Structure of the central core domain of TFIIEbeta with a novel double-stranded DNA-binding surface

Okuda, Masahiko

doi:10.1093/emboj/19.6.1346

Cited by 58 publications

(63 citation statements)

References 73 publications

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“…7 and 8). These results, together with those of protein-DNA cross-linking experiments (49) and our recent structural study of the hTFIIE␤ core dsDNA-binding domain (43), strongly suggest the following model for the different functional roles of each subunit. TFIIE␤ may play a fundamental role: it is located inside the PIC and makes contact with the promoter region at the point where the dsDNA starts to open (between positions Ϫ14 and Ϫ2 with the transcription start site defined as ϩ1).…”

Section: Discussionsupporting

confidence: 65%

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Studies of Nematode TFIIE Function Reveal a Link between Ser-5 Phosphorylation of RNA Polymerase II and the Transition from Transcription Initiation to Elongation

Yamamoto

Watanabe

Spek

et al. 2001

Molecular and Cellular Biology

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The general transcription factor TFIIE plays important roles in transcription initiation and in the transition to elongation. However, little is known about its function during these steps. Here we demonstrate for the first time that TFIIH-mediated phosphorylation of RNA polymerase II (Pol II) is essential for the transition to elongation. This phosphorylation occurs at serine position 5 (Ser-5) of the carboxy-terminal domain (CTD) heptapeptide sequence of the largest subunit of Pol II. In a human in vitro transcription system with a supercoiled template, this process was studied using a human TFIIE (hTFIIE) homolog from Caenorhabditis elegans (ceTFIIE␣ and ceTFIIE␤). ceTFIIE␤ could partially replace hTFIIE␤, whereas ceTFIIE␣ could not replace hTFIIE␣. We present the studies of TFIIE binding to general transcription factors and the effects of subunit substitution on CTD phosphorylation. As a result, ceTFIIE␣ did not bind tightly to hTFIIE␤, and ceTFIIE␤ showed a similar profile for binding to its human counterpart and supported an intermediate level of CTD phosphorylation. Using antibodies against phosphorylated serine at either Ser-2 or Ser-5 of the CTD, we found that ceTFIIE␤ induced Ser-5 phosphorylation very little but induced Ser-2 phosphorylation normally, in contrast to wild-type hTFIIE, which induced phosphorylation at both Ser-2 and Ser-5. In transcription transition assays using a linear template, ceTFIIE␤ was markedly defective in its ability to support the transition to elongation. These observations provide evidence of TFIIE involvement in the transition and suggest that Ser-5 phosphorylation is essential for Pol II to be in the processive elongation form.In eukaryotes, transcription of protein-encoding genes by RNA polymerase II (Pol II) is the first step in expression of those genes (for reviews, see references 4, 35, 44, and 51). Two sequential stages are now recognized in the establishment of Pol II processivity: transcription initiation and the transition from initiation to elongation. At initiation, five general transcription factors (TFIIB, TFIID, TFIIE, TFIIF, and TFIIH) together with Pol II form the preinitiation complex (PIC) on the core promoter. Two models of PIC formation have been proposed on the basis of recent analyses. One model involves stepwise association of the general transcription factors and Pol II on promoter DNA, while the other model entails promoter sequences binding to a preassembled Pol II holoenzyme that contains most of the general transcription factors as well as SRB (suppressor of RNA polymerase B)-and Med-containing complex (reviewed in references 3 and 22). In vitro analyses of stepwise assembly of the PIC using purified factors have demonstrated that TFIIE joins the complex at a position near the transcription start site (between positions Ϫ14 and Ϫ2), after Pol II and TFIIF have joined the complex (25, 49). TFIIE then recruits TFIIH, and these two factors stabilize and activate the PIC, resulting in isomerization of double-stranded (ds) promoter DNA (promoter me...

