Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “transcript”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,513 records · Page 84Linked to original sources
PubMed2005

The transcriptional coactivator PC4/Sub1 has multiple functions in RNA polymerase II transcription.

Transcription and processing of mRNA precursors are coordinated events that require numerous complex interactions to ensure that they are successfully executed. We described previously an unexpected association between a transcription factor, PC4 (or Sub1 in yeast), and an mRNA polyadenylation factor, CstF-64 (Rna15 in yeast), and provided evidence that this was important for efficient transcription elongation. Here we provide insight into the mechanism by which this occurs. We show that Sub1 and Rna15 are recruited to promoters and present along the length of several yeast genes. Allele-specific genetic interactions between SUB1 and genes encoding an RNA polymerase II (RNAP II)-specific kinase (KIN28) and phosphatase (FCP1) suggest that Sub1 influences and/or is sensitive to the phosphorylation status of elongating RNAP II. Remarkably, we find that cells lacking Sub1 display decreased accumulation of Fcp1, altered RNAP II phosphorylation and decreased crosslinking of RNAP II to transcribed genes. Our data provide evidence that Rna15 and Sub1 are present along the length of several genes and that Sub1 facilitates elongation by influencing enzymes that modify RNAP II.

Chromatin↗
PubMed2000

AP-2 transcription factors in the regulation of ERBB2 gene transcription by oestrogen.

Transcription of the ERBB2 oncogene is repressed by oestrogen in human breast cancer cells. We show that a 218 bp fragment of the human ERBB2 gene promoter is responsive to oestrogen in transient transfection in ZR75.1 and SKBR.3 cells when the oestrogen receptor is expressed. Deletion analysis of this fragment shows that a sequence located at the 5' end, which is known to mediate ERBB2 overexpression in breast cancer, is also responsible for the oestrogen response. This sequence binds AP-2 transcription factors and appears functionally identical to an element of the oestrogen-dependent enhancer described in the first intron of human ERBB2. We observed that oestrogen treatment down-regulates expression of AP-2 proteins but does not affect the DNA binding activity of AP-2. Constitutive expression of AP-2beta or AP-2gamma, but not AP-2alpha, abrogates the estrogenic repression. Our results demonstrate that AP-2 transcription factors are implicated in the oestrogenic regulation of ERBB2 gene expression and suggest a complex interplay involving the different AP-2 isoforms and other unidentified factors.

Breast Neoplasms↗
PubMed2003

CDX2, a homeobox transcription factor, upregulates transcription of the p21/WAF1/CIP1 gene.

The CDX2 homeobox transcription factor plays key roles in intestinal development and homeostasis. CDX2 is downregulated during colorectal carcinogenesis, whereas overexpression of CDX2 results in growth inhibition and differentiation of colon carcinoma and intestinal cells. However, the means by which CDX2 functions remain poorly understood. p21/WAF1/CIP1 is one of the cyclin-dependent kinase inhibitors. In addition to its role in cell cycle control, p21 plays critical roles in differentiation and tumor suppression. The overlapping in both the expression and function of CDX2 and p21 in the small intestine and colon strongly suggests a link between these two genes. By means of luciferase reporter and electrophoretic mobility shift assays, we show here that CDX2 transactivated and physically interacted with the promoter of p21 in a p53-independent manner. Moreover, overexpression of CDX2 increased the mRNA expression of p21 in HT-29 colon carcinoma cells, as demonstrated by reverse transcription-polymerase chain reaction. These data suggest that p21 is a transcriptional target of CDX2. Our results may thus provide a new mechanism underlying the functions of CDX2.

Animals↗
PubMed2001

Transcriptional regulation of the human cystathionine beta-synthase -1b basal promoter: synergistic transactivation by transcription factors NF-Y and Sp1/Sp3.

