transitions by dephosphorylating and activating Cyclin E/Cdk2 and Cyclin B/Cdk1 complexes, respectively (Strausfeld et al., 1991; Blomberg and Hoffmann, 1999). Several reports have observed shortening of G1 phase and increased cyclin E- and cyclin A-associated kinase activity following overexpression of Cdc25A (Blomberg and Hoffmann, 1999; Sexl et al., 1999). Therefore, the increased total levels (Figure 7E) and impaired degradation of Cdc25A during interphase (Figure 7F) observed in SENP8 knockout cells could potentially play a significant role in promoting the unscheduled dephosphorylation of CDK complexes and accelerated cell cycle progression phenotypes.

Figure 8

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Other SCF^βTrCP substrates with differential ubiquitylation status in SENP8 knockout cells identified through our SILAC-MS screen (Figure 7B) were β-catenin and UHRF1. Aberrant ubiquitylation and degradation of β-catenin in SENP8-deficient cells may have a significant influence on cell growth by affecting transcription of downstream Wnt target genes such as c-Myc and Cyclin D1 (He et al., 1998; Clevers and Nusse, 2012). UHRF1 (ICBP90), an E2F1 target, also plays a critical role in controlling the G1/S transition by methylating and suppressing certain tumor suppressor genes (Unoki et al., 2004; Jeanblanc et al., 2005; Alhosin et al., 2011; Taylor et al., 2013; Mudbhary et al., 2014). Interestingly, UHRF1 has been shown to bind to the methylated promoter region of the p14^ARF tumor suppressor (Unoki et al., 2004), which we also found was significantly downregulated in SENP8-deficient cells (Figure 7E and F). Whether the elevated UHRF1 levels in SENP8 knockout cells account for this reduced expression of p14^ARF is unclear. However, deregulated p14^ARF levels may have a significant impact on the accelerated G1/S progression in SENP8 knockout cells, as p14^ARF plays a critical role as a tumor suppressor protein by blocking mouse double minute (MDM2)-induced degradation of p53 (Stott et al., 1998), and inhibiting E2F transcriptional activity (Eymin et al., 2001). The combined effects of p14^ARF levels, deregulated CRL substrates identified in our SILAC-MS screen (Figure 7B), and undiscovered substrates (including unknown CUL5 targets, which are largely uncharacterized) are likely to contribute to the deregulated cell cycle progression observed in SENP8-deficient cells.

It is also apparent from our studies that the decreased CRL function accompanying SENP8 loss does not affect all CRL substrates uniformly and is likely to have substrate-specific effects. For example, SENP8 depletion does not entirely phenocopy the effects of complete inhibition of CRL activity following MLN4924 treatment, as we did not see a significant increase in cells with >4C DNA content (indicative of cells undergoing multiple rounds of replication without cell division, or re-replication) or induction of a DNA damage response in SENP8 knockout cells (data not shown). Similarly, using a CSN5-specific inhibitor developed by Novartis, it was recently shown that CSN inhibition does not have pleiotropic effects on all CRL substrates as MLN4924 treatment does (Schlierf et al., 2016), but results in substrate-specific stabilization. This effect was shown to be dependent on the auto-ubiquitylation of unique substrate recognition modules of CRLs following sustained neddylation and activation of CRL complexes. It is likely that SENP8 depletion has similar substrate-specific effects related to the dynamics of CRL activation and substrate-adapter auto-ubiquitylation influenced via aberrant neddylation of Ubc12.

An outstanding question from our study is concerning the biological role for auto-neddylation of NEDD8 pathway components: Is auto-neddylation simply a consequence of prolonged SENP8 depletion, or does it also occur in physiological conditions as a feedback mechanism regulating overstimulation of the NEDD8 pathway? Several recent studies have observed a correlation between overactivation of NEDD8 pathway components, including NEDD8, UBA3, NAE1, and Ubc12, with cancer progression and poor prognosis in human lung cancers (Li et al., 2014), colorectal cancers (Xie et al., 2014), and glioblastomas (Hua et al., 2015). Thus, auto-neddylation within the NEDD8 pathway may represent a deliberate quality control mechanism to limit adverse consequences associated with elevated NEDD8 conjugation and CRL activity. Consistent with this notion, we observed in our own study that NEDD8 conjugation to Ubc12 and other components occurs at defined Lys residues, which are evolutionarily conserved across species. If auto-neddylation of NEDD8 pathway components does in fact occur upon hyper-activation of the neddylation machinery in vivo, this also raises the question of how SENP8 deneddylating activity is regulated under such circumstances. Furthermore, what defines the unique substrate preferences of SENP8 versus those of CSN, the other major deneddylating enzyme in human cells? These issues and others will require further interrogation of SENP8 regulation to resolve. Nevertheless, our study characterizes the SENP8 protease as a key regulator of the NEDD8 pathway, implicating SENP8 as a potential pharmacological target to control CRL-mediated processes in cells.

1. 1. Coleman KE
   2. Bekes M
   3. Chapman JR
   4. Crist SB
   5. Jones MJ
   6. Ueberheide BM
   7. Huang TT

   (2017) SENP8 limits aberrant neddylation of NEDD8 pathway components to promote cullin-RING ubiquitin ligase function

   Publicly available at Proteome Xchange (accession no. PXD006288; MassIVE ID MSV000080917).

   http://proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD006288

1. 1. Abbas T
   2. Sivaprasad U
   3. Terai K
   4. Amador V
   5. Pagano M
   6. Dutta A

   (2008) PCNA-dependent regulation of p21 ubiquitylation and degradation via the CRL4Cdt2 ubiquitin ligase complex

   Genes and Development 22:2496–2506.

   https://doi.org/10.1101/gad.1676108

   • PubMed
   • Google Scholar
2. 1. Abbas T
   2. Shibata E
   3. Park J
   4. Jha S
   5. Karnani N
   6. Dutta A

