Biomedical subjects

M Horikoshi

Publications and source records attributed to M Horikoshi.

At least 109 records · Page 6Linked to original sources

PubMed1990

Highly conserved core domain and unique N terminus with presumptive regulatory motifs in a human TATA factor (TFIID).

The factor TFIID is one of several general factors that are necessary and sufficient for transcription initiation by mammalian RNA polymerase II. Stable interactions with the common TATA element lead both to template commitment and to the assembly of the other general factors into a functional preinitiation complex. Consistent with its key role in the promoter activation pathway, human TFIID also seems to be a target for some regulatory factors, as evidenced both by physical and functional studies of interactions between these components. The evolutionary conservation of functional properties led to the purification and cloning of yeast TFIID, the identification of presumptive structural motifs, and direct structure-function studies. Here we report the cloning of a complementary DNA encoding a functional human TFIID. This reveals an evolutionarily conserved core which corresponds precisely to the 180-residue DNA binding/activation domain determined for yeast TFIID, a near absolute conservation of component structural motifs (direct repeats, central basic core/lysine repeat, and sigma homology), providing further support for their functional importance, and a unique N-terminal structure that suggests involvement in species-specific regulatory factor interactions.

Amino Acid Sequence↗

PubMed1990

Arabidopsis thaliana contains two genes for TFIID.

The general transcription initiation factor TFIID plays a primary part in the activation of eukaryotic genes transcribed by RNA polymerase II. Binding of TFIID to the TATA box initiates the assembly of other general transcription factors as well as RNA polymerase II at the promoter resulting in a preinitiation complex capable of accurate transcription initiation in vitro. Human TFIID has been shown to interact with various regulatory factors. The observation that stimulation of transcription by different trans-acting factors is mediated through distinct TATA elements led to the suggestion that different types of TFIID may exist in yeast, humans and plants. Here we report the cloning and characterization of two distinct TFIID complementary DNA clones from Arabidopsis thaliana. Furthermore, we have found that TFIID from Arabidopsis and other organisms shows homology to helix-loop-helix proteins.

Amino Acid Sequence↗

PubMed1990

Analysis of structure-function relationships of yeast TATA box binding factor TFIID.

A systematic series of N-terminal, C-terminal, and internal deletion mutants of S. cerevisiae TFIID were expressed in vitro and tested for TATA box binding and basal level transcription activities using, respectively, DNA mobility shift and in vitro transcription assays. The domains responsible for these activities were colocalized to a surprisingly large region containing C-terminal residues 63-240. This region was noted previously to contain potentially interesting structural motifs (central basic core, direct repeats, and sigma factor homology) and, more recently, to be highly conserved among TFIID from different species. Deletion mutant cotranslation studies revealed that TFIID binds DNA as a monomer. The implications of these results for TFIID structure and function are discussed.

Base Sequence↗

PubMed1990

Binding site-dependent direct activation and repression of in vitro transcription by Drosophila homeodomain proteins.

Fushi tarazu and engrailed are two of the genes required for proper segmentation of the Drosophila embryo. Their protein products Fushi tarazu and Engrailed (Ftz and En) each contain a homeodomain and have been shown to act as transcriptional regulators in transient expression experiments in a Drosophila cell culture system. We used an in vitro transcription system to test whether the effects of Ftz and En on transcription were direct or indirect. Purified Ftz directly activates in vitro transcription by binding to homeodomain binding sites inserted upstream of the TATA box of the Drosophila hsp70 promoter. Equimolar amounts of purified En repress this activation by competition with Ftz for binding to these sites. These results indicate that Ftz and En act directly as transcription factors and suggest that such homeodomain proteins regulate development by combinatorial transcriptional control.

Animals↗

PubMed1990

cDNA clone encoding Drosophila transcription factor TFIID.

Proper initiation of transcription by RNA polymerase II requires the TATA-consensus-binding transcription factor TFIID. A cDNA clone encoding the Drosophila TFIID protein has been isolated and characterized. The deduced amino acid sequence reveals an open reading frame of 353 residues. The carboxyl-terminal 180 amino acids are approximately 80% identical to yeast TFIID and 88% identical to human TFIID. The amino-terminal portions of the yeast and Drosophila TFIID proteins lack appreciable homology, whereas the Drosophila and human amino termini appear qualitatively similar. In addition, the amino-terminal region of the Drosophila TFIID contains several sequence motifs that are found in other Drosophila proteins which appear to regulate transcription.

