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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Escherichia coli cytR-encoded repressor protein (CytR) controls the expression of several genes involved in nucleoside and deoxynucleoside uptake and metabolism. The cytR promoter was identified by determining the transcriptional initiation site of the cytR gene. A chromosomal cytR-lacZ+ operon fusion was isolated and used to study the regulation of cytR. We show that cytR expression is negatively controlled by the CytR protein and positively affected by the cAMP/CAP complex. Footprinting studies with purified CAP protein revealed two CAP binding sites upstream of the cytR promoter. A previously described mutation (cytR*) in the cloned cytR gene, which results in the phenotypic suppression of a CytR operator mutation in the tsx P2 promoter, was analysed. DNA sequence analysis of the cytR* mutation revealed a G-C to an A-T base pair transition at position -34 bp relative to the translational initiation site of cytR. This point mutation activates a cryptic promoter that is stronger than the wild-type cytR promoter and leads to overproduction of the CytR repressor.
Mol Microbiol 1990 Mar
PMID:Transcriptional regulation of the cytR repressor gene of Escherichia coli: autoregulation and positive control by the cAMP/CAP complex. 216 67

The hormonal regulation of the human urokinase type plasminogen activator (uPA) gene has been studied by introducing into mouse and rat Sertoli cell primary cultures a recombinant plasmid, in which the transcription regulatory elements of the cloned human uPA gene drive the expression of the bacterial chloramphenicol-acetyl-transferase gene. It was found to be expressed and regulated by FSH and (Bu)2cAMP in the mouse cells only, in agreement with data on the expression of the endogenous gene in rat and mouse gonads. The stimulation of transcription by FSH was evident in cultures from 13-day-old but not from 18-day-old mice, even though (Bu)2cAMP induction could be observed at both ages. Phorbol-myristate acetate was found to activate the human uPA promoter in Sertoli cell cultures from mice of both ages, even though the effect was less evident in cultures of 18-day-old animals. Deletion analysis of the human uPA 5'-flanking region showed that the distal enhancer element is not needed for (Bu)2cAMP induction, and that at least two promoter regions are involved in (Bu)2cAMP induced transcription. One of these cAMP responsive regions lies between nucleotides -72 and -29 from the CAP site. The sequence of this region would suggest the binding of transcription factor AP-2, a cell-specific mediator of both cAMP and phorbol esters action on gene expression. However, these sequences do not mediate phorbol ester activation of human uPA promoter in mouse Sertoli cells.
Mol Endocrinol 1990 Jun
PMID:Follicle-stimulating hormone and cyclic AMP induce transcription from the human urokinase promoter in primary cultures of mouse Sertoli cells. 217 96

The role of solvation on the sequence dependent conformational variabilities in DNA has been studied by calculating hydration free energies from solvent accessible surface areas for several base steps, as a function of various helical parameters, roll, twist and propeller twist. The results of roll calculations suggest opposite trends for AA and GG steps, with the former tending to have a compressed minor groove and the latter a compressed major groove. These trends are consistent with the experimental findings on sequence preferences and the nature of anisotropic bending of DNA observed in nucleosomes (Drew, H.R. and Travers, A.A., J. Mol. Biol. 186, 773-790 (1985); Satchwell, S.C., Drew, H.R. and Travers, A.A., J. Mol. Biol. 191, 659-675 (1986)) and CAP-DNA interactions (Gartenberg, M.R. and Crothers, D.M., Nature 333, 824-829, (1988)). Solvation energy profiles also indicate preferences for the base pairs in GG and AA steps to adopt low and high propeller twists, respectively. Such agreements may either reflect a coincidence of solvation effects with other energy terms or a dominance of solvent effects. The results are discussed in the context of the crystallographic observations of structural tendencies.
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PMID:Solvation effects on the sequence variability of DNA double helical conformations. 227 93

