Mechanism of synergy between TATA and initiator: synergistic binding of TFIID following a putative TFIIA-induced isomerization

Abstract

The TFIID complex interacts with at least three types of core promoter elements within protein-coding genes, including TATA, initiator (Inr), and downstream promoter elements. We have begun to explore the mechanism by which the TFIID-Inr interaction leads to functional synergy between TATA and Inr elements during both basal and activated transcription. In DNase I footprinting assays, GAL4-VP16 recruited TFIID-TFIIA to core promoters containing either a TATA box, an Inr, or both TATA and Inr elements, with synergistic interactions apparent on the TATA-Inr promoter. Appropriate spacing between the two elements was essential for the synergistic binding. Despite the sequence-specific TFIID-Inr interactions, gel shift experiments revealed that TFIID alone possesses similar affinities for the TATA-Inr and TATA promoters. Interestingly, however, recombinant TFIIA strongly and selectively enhanced TFIID binding to the TATA-Inr promoter, with little effect on binding to the TATA promoter. Studies of the natural adenovirus major late promoter confirmed these findings, despite the existence of specific but nonfunctional TFIID interactions downstream of the Inr in that promoter. These results suggest that a TFIIA-induced conformational change is essential for the sequence-specific TFIID-Inr interaction to occur with sufficient affinity to support the functional synergism between TATA and Inr elements.

Taspase1 processing alters TFIIA cofactor properties in the regulation of TFIID

“…12 Moreover, TAF interactions with the INR element enhance the promoter binding activity of isolated TFIID at TATA-containing promoters in a TFIIA-dependent manner. 13 These observations clearly establish an important role of TAFs in the functional readout of the INR element. However, TAF/promoter interactions are not sufficient to mediate the strong functional synergy of TATA and INR elements observed in crude nuclear extracts.…”

“…1 TFIIA stabilizes TBP/TATA complexes through interactions with TBP opposite of the binding site for TFIIB, and with promoter DNA upstream of TATA 20,36-39 and counteracts NC2 repression by competing for binding to TBP. 1 Importantly, TFIIA also promotes interactions between TFIID TAFs and core promoter DNA sequences downstream of the TATA region 13,19,20 and has been shown to be an essential cofactor for TAF-dependent INR function both in the absence and presence of a TATA element. 14 Consistent with these observations, results of site-specific DNA crosslinking studies provided first evidence that TFIIA affects the positioning of individual TFIID TAFs relative to downstream promoter DNA within TFIID nucleoprotein complexes.…”

“…27 Given the stringent conditions of the purification protocol, TFIIA in f:TFIID preparations must be tightly associated with the TFIID complex. Because TFIIA is required for synergistic binding of TFIID to TATA and INR core promoter elements 13 and for INR-dependent basal transcription in vitro, 14 we speculated that the subpopulation of TFIID complexes in our f:TFIID preparations that is resistant to NC2 repression in the presence of an INR element is stably associated with TFIIA. To further investigate this possibility, we carried out immunoblot analyses and identified two independent f:TFIID preparations (A) and (B), that differ significantly in TFIIA content.…”

“…1 TFIIA further modulates the promoter-binding activity of TFIID by stabilizing TBP/ TATA interactions 1 and by promoting interactions between TFIID TAFs and DNA sequences downstream of TATA, such as the INR element. 13,19,20 Indeed, TFIIA has been shown to be absolutely required for INR-mediated and TAF-dependent transcription initiation in the absence of a TATA box as well as for TAF-dependent INR-mediated stimulation of basal transcription from TATA-containing promoters. 14 TFIIA appears to modulate interactions of TAFs with specific core promoter DNA sequences by stabilizing specific TFIID complex conformations.…”

“…14 TFIIA appears to modulate interactions of TAFs with specific core promoter DNA sequences by stabilizing specific TFIID complex conformations. 13,20 Recent single particle electron studies of TFIID revealed that TFIID can exist in at least 2 structurally distinct states, a canonical state observed predominantly in the absence of DNA and a 'rearranged' state observed in the presence of promoter DNA. 21 Interestingly, these studies suggest a dual role of TFIIA in maintaining both the canonical state of TFIID in the absence of DNA and in stabilizing the re-arranged state of TFIID when promoterbound.…”