Amino Acid Sequence↗

PubMed1990

Recombinant yeast TFIID, a general transcription factor, mediates activation by the gene-specific factor USF in a chromatin assembly assay.

The TATA box-binding transcription factor TFIID from Saccharomyces cerevisiae was tested for its ability to mediate regulatory factor functions both in a cell-free system reconstituted with other general initiation factors (purified from HeLa cells) and in a combined nucleosome assembly-transcription system. In the latter assay recombinant yeast TFIID, expressed in and purified from bacteria, was sufficient to prevent nucleosome assembly-mediated repression and to mediate transcriptional enhancement of the adenovirus major late promoter by the gene-specific activator USF. In contrast, recombinant yeast TFIID was unable to mediate activation by USF in the system reconstituted only with purified general factors. Under the same conditions a partially purified natural yeast TFIID was able to mediate activation by both USF and Sp1 (assayed with the human immunodeficiency virus promoter), but to a lesser extent than observed with a partially purified natural human TFIID. The implications of these findings are discussed with respect to the structure of the yeast and human TATA factors and the possible involvement either of specific TFIID modifications or of coactivators.

Animals↗

PubMed1990

Factors involved in specific transcription by mammalian RNA polymerase II: identification of general transcription factor TFIIG.

We have identified and partially characterized another human general transcription factor, TFIIG. Using a reconstituted in vitro system comprised of purified RNA polymerase II, TFIIB, TFIID, TFIIE, and TFIIF, we found that TFIIG was essential for specific initiation from all class II genes tested. In this system TFIIA could partially replace TFIIG; however, even at saturating concentrations of TFIIA, addition of TFIIG further stimulated transcription. Since the chromatographic properties of TFIIG differed significantly from those of TFIIA, we concluded that TFIIA and TFIIG are distinct but functionally related transcription factors. Heparin challenge assays showed that TFIIG is required for the assembly of a functional preinitiation complex. However, it must act after template commitment by TFIID, since this step did not require, and was unaffected by, either TFIIG or TFIIA.

Cell Nucleus↗

PubMed1990

Factors involved in specific transcription by mammalian RNA polymerase II: purification and characterization of general transcription factor TFIIE.

Human transcription factor TFIIE, a ubiquitous factor required for transcription initiation by RNA polymerase II, was purified to homogeneity by a combination of conventional and HPLC steps. The purified TFIIE contained equimolar amounts of 57-kDa (TFIIE-alpha) and 34-kDa (TFIIE-beta) polypeptides that were judged to be functional subunits on the basis of their copurification with transcriptional activity and the recovery of activity following renaturation of polypeptides separated by reverse-phase HPLC. TFIIE-alpha had an independent TFIIE activity whereas TFIIE-beta had no activity alone but enhanced the activity of TFIIE-alpha. In conjunction with gel filtration studies, which indicated a molecular mass of approximately 180 kDa for the native protein, these results suggested that TFIIE is a heterotetramer containing two alpha and two beta polypeptides. Functional studies with the purified TFIIE demonstrated that it is a general initiation factor, required for all of the genes tested, but it failed to show any DNA-dependent ATPase activity.

Cell Nucleus↗

PubMed1990

Engrailed, a homeodomain protein, can repress in vitro transcription by competition with the TATA box-binding protein transcription factor IID.

Engrailed (En) is a homeodomain protein that binds to a consensus sequence (NP) and plays an important role during Drosophila development. Purified En, which is produced in Escherichia coli, binds not only to this consensus sequence but also to the TATA box of the Drosophila Hsp70 promoter and of other eukaryotic promoters. Interestingly, En represses transcription of these promoters in an in vitro-reconstituted mammalian transcription system and footprint analyses show that En competes with the TATA box-binding protein transcription factor IID for binding to the TATA box. In contrast, a stable template-committed complex formed by preincubation of transcription factor IID with the promoter is not disrupted by addition of En, and in this case transcription is not repressed. These in vitro studies suggest a transcriptional repression mechanism, involving competition between En and transcription factor IID for TATA box binding, that may be involved in En-mediated repression in vivo.