   (2010) CRL4(Cdt2) regulates cell proliferation and histone gene expression by targeting PR-Set7/Set8 for degradation

   Molecular Cell 40:9–21.

   https://doi.org/10.1016/j.molcel.2010.09.014

   • PubMed
   • Google Scholar
3. 1. Alhosin M
   2. Sharif T
   3. Mousli M
   4. Etienne-Selloum N
   5. Fuhrmann G
   6. Schini-Kerth VB
   7. Bronner C

   (2011) Down-regulation of UHRF1, associated with re-expression of tumor suppressor genes, is a common feature of natural compounds exhibiting anti-cancer properties

   Journal of Experimental and Clinical Cancer Research 30:41.

   https://doi.org/10.1186/1756-9966-30-41

   • PubMed
   • Google Scholar
4. 1. Aoki I
   2. Higuchi M
   3. Gotoh Y

   (2013) NEDDylation controls the target specificity of E2F1 and apoptosis induction

   Oncogene 32:3954–3964.

   https://doi.org/10.1038/onc.2012.428

   • PubMed
   • Google Scholar
5. 1. Blomberg I
   2. Hoffmann I

   (1999) Ectopic expression of Cdc25A accelerates the G(1)/S transition and leads to premature activation of cyclin E- and cyclin A-dependent kinases

   Molecular and Cellular Biology 19:6183–6194.

   https://doi.org/10.1128/MCB.19.9.6183

   • PubMed
   • Google Scholar
6. 1. Bornstein G
   2. Bloom J
   3. Sitry-Shevah D
   4. Nakayama K
   5. Pagano M
   6. Hershko A

   (2003) Role of the SCFSkp2 ubiquitin ligase in the degradation of p21Cip1 in S phase

   Journal of Biological Chemistry 278:25752–25757.

   https://doi.org/10.1074/jbc.M301774200

   • PubMed
   • Google Scholar
7. 1. Bradley SE
   2. Johnson AE
   3. Le IP
   4. Oosterhouse E
   5. Hledin MP
   6. Marquez GA
   7. Burnatowska-Hledin M

   (2010) Phosphorylation of VACM-1/Cul5 by protein kinase A regulates its neddylation and antiproliferative effect

   Journal of Biological Chemistry 285:4883–4895.

   https://doi.org/10.1074/jbc.M109.085225

   • PubMed
   • Google Scholar
8. 1. Brown JS
   2. Jackson SP

   (2015) Ubiquitylation, neddylation and the DNA damage response

   Open Biology 5:150018.

   https://doi.org/10.1098/rsob.150018

   • PubMed
   • Google Scholar
9. 1. Brownell JE
   2. Sintchak MD
   3. Gavin JM
   4. Liao H
   5. Bruzzese FJ
   6. Bump NJ
   7. Soucy TA
   8. Milhollen MA
   9. Yang X
   10. Burkhardt AL
   11. Ma J
   12. Loke HK
   13. Lingaraj T
   14. Wu D
   15. Hamman KB
   16. Spelman JJ
   17. Cullis CA
   18. Langston SP
   19. Vyskocil S
   20. Sells TB
   21. Mallender WD
   22. Visiers I
   23. Li P
   24. Claiborne CF
   25. Rolfe M
   26. Bolen JB
   27. Dick LR

   (2010) Substrate-assisted inhibition of ubiquitin-like protein-activating enzymes: the NEDD8 E1 inhibitor MLN4924 forms a NEDD8-AMP mimetic in situ

   Molecular Cell 37:102–111.

   https://doi.org/10.1016/j.molcel.2009.12.024

   • PubMed
   • Google Scholar
10. 1. Buchwalter A
    2. Van Dort C
    3. Schultz S
    4. Smith R
    5. Le IP
    6. Abbott JL
    7. Oosterhouse E
    8. Johnson AE
    9. Hansen-Smith F
    10. Burnatowska-Hledin M

    (2008) Expression of VACM-1/cul5 mutant in endothelial cells induces MAPK phosphorylation and maspin degradation and converts cells to the angiogenic phenotype

    Microvascular Research 75:155–168.

    https://doi.org/10.1016/j.mvr.2007.08.004

    • PubMed
    • Google Scholar
11. 1. Burnatowska-Hledin
    2. Zeneberg A
    3. Roulo A
    4. Grobe J
    5. Zhao P
    6. Lelkes PI
    7. Clare P
    8. Barney C

    (2001) Expression of VACM-1 protein in cultured rat adrenal endothelial cells is linked to the cell cycle

    Endothelium 8:49–63.

    https://doi.org/10.3109/10623320109063157

    • PubMed
    • Google Scholar
12. 1. Busino L
    2. Donzelli M
    3. Chiesa M
    4. Guardavaccaro D
    5. Ganoth D
    6. Dorrello NV
    7. Hershko A
    8. Pagano M
    9. Draetta GF

    (2003) Degradation of Cdc25A by beta-TrCP during S phase and in response to DNA damage

    Nature 426:87–91.

    https://doi.org/10.1038/nature02082

    • PubMed
    • Google Scholar
13. 1. Békés M
    2. Prudden J
    3. Srikumar T
    4. Raught B
    5. Boddy MN
    6. Salvesen GS

    (2011) The dynamics and mechanism of SUMO chain deconjugation by SUMO-specific proteases

    Journal of Biological Chemistry 286:10238–10247.

    https://doi.org/10.1074/jbc.M110.205153

    • PubMed
    • Google Scholar
14. 1. Békés M
    2. Okamoto K
    3. Crist SB
    4. Jones MJ
    5. Chapman JR
    6. Brasher BB
    7. Melandri FD
    8. Ueberheide BM
    9. Denchi EL
    10. Huang TT

    (2013) DUB-resistant ubiquitin to survey ubiquitination switches in mammalian cells

    Cell Reports 5:826–838.