Cystathionine beta-synthase (CBS) catalyses the condensation of serine and homocysteine to form cystathionine, an intermediate step in the synthesis of cysteine. Human CBS encodes five distinct 5' non-coding exons, the most frequent termed CBS -1a and CBS -1b, each transcribed from its own unique GC-rich TATA-less promoter. The minimal transcriptional region (-3792 to -3667) of the CBS -1b promoter was defined by 5'- and 3'-deletions, and transient transfections of reporter gene constructs in HepG2 cells, characterized by CBS transcription exclusively from the -1b promoter. Included in this 125 bp region are 3 GC-boxes (termed GC-a, GC-b and GC-c), an inverted CAAT-box and an E-box. By gel-shift and supershift assays, binding of specificity protein (Sp)1 and Sp3 to the GC-box elements, upstream stimulatory factor 1 (USF-1) to the E-box, and both nuclear factor (NF)-Y and an NF-1-like factor to the CAAT box could be demonstrated. By transient trans fections and reporter gene assays in HepG2 and Drosophila SL2 cells, a functional interplay was indicated between NF-Y binding to the CAAT-box, or between USF-1 binding to the E-box, and Sp1/Sp3 binding to the GC-box elements. In SL2 cells, NF-Y and Sp1/Sp3 were synergistic. Furthermore, both Sp1 and the long Sp3 isoform transactivated the CBS -1b minimal promoter; however, the short Sp3 isoforms were potent repressors. These results may explain the cell- or tissue-specific regulation of CBS transcription, and clarify the bases for alterations in CBS gene expression in human disease and Down's syndrome.

Base Sequence↗
PubMed2006

Structural studies on the RNA-recognition motif of NELF E, a cellular negative transcription elongation factor involved in the regulation of HIV transcription.

The elongation of transcription of HIV RNA at the TAR (transactivation-response element) is highly regulated by positive and negative factors. The cellular negative transcription elongation factor NELF (negative elongation factor) was suggested to be involved in transcriptional regulation of HIV-1 (HIV type 1) by binding to the stem of the viral TAR RNA which is synthesized by cellular RNA polymerase II at the viral long terminal repeat. NELF is a heterotetrameric protein consisting of NELF A, B, C or the splice variant D, and E. In the present study, we determined the solution structure of the RRM (RNA-recognition motif) of the RNA-binding subunit NELF E and studied its interaction with the viral TAR RNA. Our results show that the separately expressed recombinant NELF E RRM has alpha-helical and beta-strand elements adopting a betaalphabetabetaalphabeta fold and is able to bind to TAR RNA. Fluorescence equilibrium titrations with fluorescently labelled double- and single-stranded oligoribonucleotides representing the TAR RNA stem imply that NELF E RRM binds to the single-stranded TAR RNAs with K(d) values in the low-micromolar range.

Amino Acid Motifs↗
PubMed1996

HALF-1, a bZIP-type protein, interacting with the wheat transcription factor HBP-1a contains a novel transcriptional activation domain.

BACKGROUND: Nuclear factors bind to cis-acting elements and mediate transcriptional regulation through protein-protein interactions with other factors. The bZIP-type wheat nuclear protein HBP-1a(17) is a putative transcriptional activator specifically binding to the Hex (ccACGTCA) and G-box (CCACGTGG) motifs, which are often found in the cis-acting elements critical for various responses in plants. RESULTS: In order to investigate the mechanisms for gene expression mediated via the Hex and G-box motifs, we attempted to isolate proteins interacting with HBP-1a(17) based on protein-protein interactions. A cDNA expression library from wheat seedlings was screened with 32P-labelled HBP-1a(17), and a bZIP-type protein, termed HALF-1 (HBP-1-associated leucine-zipper factor-1), was isolated. GST-pulldown assay, yeast two-hybrid system and EMSA showed that HALF-1 and HBP-1a(17) interact with each other through their leucine-zipper regions. Dissection experiments showed that HALF-1 has at least one potential trans-activation domain which includes a nine amino acid motif conserved between several plant bZIP-type proteins. This motif, named GCB (GBF-conserved box; consensus, NLNIGMDXW), activated the expression of a reporter gene, when fused to the GAL4 DNA-binding domain. The corresponding region of Arabidopsis GBF1 also stimulated transcription. However, the trans-activation domain of HALF-1 did not function in yeast. CONCLUSIONS: We identified a novel trans-activation domain which contains the GCB motif conserved among plant bZIP-type factors. The trans-activation appears to be mediated by interaction between the GCB motif and a factor conserved in plants.

