Distinct subdomains of human TAFII130 are required for interactions with glutamine-rich transcriptional activators (original) (raw)
Related papers
Molecular and cellular biology, 1994
We have used protein-blotting and protein affinity chromatography to demonstrate that each of the two glutamine-rich activation domains of the human transcription factor Spl can bind specifically and directly to the C-terminal evolutionarily conserved domain of the human TATA box-binding protein (TBP). These activation domains of Spl also bind directly to Drosophila TBP but bind much less strongly to TBP from the yeast Saccharomyces cerevisiae. The abilities of the Spl activation domains to interact directly with the TBPs of various species correlate well with their abilities to activate transcription in extracts derived from the same species. We also show that a glutamine-rich transcriptional activating region of the Drosophila protein Antennapedia binds directly to TBP in a species-specific manner that reflects its ability to activate transcription in vivo. These results support the notion that TBP is a direct and important target of glutamine-rich transcriptional activators.
Molecular cloning and analysis of two subunits of the human TFIID complex: hTAFII130 and hTAFII100
Proceedings of the National Academy of Sciences, 1996
Transcription factor TFIID is a multiprotein complex composed of the TATA box-binding protein (TBP) and multiple TBP-associated factors (TAFs). TFIID plays an essential role in mediating transcriptional activation by genespecific activators. Numerous transcriptional activators have been characterized from mammalian cells; however, molecular analysis of the components of mammalian TFIID has been incomplete. Here we describe isolation of cDNAs encoding two TAF subunits of the human transcription factor TFIID. The first cDNA is predicted to encode the C-terminal 947 residues of the 130-kDa human TAF subunit, hTAF II 130. The second cDNA encodes the C-terminal 801 residues of the 100-kDa subunit, hTAF II 100.
Molecular and Cellular Biology, 2002
Surfaces of human TATA box-binding protein (hsTBP) required for activated transcription in vivo were defined by constructing a library of surface residue substitution mutations and assaying them for their ability to support activated transcription in transient-transfection assays. In earlier work, three regions were identified where mutations inhibited activated transcription without interfering with TATA box DNA binding. One region is on the upstream surface of the N-terminal TBP repeat with respect to the direction of transcription and corresponds to the TBP surface that interacts with TFIIA. A second region on the stirrup of the C-terminal TBP repeat corresponds to the TFIIB-binding surface. Here we report that the third region where mutations inhibit activated transcription in mammalian cells, the convex surface of the N-terminal repeat, corresponds to a surface on TBP that interacts with hsTAF1, the major scaffold subunit of TFIID. Since mutations at the center of the hsTAF1-in...
Genes & Development, 1993
TFIID is a multisubunit protein containing the TATA box-binding polypeptide (TBP) and associated factors (TFIID-TAFs) required for activated transcription by RNA polymerase II. TBPs from different eukaryotes contain a highly conserved carboxy-terminal domain and very divergent amino-terminal domains. Earlier studies proposed that the amino-terminal domains of metazoan TBPs are required for activated transcription. However, we report that a human TFIID complex containing an amino-terminal truncated TBP contains all the major TFIID-TAFs and supports in vitro transcriptional stimulation by different classes of activation domains and from a TATA-less promoter. Protein blotting experiments revealed direct interactions between the conserved domain of TBP and the two largest TAFs. The results suggest a model for the interaction of TFIID-TAFs with TBP.
Genes & Development, 1996
Regions on the surface of human TATA-box binding protein (TBP) required for activated transcription in vivo were defined by construction of a library of 89 surface residue mutants with radical substitutions that were assayed for their ability to support activated transcription in vivo, basal transcription in vitro, and TFIIA and TFIIB binding in vitro. Four epitopes were identified in which substitutions in two to four neighboring surface residues greatly inhibited activated transcription in vivo. One epitope in which substitutions inhibited both basal and activated transcription (E284, L287) is the interface between TBP and TFIIB. Another (A184, N189, E191, R205) is the recently determined interface between TBP and TFIIA. Mutations in residues in this TFIIA interface greatly inhibit activated, but not basal transcription, demonstrating a requirement for the TFIIA-TBP interaction for activated transcription in vivo in mammalian cells. The remaining two activation epitopes (TBP helix...
