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Role of UbL Family Modifiers and Their Binding Proteins in Cell Signaling

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Ubiquitin Family Modifiers and the Proteasome

Part of the book series: Methods in Molecular Biology ((MIMB,volume 832))

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Abstract

The versatile function of ubiquitin (Ub) is powerfully illustrated by its appearance in multiple forms and shapes, like polymeric ubiquitin chains. These chains, when recognized by specific ubiquitin-binding domains (UBDs), give rise to extraordinary complex signaling networks that regulate virtually every cellular function. At the heart of our understanding of this complexity is the evolution and adaptation of technologies and methods to analyze ubiquitin biochemistry, e.g., covalent Ub–substrate conjugates as well as transient Ub–UBD interactions. Here, we describe seminal developments in those methodologies that have paved the way to our understanding of the diversity of Ub signals as well as their recognition and interpretation by UBD-containing proteins.

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References

  1. Hershko A, Ciechanover A (1998) The ubiquitin system. Annu Rev Biochem 67:425–479.

    Article  PubMed  CAS  Google Scholar 

  2. Varshavsky A (2005) Regulated protein degradation. Trends Biochem Sci 30:283–286.

    Article  PubMed  CAS  Google Scholar 

  3. Schulman, B A, Harper, J W (2009) Ubiquitin-like protein activation by E1 enzymes: the apex for downstream signalling pathways. Nat Rev Mol Cell Biol 10:319–331.

    Article  PubMed  CAS  Google Scholar 

  4. van Wijk SJ, Timmers HT (2010) The family of ubiquitin-conjugating enzymes (E2s): deciding between life and death of proteins. FASEB J 24:981–993.

    Article  PubMed  Google Scholar 

  5. Deshaies RJ, Joazeiro CA (2009) RING domain E3 ubiquitin ligases. Annu Rev Biochem 78:399–434.

    Article  PubMed  CAS  Google Scholar 

  6. Ravid T, Hochstrasser M (2008) Diversity of degradation signals in the ubiquitin-proteasome system. Nat Rev Mol Cell Biol 9:679–690.

    Article  PubMed  CAS  Google Scholar 

  7. Ikeda F, Dikic I (2008) Atypical ubiquitin chains: new molecular signals ‘Protein Modifications: Beyond the Usual Suspects’ review series. EMBO Rep 9:536–542.

    Article  PubMed  CAS  Google Scholar 

  8. Dikic I, Wakatsuki S, Walters KJ (2009) Ubiquitin-binding domains - from structures to functions. Nat Rev Mol Cell Biol 10:659–671.

    Article  PubMed  CAS  Google Scholar 

  9. Broemer M, Meier P (2009) Ubiquitin-mediated regulation of apoptosis. Trends Cell Biol 19:130–140.

    Article  PubMed  CAS  Google Scholar 

  10. Ulrich HD, Walden H (2010) Ubiquitin signalling in DNA replication and repair. Nat Rev Mol Cell Biol 11:479–489.

    Article  PubMed  CAS  Google Scholar 

  11. Bergink S, Jentsch S (2009) Principles of ubiquitin and SUMO modifications in DNA repair. Nature 458:461–467.

    Article  PubMed  CAS  Google Scholar 

  12. Hirsch C, Gauss R, Horn SC, et al (2009) The ubiquitylation machinery of the endoplasmic reticulum. Nature 458:453–460.

    Article  PubMed  CAS  Google Scholar 

  13. Wertz IE, Dixit VM (2010) Regulation of death receptor signaling by the ubiquitin system. Cell Death Differ 17:14–24.

    Article  PubMed  CAS  Google Scholar 

  14. Raiborg C, Stenmark H (2009) The ESCRT machinery in endosomal sorting of ubiquitylated membrane proteins. Nature 458:445–452.

    Article  PubMed  CAS  Google Scholar 

  15. Wickliffe K, Williamson A, Jin L, Rape M (2009) The multiple layers of ubiquitin-dependent cell cycle control. Chem Rev 109:1537–1548.