Amino Acid Sequence↗
PubMed2001

The 5' untranslated region of the At-P5R gene is involved in both transcriptional and post-transcriptional regulation.

The steady-state level of transcripts coding for the pyrroline-5-carboxylate reductase of Arabidopsis (At-P5R) increased under salt and heat stress, mainly because of an enhanced mRNA stability. However, the At-P5R protein level was not induced, and its translation was inhibited at initiation stage and probably also at later stages. Replacement of the 5' untranslated region (5'UTR) and beta-glucuronidase (gus) fusion analysis revealed that the first 92 bp region of the At-P5R 5'UTR was sufficient to mediate transcript stabilization and translation inhibition during salt and heat stresses. Furthermore, the first 92 bp region of the At-P5R 5'UTR was also involved in transcription efficiency in a promoter-dependent manner. The results demonstrated that the stress regulation of At-P5R is complex and involves the 5'UTR which acts at three levels, partly in opposing directions.

5' Untranslated Regions↗
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↗
PubMed1992

Simple derivation of TFIID-dependent RNA polymerase II transcription systems from Schizosaccharomyces pombe and other organisms, and factors required for transcriptional activation.

Resolution of whole cell extract through two chromatographic steps yields a single protein fraction requiring only the addition of TFIID for the initiation of transcription at RNA polymerase II promoters. This approach allows the convenient generation of RNA polymerase II transcription systems from Saccharomyces cerevisiae, human lymphocytes, and Schizosaccharomyces pombe. TFIIDs from all three organisms are interchangeable among all three systems. The S. cerevisiae and Sch. pombe systems support effects of acidic activator proteins, provided a further protein fraction from S. cerevisiae is supplied. This further fraction is distinct from the mediator of transcriptional activation described previously and represents a second component in addition to general initiation factors that may facilitate a response to acidic activators.

Cell Line↗
PubMed1992

A role for the TATA-box-binding protein component of the transcription factor IID complex as a general RNA polymerase III transcription factor.

The major class of vertebrate genes transcribed by RNA polymerase (EC 2.7.7.6) III, which includes 5S rRNA genes, tRNA genes, and the adenovirus VA genes, is characterized by split internal promoters and no absolute dependence upon specific upstream sequences. Fractionation experiments have shown that transcription of such genes requires two general RNA polymerase III-specific factors, TFIIIB and TFIIIC. We now demonstrate that a third general factor is also employed by these genes. This is the TATA-box-binding protein originally identified as being a component of the general RNA polymerase II transcription factor TFIID. This protein is involved in the transcription by RNA polymerase III of every template tested, even though the promoters of VA and most vertebrate tRNA and 5S rRNA genes do not contain recognizable TATA elements.

Base Sequence↗
PubMed1993

The activation domain of transcription factor PU.1 binds the retinoblastoma (RB) protein and the transcription factor TFIID in vitro: RB shows sequence similarity to TFIID and TFIIB.

The retinoblastoma (RB) tumor suppressor protein and the TATA-box-binding protein TFIID form contacts with a number of viral transactivator proteins. One of these, the adenovirus E1A protein, can bind to both proteins. Here we present evidence that the cellular transcription factor PU.1 can bind to both RB and TFIID. Like E1A, PU.1 binds to the conserved C-terminal domain of TFIID and to the RB "pocket" domain. The PU.1 sequences required to bind either protein lie within a 75-amino acid region which functions as an independent activation domain in vivo. The ability of PU.1 to contact directly both RB and TFIID through the same 75-residue domain prompted us to look for sequence similarity between these two proteins. We find that the previously defined domain A of the RB pocket shows sequence similarity to the conserved C terminus of TFIID, whereas domain B shows sequence similarity to a second general transcription factor, TFIIB. The potential for RB to influence transcription by using TFIID- and TFIIB-related functions is discussed.

Amino Acid Sequence↗
PubMed1996

Constitutively active human Notch1 binds to the transcription factor CBF1 and stimulates transcription through a promoter containing a CBF1-responsive element.

