The Yeast TAF145 Inhibitory Domain and TFIIA Competitively Bind to TATA-Binding Protein

Abstract

The Drosophila 230-kDa TFIID subunit (dTAF230) interacts with the DNA binding domain of TATA boxbinding protein (TBP) which exists in the same complex. Here, we characterize the inhibitory domain in the yeast TAF145 (yTAF145), which is homologous to dTAF230. Mutation studies show that the N-terminal inhibitory region (residues 10 to 71) can be divided into two subdomains, I (residues 10 to 37) and II (residues 46 to 71). Mutations in either subdomain significantly impair function. Acidic residues in subdomain II are important for the interaction with TBP. In addition, yTAF145 interaction is impaired by mutating the basic residues on the convex surface of TBP, which are crucial for interaction with TFIIA. Consistently, TFIIA and yTAF145 bind competitively to TBP. A deletion of the inhibitory domain of yTAF145 leads to a temperaturesensitive growth phenotype. Importantly, this phenotype is suppressed by overexpression of the TFIIA subunits, indicating that the yTAF145 inhibitory domain is involved in TFIIA function.Transcription factor TFIID is a multisubunit protein complex found in various organisms including Drosophila melanogaster (17, 40), human (12,59,61,74), and more recently, the budding yeast Saccharomyces cerevisiae (23,(52)(53)(54). Holo-TFIID is composed of the highly conserved TATA box-binding polypeptide protein (TBP) and a number of associated polypeptides (TBP-associated factors [TAFs]). In vitro transcription studies revealed an important functional difference between holo-TFIID and TBP. Holo-TFIID mediates activator regulated transcription, whereas TBP itself mediates only basal levels of transcription. Thus, at least one or more TAFs included in holo-TFIID are essential for transmitting signals from various activators to the basal transcriptional machinery. Within the past 5 years, cDNAs encoding TAFs have been cloned from

A role of transcriptional activators as antirepressors for the autoinhibitory activity of TATA box binding of transcription factor IID

“…These results strongly suggest that this inhibitory activity could be intrinsic to TFIID and derived from TAF(s) and is sensitive to proteases. Consistent with this idea, Drosophila (d)TAF230, or the homologous yTAF145, inhibits TBP binding to the TATA box when these TAFs are mixed with TBP in vitro (16)(17)(18). Mutational analyses of dTAF230 indicate that the N-terminal 156 residues inhibit TATA box binding through direct interaction with TBP (19,20).…”

Mot1 activates and represses transcription by direct, ATPase-dependent mechanisms

“…3) and the interplay between TFIID, NC2, Mot1, TFIIA, and other TBPassociated transcription factors that stabilize or destabilize TBP binding to DNA is not well understood. TAF145 interacts with the DNA binding surface of TBP (40) as well as the convex surface of TBP in a region that overlaps with the interaction surface for TFIIA and Mot1 (35,36,41). Surprisingly, although biochemical and structural studies indicate that Mot1, TAF145, and TFIIA bind competitively to TBP, BNA1, URA1, and YDR539W transcription each depend on all three of these factors for full expression.…”

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

“…In many cases, activators increase the binding of TBP to promoters in direct proportion to their activity in transcription activation (Kuras and Struhl 1999;Li et al 1999). In contrast, factors such as Mot1 or the Taf1 N-terminal domain (TAND) inhibit the DNA binding activity of TBP (Auble et al 1994;Kokubo et al 1998;Cang et al 1999). The general factor TFIIA interacts directly with TBP and stabilizes TBP-DNA interactions (Weideman et al 1997;Hampsey 1998;Liu et al 1999).…”

“…TFIIA also stimulates and stabilizes the binding of TFIID to DNA as part of an activator-TFIID-TFIIA-DNA complex, a rate-limiting intermediate in the transcription of certain promoters (Wang et al 1992;Lieberman and Berk 1994;Chi et al 1995). 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).…”