    Article  PubMed  CAS  Google Scholar 

  16. Dammer E, Peng J (2010) At the crossroads of ubiquitin signaling and mass spectrometry. Expert Rev Proteomics 7643–645.

    Google Scholar 

  17. Danielsen JM, Sylvestersen KB, Bekker-Jensen S, et al (2011) Mass spectrometric analysis of lysine ubiquitylation reveals promiscuity at site level. Mol Cell Proteomics 10:M110.003590.

    Google Scholar 

  18. Phu L, Izrael-Tomasevic A, Matsumoto ML, et al (2010) Improved quantitative mass spectrometry methods for characterizing complex ubiquitin signals. Mol Cell Proteomics doi: 10.1074/mcp.M110.003756.

    Google Scholar 

  19. Newton K, Matsumoto ML, Wertz IE, et al (2008) Ubiquitin chain editing revealed by polyubiquitin linkage-specific antibodies. Cell 134:668–678.

    Article  PubMed  CAS  Google Scholar 

  20. Matsumoto, M L, Wickliffe, K E, Dong, K C, et al (2010) K11-linked polyubiquitination in cell cycle control revealed by a K11 linkage-specific antibody. Mol Cell 39:477–484.

    Article  PubMed  CAS  Google Scholar 

  21. Perica T, Chothia C (2010) Ubiquitin--molecular mechanisms for recognition of different structures. Curr Opin Struct Biol 20:367–376.

    Article  PubMed  CAS  Google Scholar 

  22. Winget JM, Mayor T (2010) The diversity of ubiquitin recognition: hot spots and varied specificity. Mol Cell 38:627–635.

    Article  PubMed  CAS  Google Scholar 

  23. Dynek JN, Goncharov T, Dueber EC, et al (2010) c-IAP1 and UbcH5 promote K11-linked polyubiquitination of RIP1 in TNF signalling. EMBO J 29:4198–4209.

    Article  PubMed  CAS  Google Scholar 

  24. Eger S, Scheffner M, Marx A, Rubini M (2010) Synthesis of defined ubiquitin dimers. J Am Chem Soc 132:16337–16339.

    Article  PubMed  CAS  Google Scholar 

  25. El Oualid F, Merkx R, Ekkebus R, et al (2010) Chemical Synthesis of Ubiquitin, Ubiquitin-Based Probes, and Diubiquitin. Angew Chem Int Ed Engl 49 :10149–10153.

    Article  PubMed  CAS  Google Scholar 

  26. Kulathu Y, Akutsu M, Bremm A, et al (2009) Two-sided ubiquitin binding explains specificity of the TAB2 NZF domain. Nat Struct Mol Biol 16:1328–1330.

    Article  PubMed  CAS  Google Scholar 

  27. Komander D, Reyes-Turcu F, Licchesi JD, et al (2009) Molecular discrimination of structurally equivalent Lys 63-linked and linear polyubiquitin chains. EMBO Rep 10:466–473.

    Article  PubMed  CAS  Google Scholar 

  28. Bosanac, I, Wertz, I E, Pan B, et al (2010) Ubiquitin binding to A20 ZnF4 is required for modulation of NF-kappaB signaling. Mol Cell 40:548–557.

    Article  PubMed  CAS  Google Scholar 

  29. Ikeda F, Crosetto N, Dikic I (2010) What determines the specificity and outcomes of ubiquitin signaling? Cell 143:677–681.

    Article  PubMed  CAS  Google Scholar 

  30. Iwai K, Tokunaga F (2009) Linear polyubiquitination: a new regulator of NF-kappaB activation. EMBO Rep 10:706–713.

    Article  PubMed  CAS  Google Scholar 

  31. Pierce NW, Kleiger G, Shan SO, Deshaies RJ (2009) Detection of sequential polyubiquitylation on a millisecond timescale. Nature 462:615–619.