Notch is a transmembrane receptor that plays a critical role in cell fate determination. In Drosophila, Notch binds to and signals through Suppressor of Hairless. A mammalian homologue of Suppressor of Hairless, named CBF1 (or RBPJk), is a ubiquitous transcription factor whose function in mammalian Notch signaling is unknown. To determine whether mammalian Notch can stimulate transcription through a CBF1-responsive element (RE), we cotransfected a CBF1-RE-containing chloramphenicol acetyltransferase reporter and N1(deltaEC), a constitutively active form of human Notch1 lacking the extracellular domain, into DG75, COS-1, HeLa, and 293T cells, which all contain endogenous CBF1. N1(deltaEC) dramatically increased chloramphenicol acetyltransferase activity in these cells, indicating functional coupling of Notch1 and CBF1. The activity was comparable to that produced by the Epstein-Barr virus protein EBNA2, a well-characterized, potent transactivator of CBF1. To test whether CBF1 and Notch1 interact physically, we tagged CBF1 with an epitope from the influenza virus hemagglutinin or with the N-terminal domain of gal4, and transfected the tagged CBF1 plus N1(deltaEC) into COS-1 cells. Cell lysates were immunoprecipitated and immunoblotted with several anti-Notch1 antibodies [to detect N1(deltaEC)] or with antibodies to hemagglutinin or gal4 (to detect CBF1). Each immunoprecipitate contained a complex of N1(deltaEC) and CBF1. In summary, we find that the truncated, active form of human Notch1, N1(deltaEC), binds CBF1 and activates transcription through a CBF1-RE-containing promoter. We conclude that CBF1 is a critical downstream protein in the human Notch1 signaling pathway.

Animals↗
PubMed1997

Cloning of the cDNA for the TATA-binding protein-associated factorII170 subunit of transcription factor B-TFIID reveals homology to global transcription regulators in yeast and Drosophila.

The human transcription factor B-TFIID is comprised of TATA-binding protein (TBP) in complex with one TBP-associated factor (TAF) of 170 kDa. We report the isolation of the cDNA for TAFII170. By cofractionation and coprecipitation experiments, we show that the protein encoded by the cDNA encodes the TAF subunit of B-TFIID. Recombinant TAFII170 has (d)ATPase activity. Inspection of its primary structure reveals a striking homology with genes of other organisms, yeast MOT1, and Drosophila moira, which belongs to the Trithorax group. Both homologs were isolated in genetic screens as global regulators of pol II transcription. This supports our classification of B-TFIID as a pol II transcription factor and suggests that specific TBP-TAF complexes perform distinct functions during development.

Adenosine Triphosphatases↗
PubMed1996

Localization of subunits of transcription factors IIE and IIF immediately upstream of the transcriptional initiation site of the adenovirus major late promoter.

The assembly of a preinitiation complex containing RNA polymerase II on promoter DNA is a complex process that involves several general transcription factors. Using 5-[N-(p-azidobenzoyl)-3-aminoallyl] photocross-linking, we previously determined the locations of the two large subunits of transcription factor (TF) IIA (A35 and A21), TATA box-binding protein (TBP), RNA polymerase II-associated protein (RAP) 30, and TFIIB along the Ad2 ML promoter. We have now localized TFIIE34 and RAP74 just upstream of the transcription start site. The two subunits of TFIIF, RAP74 and RAP30, cross-linked to nucleotides that probed adjacent spaces on the same face of the DNA helix beginning just downstream of TBP at -19 and extending to -5. Specific photocross-linking of TFIIE34 required the presence TFIIE56. In addition, TFIIE and RAP74 strongly stimulated cross-linking of RAP30 and the large subunits of RNA polymerase II to position -19. Our topological data support the idea that RAP74 and TFIIE34 may be involved in melting of the promoter DNA upstream of the initiation site.

Adenoviruses, Human↗
PubMed1996

Insulin-like growth factor-I regulates transcription of the elastin gene through a putative retinoblastoma control element. A role for Sp3 acting as a repressor of elastin gene transcription.

