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Bilateral inhibition of HAUSP deubiquitinase by a viral interferon regulatory factor protein

Abstract

Herpesvirus-associated ubiquitin-specific protease (HAUSP) regulates the stability of p53 and the p53-binding protein MDM2, implicating HAUSP as a therapeutic target for tuning p53-mediated antitumor activity. Here we report the structural analysis of HAUSP with Kaposi's sarcoma–associated herpesvirus viral interferon (IFN) regulatory factor 4 (vIRF4) and the discovery of two vIRF4-derived peptides, vif1 and vif2, as potent and selective HAUSP antagonists. This analysis reveals a bilateral belt-type interaction that results in inhibition of HAUSP. The vif1 peptide binds the HAUSP TRAF domain, competitively blocking substrate binding, whereas the vif2 peptide binds both the HAUSP TRAF and catalytic domains, robustly suppressing its deubiquitination activity. Peptide treatments comprehensively blocked HAUSP, leading to p53-dependent cell-cycle arrest and apoptosis in culture and to tumor regression in xenograft mouse model. Thus, the virus has developed a unique strategy to target the HAUSP–MDM2–p53 pathway, and these virus-derived short peptides represent biologically active HAUSP antagonists.

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Figure 1: vIRF4 interacts with HAUSP.
Figure 2: Structural basis for the interaction between HAUSP and vIRF4.
Figure 3: Bilateral interaction of vIRF4 with HAUSP.
Figure 4: Effect of vIRF4 peptides on HAUSP DUB enzymatic activity.
Figure 5: Cytotoxic effect of the TAT-vif1 or TAT-vif2 peptide on PEL cells.
Figure 6: TAT-vif1 and TAT-vif2 induce tumor suppression in vivo.
Figure 7: Combination therapeutic effects of a low dose of TAT-vif1 and TAT-vif2 peptides.

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Acknowledgements

This work was partly supported by grants CA82057, CA31363, CA115284, CA147868, CA148616, DE019085, the Hastings Foundation, the Fletcher Jones Foundation, the William Lawrence and Blanche Hughes (WLBH) Foundation and the Global Research Laboratory (GRL) Program (K20815000001) from the National Research Foundation of Korea (J.U.J.); and the 21C Frontier Microbial Genomics and Applications Center Program and the National Research Foundation of Korea grant (NRF-M1AXA002-2010-0029767) funded by the Ministry of Education, Science and Technology, Korea (M.H.K.). We thank the staff at the 4A and 6B beamlines, Pohang Accelerator Laboratory, Korea, for help with data collection. We thank V. Lombardi, J. Jeong and J.S. Lee for their help. Finally, we thank all lab members for their support and discussions.

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Contributions

H.-R.L. performed all aspect of this study; W.-C.C., J.H. and M.H.K. did X-ray crystallographic and biochemical studies; S.L. and Z.T. assisted with the experimental design and collected the data; E.H. and Y.H.J. carried out NMR studies; K.G. did protein purification; J.H. provided the KSHV library; H.-R.L., W.-C.C., T.-K.O., M.H.K. and J.U.J. organized this study and wrote the manuscript. All authors discussed the results and commented on the manuscript.

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Correspondence to Tae-Kwang Oh, Myung Hee Kim or Jae U Jung.

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The authors declare no competing financial interests.

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Lee, HR., Choi, WC., Lee, S. et al. Bilateral inhibition of HAUSP deubiquitinase by a viral interferon regulatory factor protein. Nat Struct Mol Biol 18, 1336–1344 (2011). https://doi.org/10.1038/nsmb.2142

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