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Molecular and Cellular Biology, October 1999, p. 6803-6814, Vol. 19, No. 10
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Cytomegalovirus IE2 Protein Stimulates Interleukin 1beta Gene Transcription via Tethering to Spi-1/PU.1

Nawarat Wara-aswapati,1,2,dagger Zhiyong Yang,1,3 Wayne R. Waterman,1 Yoshinobu Koyama,1,3 Sotirios Tetradis,1,Dagger Bob K. Choy,1,3 Andrew C. Webb,4 and Philip E. Auron1,3,*

The New England Baptist Bone & Joint Institute, Beth Israel Deaconess Medical Center,1 Department of Periodontology, Harvard School of Dental Medicine,2 and Department of Medicine, Harvard Medical School,3 Boston, Massachusetts 02115, and Department of Biological Sciences, Wellesley College, Wellesley, Massachusetts 021814

Received 23 March 1999/Returned for modification 17 May 1999/Accepted 28 July 1999

Potent induction of the gene coding for human prointerleukin 1beta (il1b) normally requires a far-upstream inducible enhancer in addition to a minimal promoter located between positions -131 and +12. The transcription factor Spi-1 (also called PU.1) is necessary for expression and binds to the minimal promoter, thus providing an essential transcription activation domain (TAD). In contrast, infection by human cytomegalovirus (HCMV) can strongly activate il1b via the expression of immediate early (IE) viral proteins and eliminates the requirement for the upstream enhancer. Spi-1 has been circumstantially implicated as a host factor in this process. We report here the molecular basis for the direct involvement of Spi-1 in HCMV activation of il1b. Transfection of Spi-1-deficient HeLa cells demonstrated both the requirement of Spi-1 for IE activity and the need for a shorter promoter (-59 to +12) than that required in the absence of IE proteins. Furthermore, in contrast to normal, enhancer-dependent il1b expression, which absolutely requires both the Spi-1 winged helix-turn-helix (wHTH) DNA-binding domain and the majority of the Spi-1 TAD, il1b expression in the presence of IE proteins does not require the Spi-1 TAD, which plays a synergistic role. In addition, we demonstrate that a single IE protein, IE2, is critical for the induction of il1b. Protein-protein interaction experiments revealed that the wing motif within the Spi-1 wHTH domain directly recruits IE2. In turn, IE2 physically associates with the Spi-1 wing and requires the integrity of at least one region of IE2. Functional analysis demonstrates that both this region and a carboxy-terminal acidic TAD are required for IE2 function. Therefore, we propose a protein-tethered transactivation mechanism in which the il1b promoter-bound Spi-1 wHTH tethers IE2, which provides a TAD, resulting in the transactivation of il1b.

* Corresponding author. Mailing address: Harvard Institutes of Medicine, Room 245, 77 Avenue Louis Pasteur, Boston, MA 02115-5727. Phone: (617) 667-0741. Fax: (617) 975-5299. E-mail: pauron{at}caregroup.med.harvard.edu.

dagger Present address: Khon Kaen University, Faculty of Dentistry, Khon Kaen 40002, Thailand.

Dagger Present address: UCLA School of Dentistry, Los Angeles, CA 90095.

Molecular and Cellular Biology, October 1999, p. 6803-6814, Vol. 19, No. 10
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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