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.