Core promoter-specific function of a mutant transcription factor TFIID defective in TATA-box binding
Proceedings of the National Academy of Sciences, 1995
In conjunction with other general initiation factors, the TATA box-binding protein (TBP) can direct basal transcription by RNA polymerase II from TATA-containing promoters, but its stable interaction with TBP-associated factors (TAFs) in the TFIID complex is required both for activator-dependent transcription and for basal transcription directed by an initiator element. We have generated a TATAbinding-defective TFIID complex containing an amino acid substitution in the DNA-binding surface of its TBP subunit. This mutated TFIID is defective in both basal and activated transcription from core promoters containing only a TATA box but supports transcription from initiator-containing promoters independently of the presence or absence of a TATA sequence. Our results show that a functional initiator element is needed to bypass the requirement for an active TATA DNA-binding surface in TFIID and imply that gene-specific transcription can be achieved by modulating distinct core promoter-specific TFIID functions-e.g., TBP-TATA versus TAF-initiator interactions.
Journal of Biological Chemistry, 2002
Initiation of transcription of protein-encoding genes by RNA polymerase II was thought to require transcription factor TFIID, a complex comprising the TATA-binding protein (TBP) and TBP-associated factors (TAFs). In the presence of TBP-free TAF complex (TFTC), initiation of polymerase II transcription can occur in the absence of TFIID. TFTC contains several subunits that have been shown to play the role of transcriptional coactivators, including the GCN5 histone acetyltransferase (HAT), which acetylates histone H3 in a nucleosomal context. Here we analyze the coactivator function of TFTC. We show direct physical interactions between TFTC and the two distinct activation regions (H1 and H2) of the VP16 activation domain, whereas the HATcontaining coactivators, p300/CBP (CREB-binding protein), interact only with the H2 subdomain of VP16. Accordingly, cell transfection experiments demonstrate the requirement of both p300 and TFTC for maximal transcriptional activation by GAL-VP16. In agreement with this finding, we show that in vitro on a chromatinized template human TFTC mediates the transcriptional activity of the VP16 activation domain in concert with p300 and in an acetyl-CoA-dependent manner. Thus, our results suggest that these two HAT-containing co-activators, p300 and TFTC, have complementary rather than redundant roles during the transcriptional activation process.
An inhibitor domain in Sp3 regulates its glutamine-rich activation domains
The EMBO journal, 1996
Sp3 is a ubiquitously expressed human transcription factor closely related to Sp1 and Sp4. All three proteins contain a highly conserved DNA binding domain and two glutamine-rich regions, suggesting that they possess similar activation functions. In our previous experiments, however, Sp3 failed to activate transcription. Instead, it repressed Sp1-mediated transcriptional activation, suggesting that it is an inhibitory member of this family of regulatory factors. Here we show that Sp3 can also act as a positive regulator of transcription. The glutamine-rich domains on their own have a strong activation function and interact with the TATA box binding protein (TBP)-associated factor dTAFII110. However, in full-length Sp3 as well as in Gal4-Sp3 fusion proteins, both activation domains are silenced by an inhibitory domain located between the second glutamine-rich region and the DNA binding domain. The inhibitory domain completely suppressed transcriptional activation when fused to a hete...
Autonomous Function of the Amino-Terminal Inhibitory Domain of TAF1 in Transcriptional Regulation
Molecular and Cellular Biology, 2004
The general transcription factor TFIID is composed of TATA-binding protein (TBP) and 14 TBP-associated factors (TAFs). TFIID mediates the transcriptional activation of a subset of eukaryotic promoters. The N-terminal domain (TAND) of TAF1 protein (Taf1p) inhibits TBP by binding to its concave and convex surfaces. This study examines the role of the TAND in transcriptional regulation and tests whether the TAND is an autonomous regulator of TBP. The TAND binds to and regulates TBP function when it is fused to the amino or carboxy terminus of Taf1p, the amino or carboxy terminus of Taf5p, or the amino terminus of Taf11p. However, a carboxy-terminal fusion of the TAND and Taf11p is not compatible with several other TAF proteins, including Taf1p, in the TFIID complex. These results indicate that there is no or minimal geometric constraint on the ability of the TAND to function normally in transcriptional regulation as long as TFIID assembly is secured.