    https://doi.org/10.1016/j.celrep.2013.10.008

    • PubMed
    • Google Scholar
15. 1. Cavadini S
    2. Fischer ES
    3. Bunker RD
    4. Potenza A
    5. Lingaraju GM
    6. Goldie KN
    7. Mohamed WI
    8. Faty M
    9. Petzold G
    10. Beckwith RE
    11. Tichkule RB
    12. Hassiepen U
    13. Abdulrahman W
    14. Pantelic RS
    15. Matsumoto S
    16. Sugasawa K
    17. Stahlberg H
    18. Thomä NH

    (2016) Cullin-RING ubiquitin E3 ligase regulation by the COP9 signalosome

    Nature 531:598–603.

    https://doi.org/10.1038/nature17416

    • PubMed
    • Google Scholar
16. 1. Centore RC
    2. Havens CG
    3. Manning AL
    4. Li JM
    5. Flynn RL
    6. Tse A
    7. Jin J
    8. Dyson NJ
    9. Walter JC
    10. Zou L

    (2010) CRL4(Cdt2)-mediated destruction of the histone methyltransferase Set8 prevents premature chromatin compaction in S phase

    Molecular Cell 40:22–33.

    https://doi.org/10.1016/j.molcel.2010.09.015

    • PubMed
    • Google Scholar
17. 1. Chan Y
    2. Yoon J
    3. Wu JT
    4. Kim HJ
    5. Pan KT
    6. Yim J
    7. Chien CT

    (2008) DEN1 deneddylates non-cullin proteins in vivo

    Journal of Cell Science 121:3218–3223.

    https://doi.org/10.1242/jcs.030445

    • PubMed
    • Google Scholar
18. 1. Chen G
    2. Li G

    (2010) Increased Cul1 expression promotes melanoma cell proliferation through regulating p27 expression

    International Journal of Oncology 37:1339–1344.

    https://doi.org/10.3892/ijo_00000786

    • PubMed
    • Google Scholar
19. 1. Chen H
    2. Ma H
    3. Inuzuka H
    4. Diao J
    5. Lan F
    6. Shi YG
    7. Wei W
    8. Shi Y

    (2013) DNA damage regulates UHRF1 stability via the SCF(β-TrCP) E3 ligase

    Molecular and Cellular Biology 33:1139–1148.

    https://doi.org/10.1128/MCB.01191-12

    • PubMed
    • Google Scholar
20. 1. Clevers H
    2. Nusse R

    (2012) Wnt/β-catenin signaling and disease

    Cell 149:1192–1205.

    https://doi.org/10.1016/j.cell.2012.05.012

    • PubMed
    • Google Scholar
21. 1. Cope GA

    (2002) Role of predicted metalloprotease motif of Jab1/Csn5 in cleavage of Nedd8 from Cul1

    Science 298:608–611.

    https://doi.org/10.1126/science.1075901

    • Google Scholar
22. 1. Cotto-Rios XM
    2. Békés M
    3. Chapman J
    4. Ueberheide B
    5. Huang TT

    (2012) Deubiquitinases as a signaling target of oxidative stress

    Cell Reports 2:1475–1484.

    https://doi.org/10.1016/j.celrep.2012.11.011

    • PubMed
    • Google Scholar
23. 1. Cox J
    2. Mann M

    (2008) MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification

    Nature Biotechnology 26:1367–1372.

    https://doi.org/10.1038/nbt.1511

    • PubMed
    • Google Scholar
24. 1. Dealy MJ
    2. Nguyen KV
    3. Lo J
    4. Gstaiger M
    5. Krek W
    6. Elson D
    7. Arbeit J
    8. Kipreos ET
    9. Johnson RS

    (1999) Loss of Cul1 results in early embryonic lethality and dysregulation of cyclin E

    Nature Genetics 23:245–248.

    https://doi.org/10.1038/13886

    • PubMed
    • Google Scholar
25. 1. Donzelli M
    2. Squatrito M
    3. Ganoth D
    4. Hershko A
    5. Pagano M
    6. Draetta GF

    (2002) Dual mode of degradation of Cdc25 A phosphatase

    The EMBO Journal 21:4875–4884.

    https://doi.org/10.1093/emboj/cdf491

    • PubMed
    • Google Scholar
26. 1. Duda DM
    2. Borg LA
    3. Scott DC
    4. Hunt HW
    5. Hammel M
    6. Schulman BA

    (2008) Structural insights into NEDD8 activation of cullin-RING ligases: conformational control of conjugation

    Cell 134:995–1006.

    https://doi.org/10.1016/j.cell.2008.07.022

    • PubMed
    • Google Scholar
27. 1. Emanuele MJ
    2. Elia AE
    3. Xu Q
    4. Thoma CR
    5. Izhar L
    6. Leng Y
    7. Guo A
    8. Chen YN
    9. Rush J
    10. Hsu PW
    11. Yen HC
    12. Elledge SJ

    (2011) Global identification of modular cullin-RING ligase substrates

    Cell 147:459–474.

    https://doi.org/10.1016/j.cell.2011.09.019

    • PubMed
    • Google Scholar
28. 1. Enchev RI
    2. Schulman BA
    3. Peter M

    (2014) Protein neddylation: beyond cullin–RING ligases

    Nature Reviews Molecular Cell Biology 16:30–44.

    https://doi.org/10.1038/nrm3919

    • Google Scholar
29. 1. Eymin B
    2. Karayan L
    3. Séité P
    4. Brambilla C
    5. Brambilla E
    6. Larsen CJ
    7. Gazzéri S

    (2001) Human ARF binds E2F1 and inhibits its transcriptional activity

    Oncogene 20:1033–1041.

    https://doi.org/10.1038/sj.onc.1204220

    • PubMed
    • Google Scholar
30. 1. Gan-Erdene T
    2. Nagamalleswari K
    3. Yin L
    4. Wu K
    5. Pan ZQ
    6. Wilkinson KD

    (2003) Identification and characterization of DEN1, a deneddylase of the ULP family

    Journal of Biological Chemistry 278:28892–28900.

