Abstract
Many intracellular proteins are metabolically unstable or can become unstable during their lifetime in a cell. The in vivo half-lives of specific proteins range from less than a minute to many days. Among the functions of intracellular proteolysis are the elimination of misfolded or otherwise abnormal proteins; maintenance of amino acid pools in cells affected by stresses such as starvation; and generation of protein fragments that act as hormones, antigens, or other effectors. One major function of proteolytic pathways is the selective destruction of proteins whose concentrations must vary with time and alterations in the state of a cell. Short in vivo half-lives of such proteins provide a way to generate their spatial gradients and to rapidly adjust their concentration or subunit composition through changes in the rate of their degradation. The regulated (and processive) degradation of intracellular proteins is carried out largely by the ubiquitin–proteasome system (Ub system), in conjunction with autophagy-lysosome pathways. Other contributors to intracellular proteolysis include cytosolic and nuclear proteases, such as caspases, calpains, and separases. They often function as “upstream” components of the Ub system, which destroys protein fragments that had been produced by these (nonprocessive) proteases. Ub, a 76-residue protein, mediates selective proteolysis through its enzymatic conjugation to proteins that contain primary degradation signals (degrons (1)), thereby marking such proteins for degradation by the 26S proteasome, an ATP-dependent multisubunit protease. Ub conjugation involves the formation of a poly-Ub chain that is linked (in most cases) to the ε-amino group of an internal Lys residue in a substrate protein. Ub is a “secondary” degron, in that Ub is conjugated to proteins that contain primary degradation signals.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Varshavsky A (1991) Naming a targeting signal. Cell 64:13–15.
Hershko A, Ciechanover A, Varshavsky A (2000) The ubiquitin system. Nat Med 10:1073–1081.
Varshavsky A (2006) The early history of the ubiquitin field. Pro Sci 15:647–654.
Varshavsky A (2008) Discovery of cellular regulation by protein degradation. J Biol Chem 283: 34469–34489.
Malynn B A, Ma A (2010) Ubiquitin makes its mark on immune regulation. Immunity 33:843–852.
Liu F, Walters K J (2010) Multitasking with ubiquitin through multivalent interactions. Trends Biochem Sci 35:352–360.
Gallastegui N, Groll M (2010) The 26S proteasome: assembly and function of a destructive machine. Trends Biochem Sci 35:634–642.
Bohn S, Beck F, Sakata E et al. (2010) Structure of the 26S proteasome from Schizosaccharomyces pombe at subnanometer resolution. Proc Natl Acad Sci USA 107:20992–20997.
Ulrich H D, Walden H (2010) Ubiquitin signalling in DNA replication and repair. Nat Rev Mol Cell Biol 11:479–489.
Stolz A, Wolf D H (2010) Endoplasmic reticulum-associated protein degradation: a chaperone-assisted journey to hell. Biochim Biophys Acta 1803:694–705.
Lu Z, Hunter T (2009) Degradation of activated protein kinases by ubiquitination. Annu Rev Biochem 78:435–475.
Hampton R Y, Garza R M (2009) Protein quality control as a strategy for cellular regulation: lessons from ubiquitin-mediated regulation of the sterol pathway. Chem Rev 109:1561–1574.
Grabbe C, Dikic I (2009) Functional roles of ubiquitin-like domain (ULD) and ubiquitin-binding domain (UBD) containing proteins. Chem Rev 109:1481–1494.
Daulni A, Tansey W P (2009) Damage control: DNA repair, transcription, and the ubiquitin-proteasome system. DNA Repair 8:444–448.
Deshaies R J, Joazeiro C A P (2009) RING domain E3 ubiquitin ligases. Annu Rev Biochem 78:399–434.
Finley D (2009) Recognition and processing of ubiquitin-protein conjugates by the proteasome. Annu Rev Biochem 78:477–513.
Reyes-Turcu F E, Ventii K H, Wilkinson K D (2009) Regulation and cellular roles of ubiquitin-specific deubiquitinating enzymes. Annu Rev Biochem 78:363–397.
Hirsch C, Gauss R, Horn S C et al. (2009) The ubiquitylation machinery of the endoplasmic reticulum. Nature 458:453–460.