    Article  PubMed  CAS  Google Scholar 

  32. Clague MJ, Urbe S (2010) Ubiquitin: same molecule, different degradation pathways. Cell 143:682–685.

    Article  PubMed  CAS  Google Scholar 

  33. Kleiger G, Saha A, Lewis S, et al (2009) Rapid E2-E3 assembly and disassembly enable processive ubiquitylation of cullin-RING ubiquitin ligase substrates. Cell 139:957–968.

    Article  PubMed  CAS  Google Scholar 

  34. Yen HC, Elledge SJ (2008) Identification of SCF ubiquitin ligase substrates by global protein stability profiling. Science 322:923–929.

    Article  PubMed  CAS  Google Scholar 

  35. Xu P, Duong DM, Seyfried NT, et al (2009) Quantitative proteomics reveals the function of unconventional ubiquitin chains in proteasomal degradation. Cell 137:133–145.

    Article  PubMed  CAS  Google Scholar 

  36. Hjerpe R, Rodriguez MS (2008) Efficient approaches for characterizing ubiquitinated proteins. Biochem Soc Trans 36:823–827.

    Article  PubMed  CAS  Google Scholar 

  37. Meierhofer D, Wang X, Huang L, Kaiser P (2008) Quantitative analysis of global ubiquitination in HeLa cells by mass spectrometry. J Proteome Res 7:4566–4576.

    Article  PubMed  CAS  Google Scholar 

  38. Shi Y, Chan DW, Jung SY, et al (2011) A dataset of human endogenous ubiquitina­tion sites. Mol Cell Proteomics doi:M110.002089.

    Google Scholar 

  39. Nielsen ML, Vermeulen M, Bonaldi T, et al (2008) Iodoacetamide-induced artifact mimics ubiquitination in mass spectrometry. Nat Methods 5:459–460.

    Article  PubMed  CAS  Google Scholar 

  40. Golebiowski F, Tatham MH, Nakamura A, Hay RT (2010) High-stringency tandem affinity purification of proteins conjugated to ubiquitin-like moieties. Nat Protoc 5:873–882.

    Article  PubMed  CAS  Google Scholar 

  41. Golebiowski F, Matic I, Tatham MH, et al (2009) System-wide changes to SUMO modifications in response to heat shock. Sci Signal 2 :ra24.

    Google Scholar 

  42. Matic I, van Hagen M, Schimmel J, et al (2008) In vivo identification of human small ubiquitin-like modifier polymerization sites by high accuracy mass spectrometry and an in vitro to in vivo strategy. Mol Cell Proteomics 7:132–144.

    PubMed  CAS  Google Scholar 

  43. Vertegaal AC, Andersen JS, Ogg SC, et al (2006) Distinct and overlapping sets of SUMO-1 and SUMO-2 target proteins revealed by quantitative proteomics. Mol Cell Proteomics 5:2298–2310.

    Article  PubMed  CAS  Google Scholar 

  44. Spence J, Sadis S, Haas AL, Finley D (1995) A ubiquitin mutant with specific defects in DNA repair and multiubiquitination. Mol Cell Biol 15:1265–1273.

    PubMed  CAS  Google Scholar 

  45. Finley D, Sadis S, Monia BP, et al (1994) Inhibition of proteolysis and cell cycle progression in a multiubiquitination-deficient yeast mutant. Mol Cell Biol 14:5501–5509.

    PubMed  CAS  Google Scholar 

  46. Xu M, Skaug B, Zeng W, Chen ZJ (2009) A ubiquitin replacement strategy in human cells reveals distinct mechanisms of IKK activation by TNFalpha and IL-1beta. Mol Cell 36:302–314.

    Article  PubMed  CAS  Google Scholar 

  47. Raasi S, Pickart CM (2003) Rad23 ubiquitin-associated domains (UBA) inhibit 26 S proteasome-catalyzed proteolysis by sequestering lysine 48-linked polyubiquitin chains. J Biol Chem 278, 8951–8959

    Article  PubMed  CAS  Google Scholar 

  48. Raasi S, Orlov I, Fleming KG, Pickart CM (2004) Binding of polyubiquitin chains to ubiquitin-associated (UBA) domains of HHR23A. J Mol Biol 341, 1367–1379.

