title: Cytomegalovirus IE2 protein stimulates interleukin 1beta gene transcription via tethering to Spi-1/PU.1.
abstract: 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.
authors.affiliation: The New England Baptist Bone & Joint Institute, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02115, USA.
authors.forename: N
authors.initials: N
authors.lastname: Wara-aswapati
authors.forename: Z
authors.initials: Z
authors.lastname: Yang
authors.forename: W R
authors.initials: WR
authors.lastname: Waterman
authors.forename: Y
authors.initials: Y
authors.lastname: Koyama
authors.forename: S
authors.initials: S
authors.lastname: Tetradis
authors.forename: B K
authors.initials: BK
authors.lastname: Choy
authors.forename: A C
authors.initials: AC
authors.lastname: Webb
authors.forename: P E
authors.initials: PE
authors.lastname: Auron
mesh_terms: D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D006367:HeLa Cells; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism*; D007375:Interleukin-1 / Q000235:genetics*; D008562:Membrane Glycoproteins; D008957:Models, Genetic; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011518:Proto-Oncogene Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D000099232:Proto-Oncogene Protein Spi-1
chemical_list: D004268:DNA-Binding Proteins; D017874:Immediate-Early Proteins; D007375:Interleukin-1; D008562:Membrane Glycoproteins; D011518:Proto-Oncogene Proteins; D015534:Trans-Activators; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D000099232:Proto-Oncogene Protein Spi-1; C088645:IE1 protein, cytomegalovirus; C069186:IE2 protein, Cytomegalovirus; C077406:UL115 protein, Human herpesvirus 5; C071440:glycoprotein H, Cytomegalovirus; C499358:glycoprotein H, Human cytomegalovirus; C439068:glycoprotein O, cytomegalovirus