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J Biol Chem, Vol. 273, Issue 26, 16509-16516, June 26, 1998

Recruitment of Human TBP Selectively Activates RNA Polymerase II TATA-dependent Promoters

From the Department of Genetics, Molecular and General Biology, University of Naples "Federico II," via Mezzocannone 8, 80134 Naples, Italy and International Institute of Genetics and Biophysics, Consiglio Nazionale delle Ricerche, via Marconi 10, 80125 Naples, Italy

An increasing body of evidence suggests that eukaryotic activators stimulate polymerase II transcription by facilitating the assembly of the functional basal machinery at the promoter. Here we describe experiments that provide added support for the idea that recruitment of TATA-binding protein (TBP) is a rate-limiting step for transcription activation in mammalian cells. We found that, in human cell lines, recruitment of TBP to a promoter, as a GAL4-TBP fusion protein, can provide a substantial activation of transcription. Activation mediated by the hTBP, tethered to promoter DNA, is strictly dependent upon the presence of a functional TATA element, and it directs faithful transcription initiation. Interestingly, GAL4-hTBP activation was not observed from initiator (Inr) -dependent TATA-less promoters. These results suggest that TBP binding to DNA is not a rate-limiting step for the initial stages of TFIID recruitment to initiator-dependent TATA-less promoters. Finally, we provide evidence that synergy between GAL4-hTBP and defined transcription domains is restricted to activators, such as VP16 and Tat, which are likely to function at steps subsequent to the TFIID recruitment. These findings strengthen the idea that recruitment of TBP represents an important mechanism of activation of TATA-dependent promoters, and on the other hand, they suggest that TBP-DNA interactions are largely dispensable for specific transcription of initiator dependent TATA-less promoters.

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