Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Consistent with the complexity of the temporally regulated processes that must occur for growth and development of higher eukaryotes, it is now apparent that transcription is regulated by the formation of multicomponent complexes that assemble on the promoters of genes. These complexes can include (in addition to the five or more general transcription factors and RNA polymerase II) DNA-binding proteins, transcriptional activators, coactivators, adaptors and various accessory proteins. The best studied example of a complex that includes a transcriptional adaptor, accessory proteins and a DNA-binding protein is that involving the herpes simplex virus VP16 protein. Evidence suggests that the adenovirus E1a protein and the cellular Sp1 and CTF/NF1 transcription factors also function through adaptors or coactivators. Each additional component of the transcription complex provides the cell with another point at which to exert control of gene expression.
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PMID:The interactions of transcription factors and their adaptors, coactivators and accessory proteins. 166 80

The sequences preceding the albumin mRNA start site are able to direct efficient transcription only upon introduction into cells expressing the endogenous albumin gene. In transient expression assays, the activity of a reporter gene (CAT) linked to this promoter is 100-fold higher in H4II differentiated hepatoma cells than in H5 dedifferentiated cells which no longer express their albumin gene. This tissue specificity depends on the very proximal promoter region, composed of a CCAAT box, the proximal element and a TATA box. Deletion of the CCAAT box leads to a two- to threefold decrease in activity, deletion of the proximal element (PE) results in loss of activity. The PE is a high-affinity binding site for HNF1/APF, a strictly liver specific trans-acting factor. When the affinity of this factor for PE is decreased by bacterial methylation (PE includes a dam methylase site), by mutation, or by its replacement with the homologous element from the alpha-fetoprotein gene (AFP), the activity of the short promoter (PE plus the TATA box) is abolished. This activity can be rescued in the presence of the more upstream elements: DEII, DEI and the CCAAT box (recognized, respectively, by the NF1/CTF, C/EBP and NFY/ACF factors) which are then absolutely required. Our results suggest that the upstream elements contribute to promoter activity by stabilizing the HNF1-PE complex and not by direct interaction with TFIID or the RNA polymerase. It is probable that these elements, essentially dispensable in already differentiated hepatoma cells, play a crucial role during development or differentiation to activate the promoter in cells that contain a low concentration of HNF1 and/or an HNF1 unable to open inactive chromatin alone.
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PMID:Anatomy of the rat albumin promoter. 218 62

The malate-aspartate shuttle, consisting of mitochondrial and cytosolic aspartate aminotransferase and mitochondrial and cytosolic malate dehydrogenase, is a major pathway for the transport of reducing equivalents from cytosol to mitochondria in mammals. To elucidate molecular mechanisms regulating metabolic coordination between the mitochondria and the cytosol, we analyzed the 5'-flanking regulatory regions of the complete set of mouse isoenzyme genes playing a pivotal role in the shuttle. Deletion analysis and an in vivo transfection assay, using NIH3T3 cells, revealed that all the promoter regions are located within the 300-base pair regions upstream from the initiation codon. Subsequently, DNase I footprinting analyses using NIH3T3 cell nuclear extracts led to identification of several protein binding sites within these promoter regions. A synthetic oligomer containing the consensus binding site sequence for CTF/NFI, a transcription factor for RNA polymerase II, competed for the binding of proteins to the promoter regions of cytosolic aspartate aminotransferase and mitochondrial and cytosolic malate dehydrogenase genes, but not for that of the mitochondrial aspartate amino-transferase gene. On the other hand, a synthetic oligomer containing the consensus binding site sequence for Sp1, which activates transcription from promoters containing properly positioned GC boxes, competed for protein(s) binding to the promoter region of the mitochondrial aspartate aminotransferase gene.
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PMID:Regulatory regions of the mitochondrial and cytosolic isoenzyme genes participating in the malate-aspartate shuttle. 229 30