DNase I footprint analysis of the core adenovirus 2 (Ad2) major late promoter (MLP) has revealed distinct patterns of protection corresponding to the assembly of transcription components during transcriptional initiation (VanDyke, M. W., Sawadogo, M., and Roeder, R. G. (1989) Mol. Cell Biol. 7, 3371-3379). By using partially purified transcription factors, DNase I protection over the TATA box element and the CAP sequence was attributed to the binding of a single factor, TFIID. We have determined, however, that protection of the CAP region results from the binding of a novel factor, designated CAP-site binding factor (CBF), which is chromatographically and functionally distinct from TFIID. DNase I footprint analysis and gel electrophoresis mobility shift competition assays confirm that distinct polypeptides bind to the Ad2 MLP upstream promoter sequence, TATA box, and CAP sequences. When the CAP sequence is mutated, transcriptional activity of the Ad2 MLP is reduced both in vitro and in vivo. The decrease in transcriptional activity correlates with decreased CBF binding activity. Nuclear extracts depleted of CBF also exhibit reduced Ad2 MLP transcriptional activity. The addition of DNA affinity purified CBF, free of TFIID or major late transcription factor, restores the activity to control levels.
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PMID:Identification and characterization of an adenovirus 2 major late promoter CAP sequence DNA-binding protein. 235 2

The Escherichia coli lac promoter mutation Pr115, an A X T to T X A transversion at +1 (the transcription initiation site of the lac wild-type and lac UV5 promoters), creates a new "-10 region"-like sequence starting at +1. We show that this mutation activates a new RNA polymerase binding site (P115) that overlaps with, and is shifted 12 base-pairs downstream from, the wild-type RNA polymerase binding site (P1). Nuclease S1 mapping studies and RNA polymerase protection experiments in vitro indicate that, in the absence of CAP-cAMP, this new site is used preferentially over the P1 site. In vivo, beta-galactosidase assays of the Pr115 mutation in combination with mutations of the P1 "-35 region" demonstrate that the P1 -35 region sequences are not involved in the interaction between RNA polymerase and P115 in the absence of CAP-cAMP; therefore P115 is an independent binding site. The presence of CAP-cAMP in vivo stimulates polymerase binding and initiation at P1, which serves to block polymerase from binding at P115.
J Mol Biol 1985 Oct 05
PMID:Lactose promoter mutation Pr115 activates an overlapping promoter within the lactose control region. 241 53

An electrophoretic procedure for the measurement of the helix unwinding induced by a sequence-specific protein is described. The method, which was applied here to EcoR I, CAP and lac repressor, involved the migration of the complexes with positively and negatively supercoiled DNA minicircles carrying a single protein binding site. Mobility shifts of complexes relative to naked DNAs appeared to be a result of i) the unwinding; of ii) an increase in the molecular frictional coefficient, which led to a retardation; of iii) bending, in the particular case of CAP, which induced an acceleration; and of iv) looping, in the case of lac repressor, which also resulted in an acceleration. Under conditions where the migration of the naked topoisomers was V-like (topoisomer mobility showed the same linear increase with both negative and positive supercoilings; Zivanovic et al. (1986) J. Mol. Biol., 192, 645-660), the protein unwinding contribution to mobility was assumed to be identical to that experimentally observed in the case of a thermal unwinding: all negatively supercoiled topoisomers were retarded and all positively supercoiled topoisomers were accelerated to the same extent. In contrast, the mobility contribution of the frictional term, as well as those of bending and looping, appeared to vary strongly with the magnitude of the supercoiling, but only weakly with its polarity. As a consequence, these latter contributions may approximately cancel when one is measuring the difference between the shifts observed for two comigrating, negatively and positively supercoiled, topoisomers, allowing the unwinding to be calculated. While estimates obtained for EcoR I, 23 +/- 3 degrees, and CAP, about 29 degrees, were in good agreement with previous measurements using topoisomerase I, the value found for lac repressor, 13 to 16 degrees, was significantly smaller.
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PMID:Protein-induced unwinding of DNA: measurement by gel electrophoresis of complexes with DNA minicircles. Application to restriction endonuclease EcoRI, catabolite gene activator protein and lac repressor. 254 54