Animals↗

PubMed1990

A plant DNA-binding protein increases the number of active preinitiation complexes in a human in vitro transcription system.

TGA1a is a tobacco DNA-binding protein that binds to the activation sequence-1 (as-1) element of the cauliflower mosaic virus 35S promoter. We have produced TGA1a in Escherichia coli, purified it from bacterial extracts, and examined its effect on transcription in a human in vitro system. Addition of TGA1a stimulates transcription by up to 20 times, and the stimulation is dependent on the presence of the as-1 element in the promoter. When transcription reinitiation is inhibited by 0.3 M KCl, activation is similar. Therefore, TGA1a activates transcription by increasing the number of active preinitiation complexes. After formation of the preinitiation complexes in the presence of TGA1a, oligonucleotides containing TGA1a-binding sites do not significantly affect the stimulated level of transcription. This result indicates that a complex remains committed to the promoter site after initiation and that this complex is used repeatedly during several initiation events. Our study demonstrates for the first time that a plant factor can activate transcription in a human in vitro system and that the activation mechanism of the plant factor is similar to that of mammalian factors.

Amino Acid Sequence↗

PubMed1990

Cloning of the Schizosaccharomyces pombe TFIID gene reveals a strong conservation of functional domains present in Saccharomyces cerevisiae TFIID.

The gene encoding the Schizosaccharomyces pombe TATA box-binding factor (TFIID) was cloned and sequenced. The gene contains three introns and codes for a polypeptide of 231 amino acids. The cDNA-expressed protein showed both TATA box-binding and basal transcription activities. The carboxy-terminal three-quarters of S. pombe TFIID shares an extraordinary degree of amino acid sequence homology with a corresponding region of Saccharomyces cerevisiae TFIID that has been shown to be necessary and sufficient for TATA box-binding and basal transcription activities. In contrast, the amino-terminal regions of the S. pombe and S. cerevisiae TFIIDs differ markedly in amino acid sequence and composition. Structure and function relationships of TFIID are discussed in light of these data.

Amino Acid Sequence↗

PubMed1989

Cloning and structure of a yeast gene encoding a general transcription initiation factor TFIID that binds to the TATA box.

The TATA sequence-binding factor TFIID plays a central role both in promoter activation by RNA polymerase II and other common initiation factors, and in promoter regulation by gene-specific factors. The sequence of yeast TFIID, which seems to be encoded by a single gene, contains interesting structural motifs that are possibly involved in these functions, and is similar to sequences of bacterial sigma factors.

Amino Acid Sequence↗

PubMed1989

Purification of a yeast TATA box-binding protein that exhibits human transcription factor IID activity.

By a series of conventional chromatographic procedures we have purified from whole-cell extracts of Saccharomyces cerevisiae yeast transcription factor IID (TFIID), which functionally substitutes for human TFIID in a complementation assay comprised of the adenovirus type 2 major late promoter and HeLa cell-derived RNA polymerase II, transcription factors IIA, IIB, and IIE. Similar to its human counterpart, yeast TFIID also exhibited specific binding to the adenovirus type 2 major late promoter TATA element, as shown by both DNase I footprinting and gel mobility shift assays. NaDodSO4/PAGE analyses showed that a 27-kDa polypeptide coeluted with TFIID complementing activity through each chromatographic step. In agreement with this result and also suggesting that the native protein is a monomer, gel-filtration experiments indicated a molecular mass of 28 kDa for TFIID under nondenaturing conditions. That the 27-kDa polypeptide represented TFIID was further demonstrated by the ability of an HPLC-purified protein to bind specifically after renaturation to the adenovirus type 2 major late promoter TATA sequence.

Antigens, Neoplasm↗

PubMed1989

A study of an in vitro model for invasion of oral squamous cell carcinoma.