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Section: Discussionsupporting

confidence: 65%

“…It has been found that TFIIE␤ binds to single-stranded (ss) DNA through the basic region near its C terminus; the other general transcription factors, TFIIB and TFIIF␤ (RAP30), bind to this region as well (42). In addition, we have recently determined the three-dimensional structure of the central core region in TFIIE␤ that binds to dsDNA (43).…”

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confidence: 99%

Studies of Nematode TFIIE Function Reveal a Link between Ser-5 Phosphorylation of RNA Polymerase II and the Transition from Transcription Initiation to Elongation

Yamamoto

Watanabe

Spek

et al. 2001

Molecular and Cellular Biology

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“…The winged-helix domains of hRAP30c and hTFIIE␤m can participate in nonspecific DNA binding, although they do not appear to use the same surface regions for these interactions (17,38). In the case of hRAP30c, the DNA interaction sites as judged by NMR are qualitatively similar to those observed by x-ray crystallography for sequence-specific DNA recognition by HNF-3␥ (33).…”

Section: Resultsmentioning

confidence: 87%

“…Winged-helix proteins in the pol II transcription initiation complex also include the Cterminal domain of the RAP30 subunit of human TFIIF (hRAP30c) (17) and the middle domain of the ␤ subunit of human TFIIE (hTFIIE␤m) (38). Our hRAP74cc crystal structure provides a third example of a winged-helix motif involved in transcription initiation by pol II.…”

Section: Resultsmentioning

confidence: 99%

Crystal structure of the C-terminal domain of the RAP74 subunit of human transcription factor IIF

Kamada

Angelis

Roeder

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

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The x-ray structure of a C-terminal fragment of the RAP74 subunit of human transcription factor (TF) IIF has been determined at 1.02-Å resolution. The ␣͞␤ structure is strikingly similar to the globular domain of linker histone H5 and the DNA-binding domain of hepatocyte nuclear factor 3␥ (HNF-3␥), making it a winged-helix protein. The surface electrostatic properties of this compact domain differ significantly from those of bona fide winged-helix transcription factors (HNF-3␥ and RFX1) and from the winged-helix domains found within the RAP30 subunit of TFIIF and the ␤ subunit of TFIIE. RAP74 has been shown to interact with the TFIIF-associated C-terminal domain phosphatase FCP1, and a putative phosphatase binding site has been identified within the RAP74 wingedhelix domain.T ranscription of class II nuclear genes in eukaryotes requires a complex assembly of proteins composed of a multisubunit RNA polymerase II (pol II) and general transcription factors (TFIIs), which are required for assembly of the preinitiation complex (PIC) on promoter DNA and initiation of transcription (1). In the most general case, PIC assembly begins with the TATA box-binding protein (TBP) subunit of TFIID recognizing the TATA element, followed by coordinated accretion of TFIIB, the nonphosphorylated form of pol II plus TFIIF, TFIIE, and TFIIH. Transcription initiation is followed by phosphorylation of the C-terminal domain (CTD) of the largest subunit of pol II, which in turn facilitates promoter clearance and productive elongation. After elongation and termination of pre-mRNA synthesis, a phosphatase recycles pol II to its nonphosphorylated form, allowing the enzyme and the requisite TFIIs to initiate the next round of transcription.Mammalian TFIIF is an ␣␤ hetero-oligomer of RAP74 and RAP30 subunits (RNA polymerase II-associating proteins; 58 kDa and 28 kDa, respectively) (2), which binds to pol II in solution and facilitates delivery of the polymerase to the TFIID-TFIIB-DNA complex on the promoter. TFIIF was originally identified on the basis of physical association with pol II, and its requirement for promoter-specific transcription initiation by this large multisubunit enzyme. Within the PIC, TFIIF stabilizes binding of pol II to TFIID and TFIIB at the promoter (3-5). Moreover, TFIIF is required for entry of TFIIE and TFIIH into the PIC, for subsequent open complex formation catalyzed by the DNA helicase subunit of TFIIH (5-7), and for synthesis of the first phosphodiester bond of nascent transcripts (8-10). It is also possible that TFIIF acts as a scaffold for binding of TFIIH to the growing PIC and͞or that TFIIF actively participates in formation of the open complex. Photocrosslinking experiments suggest that TFIIF interacts with promoter DNA to induce a dramatic conformational change that results in wrapping of DNA around pol II and the class II general transcription factors within the PIC (11-14).Biochemical properties of TFIIF have been attributed to distinct polypeptide chain segments of each of subunit. Mammalian RAP30 can b...