Optimized weight matrices defining four major eukaryotic promoter elements, the TATA-box, cap signal, CCAAT-, and GC-box, are presented; they were derived by comparative sequence analysis of 502 unrelated RNA polymerase II promoter regions. The new TATA-box and cap signal descriptions differ in several respects from the only hitherto available base frequency Tables. The CCAAT-box matrix, obtained with no prior assumption but CCAAT being the core of the motif, reflects precisely the sequence specificity of the recently discovered nuclear factor NY-I/CP1 but does not include typical recognition sequences of two other purported CCAAT-binding proteins, CTF and CBP. The GC-box description is longer than the previously proposed consensus sequences but is consistent with Sp1 protein-DNA binding data. The notion of a CACCC element distinct from the GC-box seems not to be justified any longer in view of the new weight matrix. Unlike the two fixed-distance elements, neither the CCAAT- nor the GC-box occurs at significantly high frequency in the upstream regions of non-vertebrate genes. Preliminary attempts to predict promoters with the aid of the new signal descriptions were unexpectedly successful. The new TATA-box matrix locates eukaryotic transcription initiation sites as reliably as do the best currently available methods to map Escherichia coli promoters. This analysis was made possible by the recently established Eukaryotic Promoter Database (EPD) of the EMBL Nucleotide Sequence Data Library. In order to derive the weight matrices, a novel algorithm has been devised that is generally applicable to sequence motifs positionally correlated with a biologically defined position in the sequences. The signal must be sufficiently over-represented in a particular region relative to the given site, but need not be present in all members of the input sequence collection. The algorithm iteratively redefines the set of putative motif representatives from which a weight matrix is derived, so as to maximize a quantitative measure of local over-representation, an optimization criterion that naturally combines structural and positional constancy. A comprehensive description of the technique is presented in Methods and Data.
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PMID:Weight matrix descriptions of four eukaryotic RNA polymerase II promoter elements derived from 502 unrelated promoter sequences. 232 77

This paper describes the structure of a 70-kb porcine gene for nuclear factor I, including its promoter region, comprising a total of 11 exons. Different mRNAs that we have isolated as cDNAs from both porcine liver and human HeLa cells presumably are generated from this gene by differential splicing events. One cDNA species from porcine liver that lacks exon 9 carries coding information for a protein of 439 amino acids. The in vitro translated protein displays all the properties of an NFI-like protein with high affinity toward the sequence element TGG(N)6GCCAA, as shown by gel shift analysis, and no or little affinity toward CCAAT box containing sequences. Cotranslation experiments with full-length and truncated variants of the protein demonstrate that it binds as a dimer to its cognate DNA recognition sequence. Its DNA-binding domain which is retained in all cDNA clones was mapped by deletion analysis to the 250 N-terminal amino acids of the protein. No structural homologies are observed between this protein and other known DNA-binding proteins; instead, the protein contains a novel alpha-helical sequence motif consisting of several lysine residues spaced at intervals of seven amino acids which we have termed the "lysine helix". The C-terminal portion of the protein derived from full-length cDNAs encodes a short amino acid sequence which is identical with the heptapeptide repeat CT7 observed in the C-terminal domain of the largest subunits of yeast and mouse RNA polymerase II. This region is removed by differential splicing in some of the NFI/CTF cDNAs and thus may be of functional significance.
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PMID:Structural and functional organization of a porcine gene coding for nuclear factor I. 251 76

Promoter-proximal downstream regions of the human immunodeficiency viruses (HIV-1 and HIV-2) mediate the action of the viral transcription activator protein, Tat. We demonstrate here that the downstream domain of each virus interacts with two RNA polymerase II transcription factors. One of these, CTF/NF I, is a multifunctional protein associated previously with activation of transcription and DNA replication. The other cellular protein, designated LBP-1 (leader-binding protein-1), recognizes repeated elements within an extended region of DNA corresponding to part of the 5'-untranslated leader. Analysis of clustered point mutants in the HIV-1 leader for DNA-binding and transcription activity in vitro and in vivo suggests a role for LBP-1 as part of the basal promoter. A complex overlapping arrangement is observed between sequences required for the interaction of LBP-1 and CTF/NF I proteins and those defined previously for regulation by the HIV-1 Tat protein.
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PMID:Structural arrangements of transcription control domains within the 5'-untranslated leader regions of the HIV-1 and HIV-2 promoters. 284 59