Marques A J, Palanimurugan R, Matias A C et al. (2009) Catalytic mechanism and assembly of the proteasome. Chem Rev 109:1509–1536.
Ravid T, Hochstrasser M (2008) Diversity of degradation signals in the ubiquitin-proteasome system. Nat Rev Mol Cell Biol 9:679–689.
Vembar S S, Brodsky J L (2008) One step at a time: endoplasmic reticulum-associated degradation. Nat Rev Mol Cell Biol 9:944–958.
Dye B T, Schulman B A (2007) Structural mechanisms underlying posttranslational modification by ubiquitin-like proteins. Annu Rev Biophys Biomol Struct 36:131–150.
Scheffner M, Staub O (2007) HECT E3s and human disease. BMC Biochemistry 8 (Suppl. I):S6.
Scott D C, Monda J K, Grace C R R et al. (2010) A dual mechanism for Rub1 ligation to Cdc53. Mol Cell 39:784–796.
Loeb K R, Haas A L (1992) The interferon-inducible 15-kDa ubiquitin homolog conjugates to intracellular proteins. J Biol Chem 267:7806–7813.
Hochstrasser M (2009) Origin and function of ubiquitin-like proteins. Nature 458:422–429.
Bawa-Khalfe T, Yeh E T (2010) SUMO losing balance: SUMO proteases disrupt SUMO homeostasis to facilitate cancer development and progression. Genes Cancer 1:748–752.
Gareau J R, Lima C D (2010) The SUMO pathway: emerging mechanisms that shape specificity, conjugation and recognition. Nat Rev Mol Cell Biol 11:861–871.
Rubenstein E M, Hochstrasser M (2010) Redundancy and variation in the ubiquitin-mediated proteolytic targeting of a transcription factor. Cell Cycle 9:4282–4285.
Merlet J, Burger J, Gomes J E et al. (2009) Regulation of cullin-RING E3 ubiquitin-ligases by neddylation and dimerization. Cell Mol Life Sci 66:1924–1938.
Bergink S, Jentsch S (2009) Principles of ubiquitin and SUMO modifications in DNA repair. Nature 458:461–467.
Burroughs A M, Balaji S, Iyer L M et al. (2007) Small but versatile: the extraordinary functional and structural diversity of the beta-grasp fold. Biol Direct 2:18.
Iyer L M, Burroughs A M, Aravind L (2006) The prokaryotic antecedents of the ubiquitin-signaling system and the early evolution of ubiquitin-like beta-grasp domains. Genome Biol 7:R60.
Uzunova K, Göttsche K, Miteva M et al. (2007) Ubiquitin-dependent proteolytic control of SUMO conjugates. J Biol Chem 282:34167–34175.
Johnson E S (2004) Protein modification by SUMO. Annu Rev Biochem 73:355–382.
Geoffroy M-C, Hay R T (2010) An additional role for SUMO in ubiquitin-mediated proteolysis. Nat Rev Mol Cell Biol 10:564–568.
Zhao C, Hsiang T Y, Kuo R L et al. (2010) ISG15 conjugation system targets the viral NS1 protein in influenza A virus-infected cells. Proc Natl Acad Sci USA 107:2253–2258.
Durfee L A, Lyon N, Seo K et al. (2010) The ISG15 conjugation system broadly targets newly synthesized proteins: implications for the antiviral function of ISG15. Mol Cell 38:722–732.
Skaug B, Chen Z J (2010) Emerging role of ISG15 in antiviral immunity. Cell 143:187–190.
Bachmair A, Finley D, Varshavsky A (1986) In vivo half-life of a protein is a function of its amino-terminal residue. Science 234:179–186.
Hwang C-S, Shemorry A, Varshavsky A (2010) N-terminal acetylation of cellular proteins creates specific degradation signals. Science 327:973–977.
Arnesen T, Van Damme P, Polevoda B et al. (2009) Proteomics analyses reveal the evolutionary conservation and divergence of N-terminal acetyltransferases from yeast to humans. Proc Natl Acad Sci USA 106:8157–8162.
