Abstract
Identification of substrates for post-translational modification with ubiquitin and ubiquitin-like molecules is a key in our understanding of the role of this modification in the regulation of biological processes. The ubiquitin-like molecule NEDD8 plays a vital role in cell growth, viability, and development. In this chapter, we describe a method for efficient isolation and detection of NEDDylated proteins in human cells.
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Kamitani T, Kito K, Nguyen HP, Yeh ET (1997) Characterization of NEDD8, a developmentally down-regulated ubiquitin-like protein. J Biol Chem 272:28557–28562.
Pozo JC, Timpte C, Tan S et al (1998) The ubiquitin-related protein RUB1 and auxin response in Arabidopsis. Science 280:1760–1763.
Osaka F, Saeki M, Katayama S et al (2000) Covalent modifier NEDD8 is essential for SCF ubiquitin-ligase in fission yeast. Embo J 19:3475–3484.
Ou CY, Lin YF, Chen YJ, Chien CT (2002) Distinct protein degradation mechanisms mediated by Cul1 and Cul3 controlling Ci stability in Drosophila eye development. Genes Dev 16:2403–2414.
Jones D, Candido EP (2000) The NED-8 conjugating system in Caenorhabditis elegans is required for embryogenesis and terminal differentiation of the hypodermis. Dev Biol 226:152–165.
Tateishi K, Omata M, Tanaka K, Chiba T (2001) The NEDD8 system is essential for cell cycle progression and morphogenetic pathway in mice. J Cell Biol 155:571–579.
Kerscher O, Felberbaum R, Hochstrasser M (2006) Modification of proteins by ubiquitin and ubiquitin-like proteins. Annu Rev Cell Dev Biol 22:159–180.
Xirodimas DP (2008) Novel substrates and functions for the ubiquitin-like molecule NEDD8. Biochem Soc Trans 36:802–806.
Rabut G, Peter M (2008) Function and regulation of protein neddylation. ‘Protein modifications: beyond the usual suspects’ review series. EMBO Rep 9:969–976.
Huang DT, Ayrault O, Hunt HW et al (2009) E2-RING expansion of the NEDD8 cascade confers specificity to cullin modification. Mol Cell 33:483–495.
Lyapina S, Cope G, Shevchenko A et al (2001) Promotion of NEDD-CUL1 conjugate cleavage by COP9 signalosome. Science 292:1382–1385.
Schwechheimer C, Serino G, Callis J et al (2001) Interactions of the COP9 signalosome with the E3 ubiquitin ligase SCFTIRI in mediating auxin response. Science 292:1379–1382.
Liakopoulos D, Doenges G, Matuschewski K, Jentsch S (1998) A novel protein modification pathway related to the ubiquitin system. Embo J 17:2208–2214.
Pan ZQ, Kentsis A, Dias DC et al (2004) Nedd8 on cullin: building an expressway to protein destruction. Oncogene 23:1985–1997.
Petroski MD, Deshaies RJ (2005) Function and regulation of cullin-RING ubiquitin ligases. Nat Rev Mol Cell Biol 6:9–20.
Skaar JR, Florens L, Tsutsumi T et al (2007) PARC and CUL7 form atypical cullin RING ligase complexes. Cancer Res 67:2006–2014.
Soucy TA, Smith PG, Milhollen MA et al (2009) An inhibitor of NEDD8-activating enzyme as a new approach to treat cancer. Nature 458:732–736.
Liu G, Xirodimas DP (2010) NUB1 promotes cytoplasmic localization of p53 through cooperation of the NEDD8 and ubiquitin pathways. Oncogene 29:2252–2261.
Acknowledgements
Research in DPX laboratory is supported by the Association for International Cancer Research (AICR) and INSERM. D.P.X. is an AICR Research Fellow.
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Leidecker, O., Xirodimas, D.P. (2012). Isolation of NEDDylated Proteins in Human Cells. In: Dohmen, R., Scheffner, M. (eds) Ubiquitin Family Modifiers and the Proteasome. Methods in Molecular Biology, vol 832. Humana Press. https://doi.org/10.1007/978-1-61779-474-2_8
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DOI: https://doi.org/10.1007/978-1-61779-474-2_8
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