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“…Based on our observations (i) that, like TFIIH action in formation of the open complex, TFIIH action in promoter escape by RNA polymerase II requires the XPB DNA helicase activity, an ATP(dATP) cofactor, and downstream DNA and (ii) that the requirement for the TFIIH XPB DNA helicase, an ATP(dATP) cofactor, and downstream DNA in transcription initiation and promoter escape by RNA polymerase II is lost when transcription is carried out with the Ad(Ϫ9͞ϩ9) template, which contains a premelted region extending to ϩ9, it is reasonable to propose that TFIIH XPB helicase could suppress arrest simply by unwinding promoter DNA downstream of the transcriptional start site. Indeed, the regions of the TFIIF RAP30 and TFIIE small subunits required for transcription initiation by RNA polymerase II in vitro have been shown to include double-stranded DNA binding domains (42)(43)(44); thus, unwinding of downstream promoter DNA by the TFIIH XPB DNA helicase could relieve a TFIIF-and TFIIE-induced impediment to promoter escape by disrupting interactions of TFIIF and TFIIE with double-stranded DNA downstream of the transcriptional start site. Further experiments will be required, however, to test this hypothesis.…”

Section: Discussionmentioning

confidence: 99%

TFIIH action in transcription initiation and promoter escape requires distinct regions of downstream promoter DNA

Spangler

Wang

Conaway

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

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TFIIH is a multifunctional RNA polymerase II general initiation factor that includes two DNA helicases encoded by the Xeroderma pigmentosum complementation group B (XPB) and D (XPD) genes and a cyclin-dependent protein kinase encoded by the CDK7 gene. Previous studies have shown that the TFIIH XPB DNA helicase plays critical roles not only in transcription initiation, where it catalyzes ATP-dependent formation of the open complex, but also in efficient promoter escape, where it suppresses arrest of very early RNA polymerase II elongation intermediates. In this report, we present evidence that ATP-dependent TFIIH action in transcription initiation and promoter escape requires distinct regions of the DNA template; these regions are well separated from the promoter region unwound by the XPB DNA helicase and extend, respectively, Ϸ23-39 and Ϸ39 -50 bp downstream from the transcriptional start site. Taken together, our findings bring to light a role for promoter DNA in TFIIH action and are consistent with the model that TFIIH translocates along promoter DNA ahead of the RNA polymerase II elongation complex until polymerase has escaped the promoter.T FIIH is a multifunctional RNA polymerase II general initiation factor that includes two DNA helicases encoded by the Xeroderma pigmentosum complementation group B (XPB) and D (XPD) genes, as well as a cyclin-dependent protein kinase encoded by the CDK7 gene (1). Previous studies have shown that the TFIIH XPB DNA helicase functions at multiple steps to promote efficient transcription initiation and promoter escape by RNA polymerase II. The TFIIH XPB DNA helicase catalyzes ATP(dATP)-dependent formation of the open complex before synthesis of the first phosphodiester bond of nascent transcripts (2), and it is required to suppress premature arrest of very early RNA polymerase II elongation intermediates at promoterproximal sites Ϸ10-12 bp downstream of the transcriptional start site before their escape from the promoter (3-5).In a previous study, we identified a requirement in transcription initiation and promoter escape by RNA polymerase II for promoter DNA extending Ϸ23-50 bp downstream from the transcriptional start site (6). In that study, we showed that synthesis of the first phosphodiester bond of nascent transcripts by RNA polymerase II requires promoter DNA extending Ϸ23-39 bp downstream from the transcriptional start site and that efficient promoter escape by the enzyme requires promoter DNA extending Ϸ39-50 bp downstream from the transcriptional start site. That study, however, did not identify which of the general initiation factors require downstream promoter DNA during these stages of transcription.In this report, we present direct biochemical evidence that TFIIH requires downstream promoter DNA for its action in transcription initiation and promoter escape by RNA polymerase II. In addition, we show that TFIIH function in synthesis of the first phosphodiester bond of nascent transcripts and in promoter escape requires distinct regions of DNA downstream of the tra...

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Structure of the central core domain of TFIIEbeta with a novel double-stranded DNA-binding surface

Cited by 58 publications

References 73 publications

Studies of Nematode TFIIE Function Reveal a Link between Ser-5 Phosphorylation of RNA Polymerase II and the Transition from Transcription Initiation to Elongation

Studies of Nematode TFIIE Function Reveal a Link between Ser-5 Phosphorylation of RNA Polymerase II and the Transition from Transcription Initiation to Elongation

Crystal structure of the C-terminal domain of the RAP74 subunit of human transcription factor IIF

TFIIH action in transcription initiation and promoter escape requires distinct regions of downstream promoter DNA

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