    Article  PubMed  CAS  Google Scholar 

  49. Hjerpe R, Aillet F, Lopitz-Otsoa F, et al (2009) Efficient protection and isolation of ubiquitylated proteins using tandem ubiquitin-binding entities. EMBO Rep 10:1250–1258.

    Article  PubMed  CAS  Google Scholar 

  50. Xu G, Paige JS, Jaffrey SR (2010) Global analysis of lysine ubiquitination by ubiquitin remnant immunoaffinity profiling. Nat Biotechnol 28:868–873.

    Article  PubMed  CAS  Google Scholar 

  51. Andersen, J S, Matic, I, Vertegaal, A C (2009) Identification of SUMO target proteins by quantitative proteomics. Methods Mol Biol 497:19–31

    Article  PubMed  CAS  Google Scholar 

  52. Bartee E, Eyster CA, Viswanathan K, et al (2010) Membrane-Associated RING-CH Proteins Associate with Bap31 and Target CD81 and CD44 to Lysosomes. PLoS One 5:e15132.

    Article  PubMed  CAS  Google Scholar 

  53. Xu P, Duong DM, Peng J (2009) Systematical optimization of reverse-phase chromatography for shotgun proteomics. J Proteome Res 8:3944–3950.

    Article  PubMed  CAS  Google Scholar 

  54. Kirkpatrick DS, Denison C, Gygi SP (2005) Weighing in on ubiquitin: the expanding role of mass-spectrometry-based proteomics. Nat Cell Biol 7:750–757.

    Article  PubMed  CAS  Google Scholar 

  55. Kirkpatrick DS, Gerber SA, Gygi SP (2005) The absolute quantification strategy: a general procedure for the quantification of proteins and post-translational modifications. Methods 35:265–273.

    Article  PubMed  CAS  Google Scholar 

  56. Del Rincon SV, Rogers J, Widschwendter M, et al (2010) Development and validation of a method for profiling post-translational modification activities using protein microarrays. PLoS One 5:e11332.

    Article  PubMed  Google Scholar 

  57. Persaud A, Alberts P, Amsen, EM, et al (2009) Comparison of substrate specificity of the ubiquitin ligases Nedd4 and Nedd4-2 using proteome arrays. Mol Syst Biol 5:333.

    Article  PubMed  Google Scholar 

  58. Gupta R, Kus B, Fladd C, et al (2007) Ubiquitination screen using protein microarrays for comprehensive identification of Rsp5 substrates in yeast. Mol Syst Biol 3:116.

    Article  PubMed  Google Scholar 

  59. Merbl Y, Kirschner MW (2009) Large-scale detection of ubiquitination substrates using cell extracts and protein microarrays. Proc Natl Acad Sci U S A 106 :2543–2548.

    Article  PubMed  CAS  Google Scholar 

  60. Fushman D, Walker O (2010) Exploring the linkage dependence of polyubiquitin conformations using molecular modeling. J Mol Biol 395:803–814.

    Article  PubMed  CAS  Google Scholar 

  61. Bremm A, Freund SM, Komander D (2010) Lys11-linked ubiquitin chains adopt compact conformations and are preferentially ­hydrolyzed by the deubiquitinase Cezanne. Nat Struct Mol Biol 17:939–947.

    Article  PubMed  CAS  Google Scholar 

  62. Varadan R, Walker O, Pickart C, Fushman D (2002) Structural properties of polyubiquitin chains in solution. J Mol Biol 324:637–647.

    Article  PubMed  CAS  Google Scholar 

  63. Rahighi S, Ikeda F, Kawasaki M, et al (2009) Specific recognition of linear ubiquitin chains by NEMO is important for NF-kappaB activation. Cell 136:1098–1109.

    Article  PubMed  CAS  Google Scholar 

  64. Wang H, Matsuzawa A, Brown SA, et al (2008) Analysis of nondegradative protein ubiquitylation with a monoclonal antibody specific for lysine-63-linked polyubiquitin. Proc Natl Acad Sci U S A 105:20197–20202.