Helbig A O, Gauci S, Raijmakers R et al. (2010) Profiling of N-acetylated protein termini provides in-depth insights into the N-terminal nature of the proteome. Mol Cell Proteom 9:928–939.
Polevoda B, Sherman F (2003) N-terminal acetyltransferases and sequence requirements for N-terminal acetylation of eukaryotic proteins. J Mol Biol 325:595–622.
Goetze S, Qeli E, Mosimann C et al. (2009) Identification and functional characterization of N-terminally acetylated proteins in Drosophila melanogaster. PLoS Biol 7:e1000236.
Moerschell R P, Hosokawa Y, Tsunasawa S et al. (1990) The specificities of yeast methionine aminopeptidase and acetylation of amino-terminal methionine in vivo. Processing of altered iso-1-cytochromes created by oligonucleotide transformation. J Biol Chem 265:19638–19643.
Frottin F, Martinez A, Peynot P et al. (2006) The proteomics of N-terminal methionine cleavage. Mol Cell Proteomics 5:2336–2349.
Mullen J R, Kayne P S, Moerschell R P et al. (1989) Identification and characterization of genes and mutants for an N-terminal acetyltransferase from yeast. EMBO J 8:2067–2075.
Park E C, Szostak J W (1992) ARD1 and NAT1 proteins form a complex that has N-terminal acetyltransferase activity. EMBO J 11:2087–2093.
Gautschi M, Just S, Mun A et al. (2003) The yeast N-alpha-acetyltransferase NatA is quantitatively anchored to the ribosome and interacts with nascent polypeptides. Mol Cell Biol 23:7403–7414.
Tasaki T, Kwon Y T (2007) The mammalian N-end rule pathway: new insights into its components and physiological roles. Trends Biochem Sci 32:520–528.
Mogk A, Schmidt R, Bukau B (2007) The N-end rule pathway of regulated proteolysis: prokaryotic and eukaryotic strategies. Trends Cell Biol 17:165–172.
Eisele F, Wolf D H (2008) Degradation of misfolded proteins in the cytoplasm by the ubiquitin ligase Ubr1. FEBS Lett 582:4143–4146.
Heck J W, Cheung S K, Hampton R Y (2010) Cytoplasmic protein quality control degradation mediated by parallel actions of the E3 ubiquitin ligases Ubr1 and San1. Proc Natl Acad Sci USA 107:1106–1111.
Hwang C-S, Varshavsky A (2008) Regulation of peptide import through phosphorylation of Ubr1, the ubiquitin ligase of the N-end rule pathway. Proc Natl Acad Sci USA 105:19188–19193.
Hwang C-S, Shemorry A, Varshavsky A (2009) Two proteolytic pathways regulate DNA repair by co-targeting the Mgt1 alkyguanine transferase. Proc Natl Acad Sci USA 106:2142–2147.
Hu R-G, Wang H, **a Z et al. (2008) The N-end rule pathway is a sensor of heme. Proc Natl Acad Sci USA 105:76–81.
Hu R-G, Sheng J, **n Q et al. (2005) The N-end rule pathway as a nitric oxide sensor controlling the levels of multiple regulators. Nature 437:981–986.
Wang H, Piatkov K I, Brower C S et al. (2009) Glutamine-specific N-terminal amidase, a component of the N-end rule pathway. Mol Cell 34:686–695.
Graciet E, Wellmer F (2010) The plant N-end rule pathway: structure and functions. Trends Plant Sci 15:447–453.
Brower C S, Varshavsky A (2009) Ablation of arginylation in the mouse N-end rule pathway: loss of fat, higher metabolic rate, damaged spermatogenesis, and neurological perturbations. PLoS One 4:e7757.
Zenker M, Mayerle J, Lerch M M et al. (2005) Deficiency of UBR1, a ubiquitin ligase of the N-end rule pathway, causes pancreatic dysfunction, malformations and mental retardation (Johanson-Blizzard syndrome). Nat Genet 37:1345–1350.
Hwang C-S S, M., Batygin O, Addor M C et al. (2011) Ubiquitin ligases of the N-end rule pathway: assessment of mutations in UBR1 that cause the Johanson-Blizzard syndrome. PLoS One 6:e24925.