Transcription factor CTF, which is responsible for selective recognition of eukaryotic promoters that contain the sequence CCAAT, was purified to apparent homogeneity by sequence-specific DNA affinity chromatography. Binding sites for CTF in the human Ha-ras and alpha-globin promoters were highly homologous to sequences recognized by nuclear factor I (NF-I), a cellular DNA-binding protein that is required for the initiation of adenovirus DNA replication in vitro. To determine the relationship between CTF and NF-I, we compared the biochemical properties of these two proteins. CTF and NF-I were found to be indistinguishable in polypeptide composition, DNA-binding properties, immunological cross-reactivity, and in vitro stimulation of DNA replication and transcription initiation. We conclude that CTF/NF-I can serve both as a transcription selectivity factor for RNA polymerase II and as an initiation factor for adenovirus DNA replication.
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PMID:A cellular DNA-binding protein that activates eukaryotic transcription and DNA replication. 302 47

The carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II consists of tandem repeats of a heptapeptide with the consensus YSPTSPS. It has been shown that the heptapeptide repeat interacts directly with the general transcription factor TFIID. We report here that the CTD activates transcription when fused to the DNA-binding domain of GAL4. More importantly, we find that the proline-rich transcriptional activation domain of the CCAAT-box-binding factor CTF/NF1 contains a sequence with striking similarity to the heptapeptide repeats of the CTD. We show that this CTD-like motif is essential for the transcriptional activator function of the proline-rich domain of CTF/NF1. Deletion of and point mutations in this CTD-like motif abolish the transcriptional activator function of the proline-rich domain, while natural CTD repeats from RNA polymerase II are fully functional in place of the CTD-like motif. We further show that the proline-rich activation domain of CTF/NF1 interacts directly with the TATA-box-binding protein (TBP), and that a mutation in the CTD-like motif that abolishes transcriptional activation reduces the affinity of the proline-rich domain for TBP. These results demonstrate that a class of proline-rich activator proteins and RNA polymerase II possess a common structural and functional component which can interact with the same target in the general transcription machinery. We discuss the implications of these results for the mechanisms of transcriptional activation in eucaryotes.
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PMID:The upstream activator CTF/NF1 and RNA polymerase II share a common element involved in transcriptional activation. 802 1

Initiation of RNA polymerase II-directed transcription is mediated by DNA sequence specific activator proteins interacting with components of the basal transcription machinery. NFI/CTF is a family of such binding proteins which have been shown to stimulate transcription via proline-rich activation domains. In order to identify residues crucial for its activator function, a pool of CTF1 mutants was cloned and fused to the bacterial repressor LexA. Transcriptional activation of these constructs was monitored in a Saccharomyces cerevisiae reporter assay. Our studies reveal the existence of a core domain in CTF1 between residues 463 and 508 essential for transcriptional activation functions. It contains the sequence motif SPTSPSYSP, which is strongly related to the heptapeptide repeat YSPTSPS present in the carboxyterminal domain (CTD) of RNA polymerase II. Removal of the entire CTD related motif, as well as substitution of key amino acids therein, abolish CTF1 mediated transcriptional activation.
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PMID:Transcriptional activation of NFI/CTF1 depends on a sequence motif strongly related to the carboxyterminal domain of RNA polymerase II. 804 23

The transcription factor CTF/NF-1 is a multifunctional cellular protein which participates in the expression of host and viral genes and in the replication of viral DNA. A procedure was developed to obtain recombinant CTF/NF-1 protein in order to help identify the binding sites of CTF/NF-1 protein in the regulatory region of viral sequences. Cloning and expression of the CTF/NF-1 gene downstream from a T7 RNA polymerase promoter resulted in an insoluble protein produced in Escherichia coli. Specific binding by the recombinant CTF/NF-1 protein to its cognate recognition site was obtained only after a denaturation and renaturation procedure. No special chromatography steps were required to obtain a rCTF/NF-1 preparation which could be used for our binding studies. Specific binding was detected with an NF-1 oligonucleotide and with viral DNA templates. Binding to the viral DNA sequences was identical to that previously published with biochemically purified cellular CTF/NF-1 protein.
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PMID:Overexpression and reactivation of binding activity of the recombinant CTF/NF-1 transcription factor. 816 74


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