    https://doi.org/10.1074/jbc.M302890200

    • PubMed
    • Google Scholar
31. 1. Hao B
    2. Oehlmann S
    3. Sowa ME
    4. Harper JW
    5. Pavletich NP

    (2007) Structure of a Fbw7-Skp1-cyclin E complex: multisite-phosphorylated substrate recognition by SCF ubiquitin ligases

    Molecular Cell 26:131–143.

    https://doi.org/10.1016/j.molcel.2007.02.022

    • PubMed
    • Google Scholar
32. 1. He TC
    2. Sparks AB
    3. Rago C
    4. Hermeking H
    5. Zawel L
    6. da Costa LT
    7. Morin PJ
    8. Vogelstein B
    9. Kinzler KW

    (1998) Identification of c-MYC as a target of the APC pathway

    Science 281:1509–1512.

    https://doi.org/10.1126/science.281.5382.1509

    • PubMed
    • Google Scholar
33. 1. Hjerpe R
    2. Thomas Y
    3. Chen J
    4. Zemla A
    5. Curran S
    6. Shpiro N
    7. Dick LR
    8. Kurz T

    (2012) Changes in the ratio of free NEDD8 to ubiquitin triggers NEDDylation by ubiquitin enzymes

    Biochemical Journal 441:927–939.

    https://doi.org/10.1042/BJ20111671

    • PubMed
    • Google Scholar
34. 1. Hua W
    2. Li C
    3. Yang Z
    4. Li L
    5. Jiang Y
    6. Yu G
    7. Zhu W
    8. Liu Z
    9. Duan S
    10. Chu Y
    11. Yang M
    12. Zhang Y
    13. Mao Y
    14. Jia L

    (2015) Suppression of glioblastoma by targeting the overactivated protein neddylation pathway

    Neuro-Oncology 17:1333–1343.

    https://doi.org/10.1093/neuonc/nov066

    • PubMed
    • Google Scholar
35. 1. Huang DT
    2. Paydar A
    3. Zhuang M
    4. Waddell MB
    5. Holton JM
    6. Schulman BA

    (2005) Structural basis for recruitment of Ubc12 by an E2 binding domain in NEDD8's E1

    Molecular Cell 17:341–350.

    https://doi.org/10.1016/j.molcel.2004.12.020

    • PubMed
    • Google Scholar
36. 1. Huang DT
    2. Hunt HW
    3. Zhuang M
    4. Ohi MD
    5. Holton JM
    6. Schulman BA

    (2007) Basis for a ubiquitin-like protein thioester switch toggling E1-E2 affinity

    Nature 445:394–398.

    https://doi.org/10.1038/nature05490

    • PubMed
    • Google Scholar
37. 1. Huang DT
    2. Zhuang M
    3. Ayrault O
    4. Schulman BA

    (2008) Identification of conjugation specificity determinants unmasks vestigial preference for ubiquitin within the NEDD8 E2

    Nature Structural and Molecular Biology 15:280–287.

    https://doi.org/10.1038/nsmb.1387

    • PubMed
    • Google Scholar
38. 1. Huang DT
    2. Ayrault O
    3. Hunt HW
    4. Taherbhoy AM
    5. Duda DM
    6. Scott DC
    7. Borg LA
    8. Neale G
    9. Murray PJ
    10. Roussel MF
    11. Schulman BA

    (2009a) E2-RING expansion of the NEDD8 cascade confers specificity to cullin modification

    Molecular Cell 33:483–495.

    https://doi.org/10.1016/j.molcel.2009.01.011

    • PubMed
    • Google Scholar
39. 1. Huang daW
    2. Sherman BT
    3. Lempicki RA

    (2009b) Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources

    Nature Protocols 4:44–57.

    https://doi.org/10.1038/nprot.2008.211

    • PubMed
    • Google Scholar
40. 1. Huang daW
    2. Sherman BT
    3. Lempicki RA

    (2009c) Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists

    Nucleic Acids Research 37:1–13.

    https://doi.org/10.1093/nar/gkn923

    • PubMed
    • Google Scholar
41. 1. Jeanblanc M
    2. Mousli M
    3. Hopfner R
    4. Bathami K
    5. Martinet N
    6. Abbady AQ
    7. Siffert JC
    8. Mathieu E
    9. Muller CD
    10. Bronner C

    (2005) The retinoblastoma gene and its product are targeted by ICBP90: a key mechanism in the G1/S transition during the cell cycle

    Oncogene 24:7337–7345.

    https://doi.org/10.1038/sj.onc.1208878

    • PubMed
    • Google Scholar
42. 1. Jin J
    2. Shirogane T
    3. Xu L
    4. Nalepa G
    5. Qin J
    6. Elledge SJ
    7. Harper JW

    (2003) SCFbeta-TRCP links Chk1 signaling to degradation of the Cdc25A protein phosphatase

    Genes and Development 17:3062–3074.

    https://doi.org/10.1101/gad.1157503

    • PubMed
    • Google Scholar
43. 1. Jin J
    2. Arias EE
    3. Chen J
    4. Harper JW
    5. Walter JC

    (2006) A family of diverse Cul4-Ddb1-interacting proteins includes Cdt2, which is required for S phase destruction of the replication factor Cdt1

    Molecular Cell 23:709–721.

    https://doi.org/10.1016/j.molcel.2006.08.010

    • PubMed
    • Google Scholar
44. 1. Jones J
    2. Wu K
    3. Yang Y
    4. Guerrero C
    5. Nillegoda N
    6. Pan ZQ
    7. Huang L

    (2008) A targeted proteomic analysis of the ubiquitin-like modifier nedd8 and associated proteins

    Journal of Proteome Research 7:1274–1287.

    https://doi.org/10.1021/pr700749v

    • PubMed
    • Google Scholar
45. 1. Kim AY
    2. Bommeljé CC
    3. Lee BE
    4. Yonekawa Y
    5. Choi L
    6. Morris LG
    7. Huang G
    8. Kaufman A
    9. Ryan RJ
    10. Hao B
    11. Ramanathan Y
    12. Singh B

