Inflammation Research

, Volume 68, Issue 12, pp 999–1010 | Cite as

Regulation of oxidized LDL-induced inflammatory process through NLRP3 inflammasome activation by the deubiquitinating enzyme BRCC36

  • Mohit Singh
  • Bhawna Kumari
  • Umesh C. S. YadavEmail author
Original Research Paper



Oxidized Low-Density Lipoprotein (oxLDL) is a well-established pro-inflammatory marker that activates the NLRP3 inflammasome. Ubiquitination plays an important role in modulating the stability and functions of various proteins. BRCC36 is a ubiquitin-modifying enzyme that plays a crucial role in protein stabilization and activation in the cytosol, but its role in OxLDL-induced NLRP3 inflammasome activation is not known. Here, we have investigated the role of deubiquitinating enzyme BRCC36 in regulating NLRP3 inflammasome during oxLDL stimulation.


Raw 264.7 murine macrophages were stimulated with oxLDL and effect of BRCC36 deubiquitination activity was assessed by fluorometric assay, and protein expression was assessed by Western blotting. The level of IL-1β measured by ELISA and LDH activity as pyroptotic cell death marker was assessed by fluorometric assay.


The results showed that oxLDL increased the level of NLRP3 in macrophages and also the level of active caspase-1 and IL-1β. It also modulated the expression of deubiquitinating enzymes and caused pyroptotic cell death as indicated by LDH release. Inhibiting the proteasomal activity by MG132 and siRNA-mediated silencing of BRCC36 in macrophages potentially suppressed oxLDL-induced NLRP3 inflammasome activation and IL-1β secretion. Furthermore, the inhibition of proteasomal deubiquitinating activity with specific BRCC36 inhibitor G5 also reduced the inflammatory cell death.


Taken together, our study suggests that deubiquitinating enzyme BRCC36 inhibition could potentially suppress oxLDL-induced inflammatory process by inhibiting NLRP3 activation and resultant IL-1β secretion.


Inflammation Deubiquitination BRCC36 NLRP3 inflammasome oxLDL Raw264.7 macrophages 



Mohit Singh acknowledges junior research fellowship (JRF) from University Grant Commission (UGC), Government of India, and Bhawna Kumari acknowledge junior research fellowship from Council of Scientific & Industrial Research (CSIR), Government of India.