An in vitro model for studying the invasion mechanism of oral squamous cell carcinoma (SCC) was developed using a specific culture matrix composed of collagen gel combined with human fibroblasts. Five SCC cell lines cultured on collagen-only gels showed stratified growth on the gels. However, all five cell lines showed invasive growth into the matrix when cultured on the fibroblast-incorporated collagen gels. Moreover, fibroblast-conditioned medium was shown to promote the invasion of HSC-3 cells into the collagen gels. These results suggest that fibroblasts play an important role in the invasion of oral SCC cells in vitro. Four cell lines, which were newly established in our laboratory, were tested in this assay system. These cell lines cultured on fibroblast-incorporated collagen gels expressed morphologic and biologic characteristics in vitro, similar to those in vivo.

Carcinoma, Squamous Cell↗

PubMed1989

Transcriptionally active immediate-early protein of pseudorabies virus binds to specific sites on class II gene promoters.

In the presence of partially purified pseudorabies virus immediate-early protein, multiple sites of DNase I protection were observed on the adenovirus major late and human hsp 70 promoters. Southwestern (DNA-protein blot) analysis demonstrated that the immediate-early protein bound directly to the sequences contained in these sites. These sequences share only limited homology, differ in their affinities for the immediate-early protein, and are located at different positions on these two promoters. In addition, the site-specific binding of a temperature-sensitive immediate-early protein was eliminated by the same heat treatment which eliminates its transcriptional activating function, whereas the binding of the wild-type protein was unaffected by heat treatment. Thus, site-specific binding requires a functionally active immediate-early protein. Furthermore, immediate-early-protein-dependent in vitro transcription from the major late promoter was preferentially inhibited by oligonucleotides which are homologous to the high-affinity binding sites on the major late or hsp 70 promoters. These observations suggest that transcriptional stimulation by the immediate-early protein involves binding to cis-acting elements.

Base Sequence↗

PubMed1989

Purification and characterization of multiple nuclear factors that bind to the TAX-inducible enhancer within the human T-cell leukemia virus type 1 long terminal repeat.

Within the human T-cell leukemia virus type I promoter, there are three copies of a 21-base-pair repeat (hereafter called the tax-responsive element [TRE]) that both contributes to basal promoter activity and mediates induction by the viral activator TAX. We have identified and separated three nuclear proteins that interact with the TRE. The TRE-binding protein designated TREB-3 bound more avidly to a multimerized TRE than to a single-copy TRE, while the other two TRE-binding proteins, TREB-1 and TREB-2, bound equally well to either TRE. TREB-1 has been purified to near homogeneity, and binding activity was localized to a protein of 35 to 43 kilodaltons. The affinity-purified TREB-1 activated transcription from the human T-cell leukemia virus type I promoter in vitro. The purified TREB-1 fraction contained activating transcription factor binding activity and showed a cooperative interaction with the TATA-binding factor (TFIID) on the adenovirus E4 promoter.

Base Sequence↗

PubMed1989

[Radiographic findings of the so-called globulomaxillary cyst].

In definite diagnosis of this cyst, it is deemed that a comprehensive diagnosis is required including operative findings upon its extraction and the pathohistological findings of the extracted matter in addition to the radiographic and clinical findings, such as one that the region of development is between the maxillary lateral incisor and the cuspid, one that the adjacent teeth are vital ones, etc. Thus, we examined 10 cases that we had defined as the subjects of so-called Globulomaxillary cyst by clinical and pathohistological findings in terms of the developmental region, size, border, etc. As a result, the following things turned out: 1. The median positions of cystic development existed 8 cases in left side and 2 cases in right side out of 10 cases. And this median positions could be divided into those between the central incisor and the lateral incisor (5 cases) and those between the lateral incisor and the cuspid (5 cases). 2. The pathohistological findings led to the following classification; 2 cases with no inflammation, 5 cases with inflammation, and 3 cases with present inflammation in the past. The border line of cyst on radiographic findings tended to be relatively unclear in the cases with inflammation in the past.

Cuspid↗

PubMed1988

Stimulation of transcription from accurate initiation sites by purified S-II.

The effects of transcription factors S-II and S-II', a phosphorylated form of S-II, on accurate transcription were compared in a reconstituted transcription system greatly depleted of S-II. S-II, but not S-II', stimulated the syntheses of run-off products of various truncated class II genes in this system, suggesting that the activity of this factor is regulated by its phosphorylation and dephosphorylation.

Adenoviruses, Human↗