    (2008) SCCRO (DCUN1D1) is an essential component of the E3 complex for neddylation

    Journal of Biological Chemistry 283:33211–33220.

    https://doi.org/10.1074/jbc.M804440200

    • PubMed
    • Google Scholar
46. 1. Kim W
    2. Bennett EJ
    3. Huttlin EL
    4. Guo A
    5. Li J
    6. Possemato A
    7. Sowa ME
    8. Rad R
    9. Rush J
    10. Comb MJ
    11. Harper JW
    12. Gygi SP

    (2011) Systematic and quantitative assessment of the ubiquitin-modified proteome

    Molecular Cell 44:325–340.

    https://doi.org/10.1016/j.molcel.2011.08.025

    • PubMed
    • Google Scholar
47. 1. Koepp DM
    2. Schaefer LK
    3. Ye X
    4. Keyomarsi K
    5. Chu C
    6. Harper JW
    7. Elledge SJ

    (2001) Phosphorylation-dependent ubiquitination of cyclin E by the SCFFbw7 ubiquitin ligase

    Science 294:173–177.

    https://doi.org/10.1126/science.1065203

    • PubMed
    • Google Scholar
48. 1. Kurz T
    2. Chou YC
    3. Willems AR
    4. Meyer-Schaller N
    5. Hecht ML
    6. Tyers M
    7. Peter M
    8. Sicheri F

    (2008) Dcn1 functions as a scaffold-type E3 ligase for Cullin neddylation

    Molecular Cell 29:23–35.

    https://doi.org/10.1016/j.molcel.2007.12.012

    • PubMed
    • Google Scholar
49. 1. Latres E
    2. Chiaur DS
    3. Pagano M

    (1999) The human F box protein beta-Trcp associates with the Cul1/Skp1 complex and regulates the stability of beta-catenin

    Oncogene 18:849–854.

    https://doi.org/10.1038/sj.onc.1202653

    • PubMed
    • Google Scholar
50. 1. Li X
    2. Zhao Q
    3. Liao R
    4. Sun P
    5. Wu X

    (2003) The SCF(Skp2) ubiquitin ligase complex interacts with the human replication licensing factor Cdt1 and regulates Cdt1 degradation

    Journal of Biological Chemistry 278:30854–30858.

    https://doi.org/10.1074/jbc.C300251200

    • PubMed
    • Google Scholar
51. 1. Li L
    2. Wang M
    3. Yu G
    4. Chen P
    5. Li H
    6. Wei D
    7. Zhu J
    8. Xie L
    9. Jia H
    10. Shi J
    11. Li C
    12. Yao W
    13. Wang Y
    14. Gao Q
    15. Jeong LS
    16. Lee HW
    17. Yu J
    18. Hu F
    19. Mei J
    20. Wang P
    21. Chu Y
    22. Qi H
    23. Yang M
    24. Dong Z
    25. Sun Y
    26. Hoffman RM
    27. Jia L

    (2014) Overactivated neddylation pathway as a therapeutic target in lung Cancer

    JNCI Journal of the National Cancer Institute 106:dju083–21.

    https://doi.org/10.1093/jnci/dju083

    • PubMed
    • Google Scholar
52. 1. Lingaraju GM
    2. Bunker RD
    3. Cavadini S
    4. Hess D
    5. Hassiepen U
    6. Renatus M
    7. Fischer ES
    8. Thomä NH

    (2014) Crystal structure of the human COP9 signalosome

    Nature 512:161–165.

    https://doi.org/10.1038/nature13566

    • PubMed
    • Google Scholar
53. 1. Loftus SJ
    2. Liu G
    3. Carr SM
    4. Munro S
    5. La Thangue NB

    (2012) NEDDylation regulates E2F-1-dependent transcription

    EMBO Reports 13:811–818.

    https://doi.org/10.1038/embor.2012.113

    • PubMed
    • Google Scholar
54. 1. Lyapina S
    2. Cope G
    3. Shevchenko A
    4. Serino G
    5. Tsuge T
    6. Zhou C
    7. Wolf DA
    8. Wei N
    9. Shevchenko A
    10. Deshaies RJ

    (2001) Promotion of NEDD-CUL1 conjugate cleavage by COP9 signalosome

    Science 292:1382–1385.

    https://doi.org/10.1126/science.1059780

    • PubMed
    • Google Scholar
55. 1. Ma C
    2. Qi Y
    3. Shao L
    4. Liu M
    5. Li X
    6. Tang H

    (2013a) Downregulation of miR-7 upregulates Cullin 5 (CUL5) to facilitate G1/S transition in human hepatocellular carcinoma cells

    IUBMB Life 65:1026–1034.

    https://doi.org/10.1002/iub.1231

    • PubMed
    • Google Scholar
56. 1. Ma T
    2. Chen Y
    3. Zhang F
    4. Yang CY
    5. Wang S
    6. Yu X

    (2013b) RNF111-dependent neddylation activates DNA damage-induced ubiquitination

    Molecular Cell 49:897–907.

    https://doi.org/10.1016/j.molcel.2013.01.006

    • PubMed
    • Google Scholar
57. 1. Mali P
    2. Yang L
    3. Esvelt KM
    4. Aach J
    5. Guell M
    6. DiCarlo JE
    7. Norville JE
    8. Church GM

    (2013) RNA-guided human genome engineering via Cas9

    Science 339:823–826.

    https://doi.org/10.1126/science.1232033

    • PubMed
    • Google Scholar
58. 1. Mendoza HM
    2. Shen LN
    3. Botting C
    4. Lewis A
    5. Chen J
    6. Ink B
    7. Hay RT

    (2003) NEDP1, a highly conserved cysteine protease that deNEDDylates cullins

    Journal of Biological Chemistry 278:25637–25643.