The DNA-binding C-terminal domains of the regulatory proteins Fnr from Escherichia coli and FixK from Rhizobium meliloti have been modelled on the basis of their homologies to the CAP protein from E. coli. Residues Glu181, Thr182 and Arg185 of CAP, which are exposed residues of the DNA-recognition helix alpha F, are conserved in Fnr and FixK. However, Arg180 and Gly184 are substituted by Val and Ser respectively in Fnr. We propose that this valine makes a Van der Waals' contact with the first thymine in the Fnr consensus TTGA-N6-TCAA, and that the serine contributes to the binding by displacing a thymine-bound water molecule. The corresponding residues in FixK, Ile and Ser allow the same interactions with a thymine. Therefore we predict that FixK may recognize the same sites as Fnr. This is supported experimentally by showing that Fnr can substitute for FixK in activating the fixN gene in E. coli.
J Mol Recognit 1989 Nov
PMID:Model-building of Fnr and FixK DNA-binding domains suggests a basis for specific DNA recognition. 256 29

In this paper, we have used filter hybridization and nucleotide sequencing to analyse the relationship between the three genes of the pelADE cluster in the Erwinia chrysanthemi (Ech) strain B374. This cluster encodes for three of the five pectate lyase proteins that are involved in the maceration and soft-rotting of plant tissue, an important trait in Ech pathogenicity. Southern hybridization revealed homology between each of the three pel genes. A 3560 bp DNA fragment containing the pelE and pelD genes was sequenced. These two genes show extensive homology in the coding regions but only low homology in the 5' and 3' non-coding regions. However both genes exhibit sequences homologous to the Escherichia coli CAP-binding site consensus sequence upstream of the start codon and an inverted repeat sequence which may act as a rho-independent transcriptional terminator after the translational stop. The pel genes of Ech B374 were also compared with the already sequenced pel genes of EC16, another Ech strain.
Mol Microbiol 1989 Oct
PMID:Relationship between the pel genes of the pelADE cluster in Erwinia chrysanthemi strain B374. 261 52

Characterization of ternary complexes containing an Escherichia coli lac promoter DNA fragment, CAP protein and RNA polymerase, separated on non-denaturing polyacrylamide gels and footprinted in the gel slice, reveals a striking stabilization of CAP against dissociation in the open complex, compared to the CAP-DNA complex lacking RNA polymerase. The stabilization is lost when half a helical turn of DNA is inserted between CAP and polymerase sites, but is partially restored with an 11 base-pair insert; stimulation of transcription parallels the stabilization effect. This behavior suggests a direct protein-protein interaction. Comparison of initiation kinetics for wild-type and a mutant in which the P2 promoter has been inactivated shows that CAP both strengthens binding in the closed complex and accelerates isomerization to the open complex; the latter effect accounts for the bulk of the observed transcriptional activation.
J Mol Biol 1989 Mar 05
PMID:Synergy between Escherichia coli CAP protein and RNA polymerase in the lac promoter open complex. 264 87

In enteric bacteria, the expression of many genes encoding various anaerobic electron transfer functions is controlled by FNR, the product of the autoregulated fnr gene. FNR is structurally and functionally homologous to CAP, the catabolite gene activator protein, and increased FNR production strongly stimulates transcription of its target genes. By analysis of RNA produced in vivo the promoters of four FNR-dependent genes were localized and shown to display a common arrangement. A 22bp dyad symmetry was found about 30 nucleotides upstream of the transcriptional startpoints and a similar sequence was shown to overlap the site of transcription initiation in the negatively controlled fnr gene. The consensus sequence for the half site recognized by FNR (AAA-TTGAT) is only slightly different from that of CAP (AA-TGTGA). Studies with two mutant frd promoters from Escherichia coli, displaying altered regulation and FNR response, provided additional evidence for recognition of this sequence by FNR.
Mol Microbiol 1989 Jul
PMID:Molecular genetic analysis of FNR-dependent promoters. 267 2


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