TFIIA Interacts with TFIID via Association with TATA-Binding Protein and TAF40

Abstract

TFIIA and TATA-binding protein (TBP) associate directly at the TATA element of genes transcribed by RNA polymerase II. In vivo, TBP is complexed with approximately 14 TBP-associated factors (TAFs) to form the general transcription factor TFIID. How TFIIA and TFIID communicate is not well understood. We show that in addition to making direct contacts with TBP, yeast TAF40 interacts directly and specifically with TFIIA. Mutational analyses of the Toa2 subunit of TFIIA indicate that loss of functional interaction between TFIIA and TAF40 results in conditional growth phenotypes and defects in transcription. These results demonstrate that the TFIIA-TAF40 interaction is important in vivo and indicate a functional role for TAF40 as a bridging factor between TFIIA and TFIID.Transcription by eukaryotic RNA polymerase II (Pol II) involves the assembly of a preinitiation complex consisting of Pol II and the general transcription factors TFIIA, TFIIB, TFIID, TFIIE, TFIIF, and TFIIH (for review, see reference 66). An important step of transcription initiation is the binding of TFIID to the core promoter. TFIID, a multisubunit protein complex that is highly conserved among eukaryotes, is composed of the TATA-binding protein (TBP) and over a dozen TBP-associated factors (TAFs) (reviewed in references 26 and 27). TBP mediates promoter recognition through the sequence-specific binding of the TATA element found at many promoters. The importance of the TBP-TATA interaction is illustrated by many studies which demonstrate that recruitment of TBP is a rate-limiting step at a majority of promoters (12,17,37,39,46,50).In yeast, 13 TAFs are required for viability, indicating essential roles for individual TAFs (75, 77; reviewed in reference 26). However, the precise functional requirements for the TAFs remain unresolved. In vitro biochemical experiments suggest that TAFs function in higher eukaryotic systems as obligatory coactivators essential for activator response (reviewed in references 9 and 84). In contrast, functional inactivation and depletion studies with certain TAFs in yeast cells demonstrate that the expression of many genes are unaffected by TAF loss, although TAF inactivation results in distinct cell cycle phenotypes (2,55,59,60,62,86). In addition, disruption of the TFIID complex with a temperature-sensitive mutation in TBP results in gene-specific transcriptional defects (71), and promoter occupancy studies indicate that TAFs are not present on certain transcriptionally active promoters in vivo (45, 49). Promoter-specific requirements for particular TAFs are further illustrated by whole-genome transcriptional profiles. For example, inactivation of TAF145/130 has no effect on the expression of a majority of genes, while transcription of a subset of genes is affected (31). This TAF dependence was mapped to the core promoter (78), indicating important TAF functions in promoter activity in vivo. In contrast to these gene-specific effects, inactivation of several other TAFs, namely TAF17 (2, 59, 61), TAF40 (44), TAF60 and TA...

Positive and negative functions of the SAGA complex mediated through interaction of Spt8 with TBP and the N-terminal domain of TFIIA

“…Numerous other Flag-tagged nuclear extracts were also tested in the cross-linking assay, none of which displayed cross-linked polypeptides of shifted mobility. Proteins of note that did not cross-link to the TFIIA NTD include Taf11, previously shown to interact with the TFIIA NTD by two-hybrid assay (Kraemer et al 2001), and Spt3, which genetically interacts with Spt8 and TBP in the SAGA complex (Eisenmann et al 1994).…”

“…Later work also showed the importance of other nearby residues on this TFIIA NTD surface. Protein-protein interaction studies using the two-hybrid and GST-pulldown assays suggested that the TFIIA NTD interacts with the TFIID-specific subunit Taf11 (Kraemer et al 2001). This interaction is blocked by mutation of Toa2 residue I27, which also causes a defect in transcription from a subset of genes in vivo (Kraemer et al 2001).…”

“…Protein-protein interaction studies using the two-hybrid and GST-pulldown assays suggested that the TFIIA NTD interacts with the TFIID-specific subunit Taf11 (Kraemer et al 2001). This interaction is blocked by mutation of Toa2 residue I27, which also causes a defect in transcription from a subset of genes in vivo (Kraemer et al 2001). In other work, it was proposed that this TFIIA NTD surface is important for transcription activation because mutation of Toa2 residue G30 suppresses the TBP mutation P109A, which is defective in activation of specific promoters (Liu et al 1999).…”

“…Finally, TFIIA can compete with negative factors such as NC2, Mot1, and the Taf1 TAND domain for binding to TBP (Hampsey 1998;Kokubo et al 1998;Ozer et al 1998b;Cang et al 1999). TFIIA has also been found to interact with at least two Tafs and some transcription activators, using two-hybrid and affinity chromatography analysis (Yokomori et al 1993;Ozer et al 1994;Kobayashi et al 1998;Kraemer et al 2001).…”

Differential expression and interaction of transcription co-activator MBF1 with TATA-binding protein (TBP) in the apicomplexan Cryptosporidium parvum

“…The binding of TBP to the TATA-box that is located upstream of the 59 untranslated region (59-UTR) initiates the general downstream gene transcription machinery by recruiting a variety of transcription factors including TFIIA, B, E, F, H and J (Kraemer et al, 2001;Ouyang et al, 2000;Pugh, 2000;Vasanthi et al, 2003;Wu & Chiang, 2001). Most of these factors can be identified from the C. parvum genome (http://cryptodb.org/) (data not shown), suggesting that the general gene transcriptional machinery is conserved between this parasite and other eukaryotes.…”