    https://doi.org/10.1074/jbc.M212948200

    • PubMed
    • Google Scholar
59. 1. Mergner J
    2. Heinzlmeir S
    3. Kuster B
    4. Schwechheimer C

    (2015) DENEDDYLASE1 deconjugates NEDD8 from non-cullin protein substrates in Arabidopsis thaliana

    The Plant Cell 27:741–753.

    https://doi.org/10.1105/tpc.114.135996

    • PubMed
    • Google Scholar
60. 1. Mikolajczyk J
    2. Drag M
    3. Békés M
    4. Cao JT
    5. Ronai Z
    6. Salvesen GS

    (2007) Small ubiquitin-related modifier (SUMO)-specific proteases: profiling the specificities and activities of human SENPs

    Journal of Biological Chemistry 282:26217–26224.

    https://doi.org/10.1074/jbc.M702444200

    • PubMed
    • Google Scholar
61. 1. Monda JK
    2. Scott DC
    3. Miller DJ
    4. Lydeard J
    5. King D
    6. Harper JW
    7. Bennett EJ
    8. Schulman BA

    (2013) Structural conservation of distinctive N-terminal acetylation-dependent interactions across a family of mammalian NEDD8 ligation enzymes

    Structure 21:42–53.

    https://doi.org/10.1016/j.str.2012.10.013

    • PubMed
    • Google Scholar
62. 1. Mudbhary R
    2. Hoshida Y
    3. Chernyavskaya Y
    4. Jacob V
    5. Villanueva A
    6. Fiel MI
    7. Chen X
    8. Kojima K
    9. Thung S
    10. Bronson RT
    11. Lachenmayer A
    12. Revill K
    13. Alsinet C
    14. Sachidanandam R
    15. Desai A
    16. SenBanerjee S
    17. Ukomadu C
    18. Llovet JM
    19. Sadler KC

    (2014) UHRF1 overexpression drives DNA hypomethylation and hepatocellular carcinoma

    Cancer Cell 25:196–209.

    https://doi.org/10.1016/j.ccr.2014.01.003

    • PubMed
    • Google Scholar
63. 1. Nishitani H
    2. Sugimoto N
    3. Roukos V
    4. Nakanishi Y
    5. Saijo M
    6. Obuse C
    7. Tsurimoto T
    8. Nakayama KI
    9. Nakayama K
    10. Fujita M
    11. Lygerou Z
    12. Nishimoto T

    (2006) Two E3 ubiquitin ligases, SCF-Skp2 and DDB1-Cul4, target human Cdt1 for proteolysis

    The EMBO Journal 25:1126–1136.

    https://doi.org/10.1038/sj.emboj.7601002

    • PubMed
    • Google Scholar
64. 1. Ong SE
    2. Mann M

    (2006) A practical recipe for stable isotope labeling by amino acids in cell culture (SILAC)

    Nature Protocols 1:2650–2660.

    https://doi.org/10.1038/nprot.2006.427

    • PubMed
    • Google Scholar
65. 1. Pierce NW
    2. Lee JE
    3. Liu X
    4. Sweredoski MJ
    5. Graham RL
    6. Larimore EA
    7. Rome M
    8. Zheng N
    9. Clurman BE
    10. Hess S
    11. Shan SO
    12. Deshaies RJ

    (2013) Cand1 promotes assembly of new SCF complexes through dynamic exchange of F box proteins

    Cell 153:206–215.

    https://doi.org/10.1016/j.cell.2013.02.024

    • PubMed
    • Google Scholar
66. 1. Resnitzky D
    2. Gossen M
    3. Bujard H
    4. Reed SI

    (1994) Acceleration of the G1/S phase transition by expression of cyclins D1 and E with an inducible system

    Molecular and Cellular Biology 14:1669–1679.

    https://doi.org/10.1128/MCB.14.3.1669

    • PubMed
    • Google Scholar
67. 1. Saha A
    2. Deshaies RJ

    (2008) Multimodal activation of the ubiquitin ligase SCF by Nedd8 conjugation

    Molecular Cell 32:21–31.

    https://doi.org/10.1016/j.molcel.2008.08.021

    • PubMed
    • Google Scholar
68. 1. Schlierf A
    2. Altmann E
    3. Quancard J
    4. Jefferson AB
    5. Assenberg R
    6. Renatus M
    7. Jones M
    8. Hassiepen U
    9. Schaefer M
    10. Kiffe M
    11. Weiss A
    12. Wiesmann C
    13. Sedrani R
    14. Eder J
    15. Martoglio B

    (2016) Targeted inhibition of the COP9 signalosome for treatment of Cancer

    Nature Communications 7:13166.

    https://doi.org/10.1038/ncomms13166

    • PubMed
    • Google Scholar
69. 1. Scott DC
    2. Monda JK
    3. Bennett EJ
    4. Harper JW
    5. Schulman BA

    (2011) N-terminal acetylation acts as an avidity enhancer within an interconnected multiprotein complex

    Science 334:674–678.

    https://doi.org/10.1126/science.1209307

    • PubMed
    • Google Scholar
70. 1. Sexl V
    2. Diehl JA
    3. Sherr CJ
    4. Ashmun R
    5. Beach D
    6. Roussel MF

    (1999) A rate limiting function of cdc25A for S phase entry inversely correlates with tyrosine dephosphorylation of Cdk2

    Oncogene 18:573–582.

    https://doi.org/10.1038/sj.onc.1202362

    • PubMed
    • Google Scholar
71. 1. Shen LN
    2. Liu H
    3. Dong C
    4. Xirodimas D
    5. Naismith JH
    6. Hay RT

    (2005) Structural basis of NEDD8 ubiquitin discrimination by the deNEDDylating enzyme NEDP1

    The EMBO Journal 24:1341–1351.

    https://doi.org/10.1038/sj.emboj.7600628

    • PubMed
    • Google Scholar
72. 1. Sigl R
    2. Wandke C
    3. Rauch V
    4. Kirk J
    5. Hunt T
    6. Geley S