Previous studies have demonstrated that insulin-like growth factor-I (IGF-I) increases elastin gene transcription in aortic smooth muscle cells and that this up-regulation is accompanied by a loss of protein binding to the proximal promoter. Sp1 has been identified as one of the factors whose binding is lost, and in the present study we show that Sp3 binding is also abrogated by IGF-I, but in a selected manner. In functional analyses using Drosophila SL-2 cells, Sp1 expression can drive transcription from the elastin proximal promoter, while co-expression of Sp3 results in a repression of Sp1 activity. Footprint and gel shift analyses position the IGF-I responsive sequences to a putative retinoblastoma control element (RCE). Mutation of the putative RCE sequence as assessed by transient transfection of smooth muscle cells results in an increase in reporter activity equal in magnitude to that conferred by IGF-I on the wild type promoter. Together these results support the hypothesis that IGF-I-mediated increase in elastin transcription occurs via a mechanism of derepression involving the abrogation of a repressor that appears to be Sp3 binding to the RCE.

Animals↗
PubMed1997

TGT3, thyroid transcription factor I, and Sp1 elements regulate transcriptional activity of the 1.3-kilobase pair promoter of T1alpha, a lung alveolar type I cell gene.

Alveolar type I epithelial cells form the major surface for gas exchange in the lung. To explore how type I cells differ in gene expression from their progenitor alveolar type II cells, we analyzed transcriptional regulation of T1alpha, a gene expressed by adult type I but not type II cells. In vivo developmental patterns of T1alpha expression in lung and brain suggest active gene regulation. We cloned and sequenced 1.25 kilobase pairs of the T1alpha promoter that can drive reporter expression in lung epithelial cell lines. Deletion analyses identified regions important for lung cell expression. The base pair (bp) -100 to -170 fragment conferred differential regulation in lung epithelial cells compared with fibroblasts. Sequence alignment of this fragment with type II-specific surfactant protein B and C promoters shows similar consensus elements arranged in a different order. Gel retardation studies with alveolar epithelial cell line nuclear extracts, thyroid transcription factor I (TTF-1) homeodomain, hepatic nuclear factor (HNF)-3beta, or Sp1 proteins, and supershift assays were used to characterize TTF-1, HNF-3 (TGT3), and Sp1/Sp3 binding sites. The TGT3 site binds factors with binding properties similar to HNF-3/Fkh (hepatic nuclear factor-3/forkhead) proteins but different from HNF-3alpha or HNF-3beta. Co-transfection with a TTF-1 expression vector moderately transactivated the -170 bp-reporter construct. Mutational analysis of these three binding sites showed reduced transcriptional activity of the -170 bp promoter. Therefore, several regulatory sequences involved in type II cell gene regulation are also present in the T1alpha promoter, suggesting that genes of the peripheral lung epithelium may be regulated by similar factors.

Adult↗
PubMed1997

Yeast Gal11 and transcription factor IIE function through a common pathway in transcriptional regulation.

The global transcription regulator Gal11, a component of RNA polymerase II holoenzyme, is required for full expression of many genes in yeast. We previously reported that Gal11 binds the small (Tfa2) and large (Tfa1) subunits of the general transcription factor (TF) IIE through Gal11 functional domains A and B, respectively. Here we demonstrate that the C-terminal basic region in Tfa2 is responsible for binding to domain A, whereas both the N-terminal hydrophobic and internal glutamic acid-rich regions in Tfa1 are responsible for binding to domain B. Yeast cells bearing a C-terminal deletion encompassing the Gal11-interacting region in each of the two TFIIE subunits, being viable, exhibited no obvious phenotype. In contrast, combination of the two deletions (TFIIE-DeltaC) showed phenotypes similar to those of gal11 null mutations. The levels of mRNA from TATA-containing genes, but not from TATA-less genes, decreased in TFIIE-DeltaC to an extent comparable to that in the gal11 null mutant. Combination of TFIIE-DeltaC with a gal11 null mutation did not result in an enhanced effect, suggesting that both TFIIE and Gal11 act in a common regulatory pathway. In a reconstituted cell-free system, Gal11 protein stimulated basal transcription in the presence of wild-type TFIIE. Such a stimulation was not seen in the presence of TFIIE-DeltaC.

Cell-Free System↗