The Tax protein encoded by human T cell leukemia virus type I transactivates the viral promoter by forming a complex with several cellular factors bound to three repeats of a specific upstream regulatory sequence. We have shown that transactivation by Tax was correlated with its ability to interact with the C-terminal moiety of the TATA box-binding protein (TBP). In the present study, the ability of The viral protein Tax strongly activates transcription of the human T cell leukemia virus type I (HTLV-I) provirus and of a specific group of cellular genes (1). Various studies have established that Tax is recruited to the Tax-responsive element 1 of the HTLV-I promoter and to the serum-responsive element of the c-fos promoter (2-5) via interactions with cellular transcription factors. In agreement with this notion, a GAL4 DNA binding domain-Tax fusion protein stimulates transcription of a reporter gene under the control of GAL4 sites (6). Using this approach, we have shown that the transcriptional activity of the GAL4-Tax fusion protein involves a direct protein-protein interaction between Tax and TATA box-binding protein (TBP) (7).However, in vitro transcription experiments have established that activation by Tax requires the transcription factor complex TFIID comprised of TBP and TBP-associated factors (TAF II s) (8). To better understand the molecular mechanisms underlying activation by Tax, we investigated the interactions between Tax and hTAF II s. These experiments show a selective interaction of Tax with hTAF II 28. Increasing the intracellular concentration of hTAF II 28 augments the activity of GAL4-Tax, and the overexpression of both TBP and hTAF II 28 resulted in an additive increase in activation. The ability of hTAF II 28 to potentiate activation by Tax was dependent on its ability to interact with TBP. These observations support the notion that Tax activates transcription by interacting with TFIID through both TBP and hTAF II 28. MATERIALS AND METHODSTransfections. HeLa cells and COS-7 cells, grown in monolayers to 40% (HeLa cells) or 80% (COS-7 cells) confluence, were transfected by the calcium phosphate coprecipitation method (7). The total amount of simian virus 40 promotercontaining plasmids was adjusted to a constant level with pSG5, and the total amount of cytomegalovirus promotercontaining plasmids was adjusted to a constant level with pXJ41.