    (2009) Loss of the mammalian APC/C activator FZR1 shortens G1 and lengthens S phase but has little effect on exit from mitosis

    Journal of Cell Science 122:4208–4217.

    https://doi.org/10.1242/jcs.054197

    • PubMed
    • Google Scholar
73. 1. Soucek T
    2. Pusch O
    3. Hengstschläger-Ottnad E
    4. Adams PD
    5. Hengstschläger M

    (1997) Deregulated expression of E2F-1 induces cyclin A- and E-associated kinase activities independently from cell cycle position

    Oncogene 14:2251–2257.

    https://doi.org/10.1038/sj.onc.1201061

    • PubMed
    • Google Scholar
74. 1. Soucy TA
    2. Smith PG
    3. Milhollen MA
    4. Berger AJ
    5. Gavin JM
    6. Adhikari S
    7. Brownell JE
    8. Burke KE
    9. Cardin DP
    10. Critchley S
    11. Cullis CA
    12. Doucette A
    13. Garnsey JJ
    14. Gaulin JL
    15. Gershman RE
    16. Lublinsky AR
    17. McDonald A
    18. Mizutani H
    19. Narayanan U
    20. Olhava EJ
    21. Peluso S
    22. Rezaei M
    23. Sintchak MD
    24. Talreja T
    25. Thomas MP
    26. Traore T
    27. Vyskocil S
    28. Weatherhead GS
    29. Yu J
    30. Zhang J
    31. Dick LR
    32. Claiborne CF
    33. Rolfe M
    34. Bolen JB
    35. Langston SP

    (2009) An inhibitor of NEDD8-activating enzyme as a new approach to treat Cancer

    Nature 458:732–736.

    https://doi.org/10.1038/nature07884

    • PubMed
    • Google Scholar
75. 1. Soucy TA
    2. Dick LR
    3. Smith PG
    4. Milhollen MA
    5. Brownell JE

    (2010) The NEDD8 conjugation pathway and its relevance in Cancer biology and therapy

    Genes and Cancer 1:708–716.

    https://doi.org/10.1177/1947601910382898

    • PubMed
    • Google Scholar
76. 1. Stott FJ
    2. Bates S
    3. James MC
    4. McConnell BB
    5. Starborg M
    6. Brookes S
    7. Palmero I
    8. Ryan K
    9. Hara E
    10. Vousden KH
    11. Peters G

    (1998) The alternative product from the human CDKN2A locus, p14(ARF), participates in a regulatory feedback loop with p53 and MDM2

    The EMBO Journal 17:5001–5014.

    https://doi.org/10.1093/emboj/17.17.5001

    • PubMed
    • Google Scholar
77. 1. Strausfeld U
    2. Labbé JC
    3. Fesquet D
    4. Cavadore JC
    5. Picard A
    6. Sadhu K
    7. Russell P
    8. Dorée M

    (1991) Dephosphorylation and activation of a p34cdc2/cyclin B complex in vitro by human CDC25 protein

    Nature 351:242–245.

    https://doi.org/10.1038/351242a0

    • PubMed
    • Google Scholar
78. 1. Sundqvist A
    2. Liu G
    3. Mirsaliotis A
    4. Xirodimas DP

    (2009) Regulation of nucleolar signalling to p53 through NEDDylation of L11

    EMBO Reports 10:1132–1139.

    https://doi.org/10.1038/embor.2009.178

    • PubMed
    • Google Scholar
79. 1. Taylor EM
    2. Bonsu NM
    3. Price RJ
    4. Lindsay HD

    (2013) Depletion of Uhrf1 inhibits chromosomal DNA replication in xenopus egg extracts

    Nucleic Acids Research 41:7725–7737.

    https://doi.org/10.1093/nar/gkt549

    • PubMed
    • Google Scholar
80. 1. Tighe A
    2. Staples O
    3. Taylor S

    (2008) Mps1 kinase activity restrains anaphase during an unperturbed mitosis and targets Mad2 to kinetochores

    The Journal of Cell Biology 181:893–901.

    https://doi.org/10.1083/jcb.200712028

    • PubMed
    • Google Scholar
81. 1. Udeshi ND
    2. Mertins P
    3. Svinkina T

    (2013) Carr, S. a

    Nature Protocols. Nature Publishing Group 8:1950–1960.

    https://doi.org/10.1038/nprot.2013.120

    • Google Scholar
82. 1. Unoki M
    2. Nishidate T
    3. Nakamura Y

    (2004) ICBP90, an E2F-1 target, recruits HDAC1 and binds to methyl-CpG through its SRA domain

    Oncogene 23:7601–7610.

    https://doi.org/10.1038/sj.onc.1208053

    • PubMed
    • Google Scholar
83. 1. Wada H
    2. Kito K
    3. Caskey LS
    4. Yeh ET
    5. Kamitani T

    (1998) Cleavage of the C-terminus of NEDD8 by UCH-L3

    Biochemical and Biophysical Research Communications 251:688–692.

    https://doi.org/10.1006/bbrc.1998.9532

    • PubMed
    • Google Scholar
84. 1. Walker MP
    2. Stopford CM
    3. Cederlund M
    4. Fang F
    5. Jahn C
    6. Rabinowitz AD
    7. Goldfarb D
    8. Graham DM
    9. Yan F
    10. Deal AM
    11. Fedoriw Y
    12. Richards KL
    13. Davis IJ
    14. Weidinger G
    15. Damania B
    16. Major MB

    (2015) FOXP1 potentiates wnt/β-catenin signaling in diffuse large B cell lymphoma

    Science Signaling 8:ra12–12.

    https://doi.org/10.1126/scisignal.2005654

    • PubMed
    • Google Scholar
85. 1. Wang Y
    2. Penfold S
    3. Tang X
    4. Hattori N
    5. Riley P
    6. Harper JW
    7. Cross JC
    8. Tyers M

