This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Ning, S.
Right arrow Articles by Pagano, J. S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ning, S.
Right arrow Articles by Pagano, J. S.

 Previous Article

Molecular and Cellular Biology, October 2008, p. 6536-6546, Vol. 28, No. 20
0270-7306/08/$08.00+0     doi:10.1128/MCB.00785-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

TRAF6 and the Three C-Terminal Lysine Sites on IRF7 Are Required for Its Ubiquitination-Mediated Activation by the Tumor Necrosis Factor Receptor Family Member Latent Membrane Protein 1{triangledown}

Shunbin Ning,1 Alex D. Campos,3 Bryant G. Darnay,3 Gretchen L. Bentz,1 and Joseph S. Pagano1,2*

Lineberger Comprehensive Cancer Center,1 Department of Microbiology and Immunology and Department of Medicine, University of North Carolina, School of Medicine, Chapel Hill, North Carolina 27599,2 Department of Experimental Therapeutics, University of Texas M. D. Anderson Cancer Center, Houston, Texas 770303

Received 15 May 2008/ Returned for modification 20 June 2008/ Accepted 30 July 2008

We have recently shown that interferon regulatory factor 7 (IRF7) is activated by Epstein-Barr virus latent membrane protein 1 (LMP1), a member of the tumor necrosis factor receptor (TNFR) superfamily, through receptor-interacting protein-dependent K63-linked ubiquitination (L. E. Huye, S. Ning, M. Kelliher, and J. S. Pagano, Mol. Cell. Biol. 27:2910-2918, 2007). In this study, with the use of small interfering RNA and TNFR-associated factor 6 (TRAF6) knockout cells, we first show that TRAF6 and its E3 ligase activity are required for LMP1-stimulated IRF7 ubiquitination. In Raji cells which are latently infected and express high levels of LMP1 and IRF7 endogenously, expression of a TRAF6 small hairpin RNA construct reduces endogenous ubiquitination and endogenous activity of IRF7. In TRAF6–/– mouse embryonic fibroblasts, reconstitution with TRAF6 expression, but not with TRAF6(C70A), which lacks the E3 ligase activity, recovers LMP1's ability to stimulate K63-linked ubiquitination of IRF7. Further, we identify IRF7 as a substrate for TRAF6 E3 ligase and show that IRF7 is ubiquitinated by TRAF6 at multiple sites both in vitro and in vivo. Most important, we determine that the last three C-terminal lysine sites (positions 444, 446, and 452) of human IRF7 variant A are essential for activation of IRF7; these are the first such sites identified. A ubiquitination-deficient mutant of IRF7 with these sites mutated to arginines completely loses transactivational ability in response not only to LMP1 but also to the IRF7 kinase I{kappa}B kinase {varepsilon}. In addition, we find that K63-linked ubiquitination of IRF7 occurs independently of its C-terminal functional phosphorylation sites. These data support our hypothesis that regulatory ubiquitination of IRF7 is a prerequisite for its phosphorylation. This is the first evidence to imply that ubiquitination is required for phosphorylation and activation of a transcription factor.

* Corresponding author. Mailing address: Lineberger Comprehensive Cancer Center, University of North Carolina, Campus Box 7295, Chapel Hill, NC 27599. Phone: (919) 966-5907. Fax: (919) 966-9673. E-mail: joseph_pagano{at}med.unc.edu

{triangledown} Published ahead of print on 8 August 2008.

Molecular and Cellular Biology, October 2008, p. 6536-6546, Vol. 28, No. 20
0270-7306/08/$08.00+0     doi:10.1128/MCB.00785-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Paz, S., Vilasco, M., Arguello, M., Sun, Q., Lacoste, J., Nguyen, T. L.-A., Zhao, T., Shestakova, E. A., Zaari, S., Bibeau-Poirier, A., Servant, M. J., Lin, R., Meurs, E. F., Hiscott, J. (2009). Ubiquitin-Regulated Recruitment of I{kappa}B Kinase {varepsilon} to the MAVS Interferon Signaling Adapter. Mol. Cell. Biol. 29: 3401-3412 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»