Abstract
Immunoproteasomes (IPs) containing the interferon-inducible subunits β1i (LMP2), β2i (MECL-1), and β5i (LMP7) alter proteasomal cleavage preference, optimise the generation of peptide ligands of MHC class I molecules, alter cytokine profile, influence T-helper cell differentiation, and play a role in T-cell survival. Small molecule inhibitors are useful tools for probing the role of the immunoproteasome in immune functions. Here, we describe different methods to characterise immunoproteasome-selective inhibitors. Thereby, we provide the methodology to analyse the specificity and cell permeability of immunoproteasome inhibitors, as well as to functionally investigate immunoproteasome inhibitors in antigen presentation.
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References
Adams J, Palombella VJ, Sausville EA, et al. (1999) Proteasome inhibitors: A novel class of potent and effective antitumor agents. Cancer Res 59:2615–2622.
Basler M, Youhnovski N, Van Den Broek M, et al. (2004) Immunoproteasomes down-regulate presentation of a subdominant T cell epitope from lymphocytic choriomeningitis virus. J Immunol 173:3925–3934.
Basler M, Moebius J, Elenich L, et al. (2006) An Altered T Cell Repertoire in MECL-1-Deficient Mice. J Immunol 176:6665–6672.
Groettrup M, Kirk CJ, Basler M. (2010) Proteasomes in immune cells: more than peptide producers? Nat Rev Immunol 10:73–78.
Muchamuel T, Basler M, Aujay MA, et al. (2009) A selective inhibitor of the immunoproteasome subunit LMP7 blocks cytokine production and attenuates progression of experimental arthritis. Nat Med 15:781–787.
van Swieten PF, Samuel E, Hernandez RO, et al. (2007) A cell-permeable inhibitor and activity-based probe for the caspase-like activity of the proteasome. Bioorg Med Chem Lett 17:3402–3405.
Ho YK, Bargagna-Mohan P, Wehenkel M, et al. (2007) LMP2-specific inhibitors: chemical genetic tools for proteasome biology. Chem Biol 14:419–430.
Kuhn DJ, Hunsucker SA, Chen Q, et al. (2009) Targeted inhibition of the immunoproteasome is a potent strategy against models of multiple myeloma that overcomes resistance to conventional drugs and nonspecific proteasome inhibitors. Blood 113:4667–4676.
Blackburn C, Gigstad KM, Hales P, et al. (2010) Characterization of a new series of non-covalent proteasome inhibitors with exquisite potency and selectivity for the 20S beta5-subunit. Biochem J 430:461–476.
Basler M, Dajee M, Moll C, et al. (2010) Prevention of experimental colitis by a selective inhibitor of the immunoproteasome. J Immunol 185:634–641.
Schwarz K, Eggers M, Soza A, et al. (2000) The proteasome regulator PA28α/β can enhance antigen presentation without affecting 20S proteasome subunit composition. Eur. J. Immunol. 30:3672–3679.
Khan S, van den Broek M, Schwarz K, et al. (2001) Immunoproteasomes largely replace constitutive proteasomes during an antiviral and antibacterial immune response in the liver. J. Immunol. 167:6859–6868.
Toes REM, Nussbaum AK, Degermann S, et al. (2001) Discrete cleavage motifs of constitutive and immunoproteasomes revealed by quantitative analysis of cleavage products. J. Exp. Med. 194:1–12.
Groettrup M, Kraft R, Kostka S, et al. (1996) A third interferon-γ-induced subunit exchange in the 20S proteasome. Eur. J. Immunol. 26:863–869.
Van Kaer L, Ashton-Rickardt PG, Eichelberger M, et al. (1994) Altered peptidase and viral-specific T cell response in LMP 2 mutant mice. Immunity 1:533–541.
Fehling HJ, Swat W, Laplace C, et al. (1994) MHC class I expression in mice lacking proteasome subunit LMP-7. Science 265:1234–1237.
Gileadi U, Gallimore A, van der Bruggen P, et al. (1999) Effect of epitope flanking residues on the presentation of N-terminal cytotoxic T lymphocyte epitopes. Eur. J. Immunol. 29:2213–2222.
Acknowledgements
This work was supported by grants from the German Research Foundation (GR 1517/12-1) and Fritz Thyssen Foundation (Az.10.10.2.122). We thank Ulrike Beck for excellent technical assistance in establishing the methods described in this chapter.
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Basler, M., Groettrup, M. (2012). Immunoproteasome-Specific Inhibitors and Their Application. 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_27
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DOI: https://doi.org/10.1007/978-1-61779-474-2_27
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