    (1999) Deletion of the Cul1 gene in mice causes arrest in early embryogenesis and accumulation of cyclin E

    Current Biology 9:1191–S2.

    https://doi.org/10.1016/S0960-9822(00)80024-X

    • PubMed
    • Google Scholar
86. 1. Willis AN
    2. Dean SE
    3. Habbouche JA
    4. Kempers BT
    5. Ludwig ML
    6. Sayfie AD
    7. Lewis SP
    8. Harrier S
    9. DeBruine ZJ
    10. Garrett R
    11. Burnatowska-Hledin MA

    (2017) Nuclear localization signal sequence is required for VACM-1/CUL5-dependent regulation of cellular growth

    Cell and Tissue Research 368:105–114.

    https://doi.org/10.1007/s00441-016-2522-7

    • PubMed
    • Google Scholar
87. 1. Wu K
    2. Yamoah K
    3. Dolios G
    4. Gan-Erdene T
    5. Tan P
    6. Chen A
    7. Lee CG
    8. Wei N
    9. Wilkinson KD
    10. Wang R
    11. Pan ZQ

    (2003) DEN1 is a dual function protease capable of processing the C terminus of Nedd8 and deconjugating hyper-neddylated CUL1

    Journal of Biological Chemistry 278:28882–28891.

    https://doi.org/10.1074/jbc.M302888200

    • PubMed
    • Google Scholar
88. 1. Wu S
    2. Wang W
    3. Kong X
    4. Congdon LM
    5. Yokomori K
    6. Kirschner MW
    7. Rice JC

    (2010) Dynamic regulation of the PR-Set7 histone methyltransferase is required for normal cell cycle progression

    Genes and Development 24:2531–2542.

    https://doi.org/10.1101/gad.1984210

    • PubMed
    • Google Scholar
89. 1. Xie P
    2. Zhang M
    3. He S
    4. Lu K
    5. Chen Y
    6. Xing G
    7. Lu Y
    8. Liu P
    9. Li Y
    10. Wang S
    11. Chai N
    12. Wu J
    13. Deng H
    14. Wang H-R
    15. Cao Y
    16. Zhao F
    17. Cui Y
    18. Wang J
    19. He F
    20. Zhang L

    (2014) The covalent modifier Nedd8 is critical for the activation of Smurf1 ubiquitin ligase in tumorigenesis

    Nature Communications 5:1–16.

    https://doi.org/10.1038/ncomms4733

    • Google Scholar
90. 1. Xirodimas DP
    2. Saville MK
    3. Bourdon JC
    4. Hay RT
    5. Lane DP

    (2004) Mdm2-mediated NEDD8 conjugation of p53 inhibits its transcriptional activity

    Cell 118:83–97.

    https://doi.org/10.1016/j.cell.2004.06.016

    • PubMed
    • Google Scholar
91. 1. Xirodimas DP
    2. Sundqvist A
    3. Nakamura A
    4. Shen L
    5. Botting C
    6. Hay RT

    (2008) Ribosomal proteins are targets for the NEDD8 pathway

    EMBO Reports 9:280–286.

    https://doi.org/10.1038/embor.2008.10

    • PubMed
    • Google Scholar
92. 1. Xu G
    2. Paige JS
    3. Jaffrey SR

    (2010) Global analysis of lysine ubiquitination by ubiquitin remnant immunoaffinity profiling

    Nature Biotechnology 28:868–873.

    https://doi.org/10.1038/nbt.1654

    • PubMed
    • Google Scholar
93. 1. Yen HC
    2. Elledge SJ

    (2008) Identification of SCF ubiquitin ligase substrates by global protein stability profiling

    Science 322:923–929.

    https://doi.org/10.1126/science.1160462

    • PubMed
    • Google Scholar
94. 1. Yuan X
    2. Srividhya J
    3. De Luca T
    4. Lee JH
    5. Pomerening JR

    (2014) Uncovering the role of APC-Cdh1 in generating the dynamics of S-phase onset

    Molecular Biology of the Cell 25:441–456.

    https://doi.org/10.1091/mbc.E13-08-0480

    • PubMed
    • Google Scholar

Author details

1. Kate E Coleman

   Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, United States

   Contribution

   KEC, Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Writing—original draft, Writing—review and editing

   Contributed equally with

   Miklós Békés

   Competing interests

   The authors declare that no competing interests exist.

   "This ORCID iD identifies the author of this article:" 0000-0003-4676-4981

2. Miklós Békés

   Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, United States

   Present address

   Nurix Inc, San Francisco, United States

   Contribution

   MB, Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Writing—review and editing

   Contributed equally with

   Kate E Coleman

   Competing interests

   The authors declare that no competing interests exist.

3. Jessica R Chapman

   Proteomics Laboratory, Division of Advanced Research Technologies, New York University School of Medicine, New York, United States

   Contribution

   JRC, Resources, Data curation, Formal analysis, Methodology, Writing—review and editing

   Competing interests

   The authors declare that no competing interests exist.

4. Sarah B Crist

   Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, United States

   Contribution

   SBC, Data curation, Investigation

   Competing interests

   The authors declare that no competing interests exist.

5. Mathew JK Jones

   Molecular Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, Unites States

   Contribution

   MJKJ, Data curation, Investigation

   Competing interests

   The authors declare that no competing interests exist.

6. Beatrix M Ueberheide

   1. Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, United States
   2. Proteomics Laboratory, Division of Advanced Research Technologies, New York University School of Medicine, New York, United States

   Contribution

   BMU, Formal analysis, Supervision, Funding acquisition, Resources

   Competing interests

   The authors declare that no competing interests exist.

7. Tony T Huang

   Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, United States

   Contribution

   TTH, Formal analysis, Supervision, Funding acquisition, Writing—review and editing

   For correspondence

   tony.huang@nyumc.org

   Competing interests

   The authors declare that no competing interests exist.

   "This ORCID iD identifies the author of this article:" 0000-0001-9291-5002

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