  1. 1.
    Liu W, et al. OxLDL-induced IL-1 beta secretion promoting foam cells formation was mainly via CD36 mediated ROS production leading to NLRP3 inflammasome activation. Inflamm Res. 2014;63(1):33–43.CrossRefGoogle Scholar
  2. 2.
    Varghese JF, Patel R, Yadav UCS. Sterol regulatory element binding protein (SREBP) -1 mediates oxidized low-density lipoprotein (oxLDL) induced macrophage foam cell formation through NLRP3 inflammasome activation. Cell Signal. 2019;53:316–26.CrossRefGoogle Scholar
  3. 3.
    Mangan MSJ, et al. Targeting the NLRP3 inflammasome in inflammatory diseases. Nat Rev Drug Discov. 2018;17:588.CrossRefGoogle Scholar
  4. 4.
    He Y, Hara H, Núñez G. Mechanism and regulation of NLRP3 inflammasome activation. Trends Biochem Sci. 2016;41(12):1012–21.CrossRefGoogle Scholar
  5. 5.
    Juliana C, et al. Non-transcriptional priming and deubiquitination regulate NLRP3 inflammasome activation. J Biol Chem. 2012;287(43):36617–22.CrossRefGoogle Scholar
  6. 6.
    Shim D-W, Lee K-H. Posttranslational regulation of the NLR family pyrin domain-containing 3 inflammasome. Front Immunol. 2018;9:1054.CrossRefGoogle Scholar
  7. 7.
    Yang J, Liu Z, Xiao TS. Post-translational regulation of inflammasomes. Cell Mol Immunol. 2017;14(1):65–79.CrossRefGoogle Scholar
  8. 8.
    Heride C, Urbe S, Clague MJ. Ubiquitin code assembly and disassembly. Curr Biol. 2014;24(6):R215–20.CrossRefGoogle Scholar
  9. 9.
    Varadan R, et al. Structural properties of polyubiquitin chains in solution. J Mol Biol. 2002;324(4):637–47.CrossRefGoogle Scholar
  10. 10.
    Ng HM, et al. The Lys63-deubiquitylating enzyme BRCC36 limits DNA break processing and repair. J Biol Chem. 2016;291(31):16197–207.CrossRefGoogle Scholar
  11. 11.
    Shintani H. LDL isolation and copper-catalysed oxidation. Pharm Anal Acta. 2013;4:247.Google Scholar
  12. 12.
    Py BF, et al. Deubiquitination of NLRP3 by BRCC3 critically regulates inflammasome activity. Mol Cell. 2013;49(2):331–8.CrossRefGoogle Scholar
  13. 13.
    Pickart CM, Eddins MJ. Ubiquitin: structures, functions, mechanisms. Biochim Biophys Acta. 2004;1695(1–3):55–72.CrossRefGoogle Scholar
  14. 14.
    Cooper EM, et al. K63-specific deubiquitination by two JAMM/MPN + complexes: BRISC-associated Brcc36 and proteasomal Poh1. EMBO J. 2009;28(6):621–31.CrossRefGoogle Scholar
  15. 15.
    Sheedy FJ, et al. CD36 coordinates NLRP3 inflammasome activation by facilitating intracellular nucleation of soluble ligands into particulate ligands in sterile inflammation. Nat Immunol. 2013;14(8):812–20.CrossRefGoogle Scholar
  16. 16.
    Binder CJ, et al. Pneumococcal vaccination decreases atherosclerotic lesion formation: molecular mimicry between Streptococcus pneumoniae and oxidized LDL. Nat Med. 2003;9(6):736–43.CrossRefGoogle Scholar
  17. 17.
    Dinarello CA. Interleukin-1 in the pathogenesis and treatment of inflammatory diseases. Blood. 2011;117(14):3720–32.CrossRefGoogle Scholar
  18. 18.
    Song H, et al. The E3 ubiquitin ligase TRIM31 attenuates NLRP3 inflammasome activation by promoting proteasomal degradation of NLRP3. Nat Commun. 2016;7:13727.CrossRefGoogle Scholar
  19. 19.
    Drews O, Taegtmeyer H. Targeting the ubiquitin-proteasome system in heart disease: the basis for new therapeutic strategies. Antioxid Redox Signal. 2014;21(17):2322–43.CrossRefGoogle Scholar
  20. 20.
    Ng H-M, et al. The Lys63-deubiquitylating enzyme BRCC36 limits DNA break processing and repair. J Biol Chem. 2016;291(31):16197–207.CrossRefGoogle Scholar
  21. 21.
    Feng L, Wang J, Chen J. The Lys63-specific deubiquitinating enzyme BRCC36 is regulated by two scaffold proteins localizing in different subcellular compartments. J Biol Chem. 2010;285(40):30982–8.CrossRefGoogle Scholar
  22. 22.
    Zeqiraj E, et al. Higher-order assembly of BRCC36-KIAA0157 is required for DUB activity and biological function. Mol Cell. 2015;59(6):970–83.CrossRefGoogle Scholar
  23. 23.
    Yoshida T, et al. Proteasome inhibitor MG132 induces death receptor 5 through CCAAT/enhancer-binding protein homologous protein. Cancer Res. 2005;65(13):5662–7.CrossRefGoogle Scholar
  24. 24.
    Möller Westerberg C, Hägglund H, Nilsson G. Proteasome inhibition upregulates Bim and induces caspase-3-dependent apoptosis in human mast cells expressing the Kit D816V mutation. Cell Death Dis. 2012;3:e417.CrossRefGoogle Scholar
  25. 25.
    Yan K, et al. The deubiquitinating enzyme complex BRISC is required for proper mitotic spindle assembly in mammalian cells. J Cell Biol. 2015;210(2):209.CrossRefGoogle Scholar
  26. 26.
    Shao G, et al. The Rap80-BRCC36 de-ubiquitinating enzyme complex antagonizes RNF8-Ubc13-dependent ubiquitination events at DNA double strand breaks. Proc Natl Acad Sci USA. 2009;106(9):3166–71.CrossRefGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  • Mohit Singh
    • 1
  • Bhawna Kumari
    • 1
  • Umesh C. S. Yadav
    • 1
    Email author
  1. 1.Metabolic Disorders and Inflammatory Pathologies Laboratory, School of Life SciencesCentral University of GujaratGandhinagarIndia

Personalised recommendations