Prasad R, Kawaguchi S, Ng D T W (2010) A nucleus-based quality control mechanism for cytosolic proteins. Mol Biol Cell 21:2117–2127.
Kurosaka S, Leu N A, Zhang F et al. (2010) Arginylation-dependent neural crest cell migration is essential for mouse development. PLoS Genet 6:e1000878.
Zhang F, Saha S, Shabalina S A et al. (2010) Differential arginylation of actin isoforms is regulated by coding sequence-dependent degradation. Science 329.
Baker R T, Varshavsky A (1991) Inhibition of the N-end rule pathway in living cells. Proc Natl Acad Sci USA 87:2374–2378.
Varshavsky A (1996) The N-end rule: functions, mysteries, uses. Proc Natl Acad Sci USA 93:12142–12149.
Buchler N E, Gerland U, Hwa T (2005) Nonlinear protein degradation and the function of genetic circuits. Proc Natl Acad Sci USA 102:9559–9564.
Lam Y W, Lamond A I, Mann M et al. (2007) Analysis of nucleolar protein dynamics reveals the nuclear degradation of ribosomal proteins. Curr Biol 17:749–760.
Singh R K, Kabbaj M-H M, Paik J et al. (2009) Histone levels are regulated by phosphorylation and ubiquitylation-dependent proteolysis. Nat Cell Biol 11:925–933.
Johnson E S, Gonda D K, Varshavsky A (1990) Cis-trans recognition and subunit-specific degradation of short-lived proteins. Nature 346:287–291.
Hochstrasser M, Varshavsky A (1990) In vivo degradation of a transcriptional regulator: the yeast MATalpha2 repressor. Cell 61:697–708.
Schrader E K, Harstad K G, Matouschek A (2009) Targeting proteins for degradation. Nat Chem Biol 5:815–822.
Collins G A, Lipford J R, Deshaies R J et al. (2010) Gal4 turnover and transcription activation. Nature 461:E7-E8.
Wang X, Muratani M, Tansey W P et al. (2010) Proteolytic instability and the action of nonclassical transcriptional activators. Curr Biol 20:868–871.
Murray A W (2004) Recycling the cell cycle: cyclins revisited. Cell 116:221–234.
Powers E T, Morimoto R I, Dillin A et al. (2009) Biological and chemical approaches to diseases of proteostasis deficiency. Annu Rev Biochem 78:959–991.
Graciet E, Hu R G, Piatkov K et al. (2006) Aminoacyl-transferases and the N-end rule pathway of prokaryotic/eukaryotic specificity in a human pathogen. Proc Natl Acad Sci USA 103:3078–3083.
Finley D, Bartel B, Varshavsky A (1989) The tails of ubiquitin precursors are ribosomal proteins whose fusion to ubiquitin facilitates ribosome biogenesis. Nature 338:394–401.
Bedford L, Lowe J, Dick L R et al. (2011) Ubiquitin-like protein conjugation and the ubiquitin-proteasome system as drug targets. Nat Rev Drug Discov 10:29–46.
Hwang C-S, Shemorry A, Varshavsky A (2010) The N-end rule pathway is mediated by a complex of the RING-type Ubr1 and HECT-type Ufd4 ubiquitin ligases. Nat Cell Biol 12:1177–1185.
**a Z, Webster A, Du F et al. (2008) Substrate-binding sites of UBR1, the ubiquitin ligase of the N-end rule pathway. J Biol Chem 283:24011–24028.
Acknowledgments
I thank R. Hoffman (University of California, San Diego, USA), C. Brower, A. Shemorry, and B. Wadas (California Institute of Technology, USA) for helpful comments on the manuscript. Studies in our laboratory are supported by grants from the National Institutes of Health and the March of Dimes Foundation.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Varshavsky, A. (2012). Three Decades of Studies to Understand the Functions of the Ubiquitin Family. 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_1
Download citation
DOI: https://doi.org/10.1007/978-1-61779-474-2_1
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-61779-473-5
Online ISBN: 978-1-61779-474-2
eBook Packages: Springer Protocols