    Article  PubMed  CAS  Google Scholar 

  65. Tokunaga F, Sakata S, Saeki Y, et al (2009) Involvement of linear polyubiquitylation of NEMO in NF-kappaB activation. Nat Cell Biol 11:123–132.

    Article  PubMed  CAS  Google Scholar 

  66. Swanson KA, Kang RS, Stamenova SD, et al (2003) Solution structure of Vps27 UIM-ubiquitin complex important for endosomal sorting and receptor downregulation. EMBO J 22:4597–4606.

    Article  PubMed  CAS  Google Scholar 

  67. Ohno A, Jee J, Fujiwara K, et al (2005) Structure of the UBA domain of Dsk2p in complex with ubiquitin molecular determinants for ubiquitin recognition. Structure 13:521–532.

    Article  PubMed  CAS  Google Scholar 

  68. Wagner S, Carpentier I, Rogov V, et al (2008) Ubiquitin binding mediates the NF-kappaB inhibitory potential of ABIN proteins. Oncogene 27:3739–3745.

    Article  PubMed  CAS  Google Scholar 

  69. Bienko M, Green C M, Crosetto N, et al (2005) Ubiquitin-binding domains in Y-family polymerases regulate translesion synthesis. Science 310:1821–1824.

    Article  PubMed  CAS  Google Scholar 

  70. Lee S, Tsai YC, Mattera R, et al (2006) Structural basis for ubiquitin recogniti.on and autoubiquitination by Rabex-5, Nat Struct Mol Biol 13:264–271

    Article  PubMed  CAS  Google Scholar 

  71. Husnjak K, Elsasser S, Zhang N, et al (2008) Proteasome subunit Rpn13 is a novel ubiquitin receptor. Nature 453:481–488.

    Article  PubMed  CAS  Google Scholar 

  72. VanDemark AP, Hofmann RM, Tsui C, et al (2001) Molecular insights into polyubiquitin chain assembly: crystal structure of the Mms2/Ubc13 heterodimer. Cell 105:711–720.

    Article  PubMed  CAS  Google Scholar 

  73. Brzovic PS, Lissounov A, Christensen DE, et al (2006) A UbcH5/ubiquitin noncovalent complex is required for processive BRCA1-directed ubiquitination. Mol Cell 21:873–880.

    Article  PubMed  CAS  Google Scholar 

  74. Bienko M, Green CM, Sabbioneda S, et al (2010) Regulation of translesion synthesis DNA polymerase eta by monoubiquitination. Mol Cell 37:396–407.

    Article  PubMed  CAS  Google Scholar 

  75. Pickart C M, Raasi S (2005) Controlled synthesis of polyubiquitin chains. Methods Enzymol 399:21–36.

    Article  PubMed  CAS  Google Scholar 

  76. Raasi S, Pickart CM (2005) Ubiquitin chain synthesis. Methods Mol Biol 301:47–55.

    PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Frankfurt Institute for Molecular Life Sciences and Institute of Biochemistry II, Goethe University School of Medicine, Frankfurt (Main), Germany

    Sjoerd J. L. van Wijk, Magda Bienko & Ivan Dikic

  2. Departments of Physics and of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA

    Magda Bienko

Authors
  1. Sjoerd J. L. van Wijk
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  2. Magda Bienko
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  3. Ivan Dikic
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Correspondence to Ivan Dikic .

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Editors and Affiliations

  1. Institut für Genetik, Universität Köln, Zülpicher Str. 47, Köln, 50674, Germany

    R. Jürgen Dohmen

  2. Fak. Biologie, Universität Konstanz, Konstanz, 78457, Germany

    Martin Scheffner

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van Wijk, S.J.L., Bienko, M., Dikic, I. (2012). Role of UbL Family Modifiers and Their Binding Proteins in Cell Signaling. 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_11

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  • DOI: https://doi.org/10.1007/978-1-61779-474-2_11

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-473-5

  • Online ISBN: 978-1-61779-474-2

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