Functional characterization of TFIID subunits (TAFs) in Saccharomyces cerevisiae

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The general transcription factor TFIID, which is composed of a TATA-binding protein (TBP) and a set of TBP-associated factors (TAFs), has been shown to be involved in both core promoter recognition and the transcriptional activation of eukaryotic genes. We isolated TAF145 temperature-sensitive mutants in yeast, in which transcription of the TUB2 gene is impaired at restrictive temperatures due to a defect in core promoter. We also showed in these mutants that the transcription of the RPS5 gene is impaired, mostly due to a defect in transcriptional activation rather than to a defect in core promoter recognition, although the latter is slightly affected as well. Surprisingly, the RPS5 core promoter can be activated by various activation domains fused to a GAL4 DNA binding domain (e.g. GAL4-RAP1, -VP16C, -EBNA2, and -GCN4), but not by the original upstream activating sequence (UAS) of the RPS5 gene. In addition, a heterologous CYC1 core promoter can be activated by RPS5-UAS at normal levels even in these mutants. These observations indicate that a distinct combination of core promoters and activators may exploit alternative activation pathways which vary in their requirement for TAF 145 function. In addition, a particular function of TAF 145 that is deleted in our mutants appears to be involved in both core promoter recognition and transcriptional activation