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: UNIPROT:P51532 (transcriptional activator)
6,546 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pterin 4a-carbinolamine dehydratase is bifunctional in mammals. In addition to playing a catalytic role in pterin recycling in the cytoplasm, it plays a regulatory role in the nucleus, where it acts as a dimerization-cofactor component (called DCoH) for the transcriptional activator HNF-1alpha. A thus far unique operon in Pseudomonas aeruginosa contains a gene encoding a homolog (PhhB) of the regulatory dehydratase, together with genes encoding phenylalanine hydroxylase (PhhA) and aromatic aminotransferase (PhhC). Using complementation of tyrosine auxotrophy in Escherichia coli as a functional test, we have found that the in vivo function of PhhA requires PhhB. Strikingly, mammalian DCoH was an effective substitute for PhhB, and either one was effective in trans. Surprisingly, the required presence of PhhB for complementation did not reflect a critical positive regulatory effect of phhB on phhA expression. Rather, in the absence of PhhB, PhhA was found to be extremely toxic in E. coli, probably due to the nonenzymatic formation of 7-biopterin or a similar derivative. However, bacterial PhhB does appear to exert modest regulatory effects in addition to having a catalytic function. PhhB enhances the level of PhhA two- to threefold, as was demonstrated by gene inactivation of phhB in P. aeruginosa and by comparison of the levels of expression of PhhA in the presence and absence of PhhB in Escherichia coli. Experiments using constructs having transcriptional and translational fusions with a lacZ reporter indicated that PhhB activates PhhA at the posttranscriptional level. Regulation of PhhA and PhhB is semicoordinate; both PhhA and PhhB are induced coordinately in the presence of either L-tyrosine or L-phenylalanine, but PhhB exhibits a significant basal level of activity that is lacking for PhhA. Immunoprecipitation and affinity chromatography showed that PhhA and PhhB form a protein-protein complex.
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PMID:PhhB, a Pseudomonas aeruginosa homolog of mammalian pterin 4a-carbinolamine dehydratase/DCoH, does not regulate expression of phenylalanine hydroxylase at the transcriptional level. 1021 69

Recent studies have established an essential role for p38 MAP kinase in UV activation of human immunodeficiency virus (HIV) gene expression. However, p38 MAP kinase is not involved in activation of NF-kappa B, a key transcriptional activator of HIV gene expression, in response to UV, suggesting that NF-kappa B acts independently of p38 MAP kinase. In this study, we have investigated whether activation of HIV gene expression occurs when p38 MAP kinase and NF-kappa B are activated by separate stress-causing treatments, each relatively specific for activating only one of the factors. Treatment of cells with sorbitol (hyperosmotic shock) strongly activates p38 MAP kinase, whereas the cytokine TNF-alpha is a poor activator of p38 MAP kinase. On the other hand, TNF-alpha is a strong activator of NF-kappa B whereas sorbitol is not. Sorbitol, however, activates AP-1 DNA binding activity in a manner similar to that of UV. Most importantly, both sorbitol and TNF-alpha are poor activators of HIV gene expression in HeLa cells stably transfected with an HIVcat reporter gene, whereas UV elicits a strong response. The combined treatment with UV and hyperosmotic shock produces an additive effect on HIV gene expression, suggesting that these agents activate at least in part by different mechanisms. The combined treatment with sorbitol and TNF-alpha activates p38 and NF-kappa B to levels similar to those with UV, yet only results in 25-30% of the CAT levels elicited by UV. Inhibition of NF-kappa B activation by the protease inhibitor N-alpha-tosyl-L-phenylalanine chloromethyl ketone (TPCK) prevents UV activation of HIV gene expression, but does not inhibit p38 MAP kinase activation. We conclude that whereas both p38 MAP kinase and NF-kappa B are important for UV activation of HIV gene expression they act independently from each other and activation of both factors is not sufficient for triggering a full HIV gene expression response. Activation of HIV gene expression by UV must therefore involve additional cellular processes, such as those triggered by DNA damage, for generation of a full gene expression response.
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PMID:Activation of NF-kappa B and p38 MAP kinase is not sufficient for triggering efficient HIV gene expression in response to stress. 1067 19

The contributions of defective mismatch repair (MMR) and the p53-response to cell killing by N-(2-chloroethyl)-N'-cyclohexyl-N-nitrosourea (CCNU) were evaluated. MMR defects were previously shown to be associated with CCNU sensitivity (G. Aquilina et al., Cancer Res., 58: 135-141, 1998). Unexpectedly, eight MMR-deficient variants of the A2780 human ovarian carcinoma cell line were 3-fold more resistant to CCNU than the MMR-proficient parental cells. The variants were members of a preexisting subpopulation of drug-resistant A2780 cells. In addition to deficient expression of the MMR protein hMLH1, an essential component of the hMutL alpha repair complex, the variants exhibited alterations in the expression of other genes that influence drug sensitivity. Although A2780 cells possess a wild-type p53 gene, all of the clones contained a heterozygous G to T tranversion at codon 172. This change resulted in a Val to Phe substitution and was associated with a constitutive production of high levels of p53, which was inactive as a transcriptional activator of bax and p21. The hMLH1/p53 defective variants displayed a less prominent cell cycle arrest and reduced apoptosis after CCNU treatment. In contrast, MMR-defective A2780 variants, which had a similar hMutL alpha defect but retained a wild-type p53, did exhibit the expected CCNU sensitivity. Expression of a dominant-negative p53val135 increased CCNU resistance of both MMR-proficient and MMR-deficient A2780 cells. Thus, defective MMR and p53 influence CCNU sensitivity in opposite directions. Their effects are independent, and sensitization by defective MMR does not require a functional p53 response.
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PMID:Mismatch repair and p53 independently affect sensitivity to N-(2-chloroethyl)-N'-cyclohexyl-N-nitrosourea. 1069 May 53

Using a modified TAIL-PCR technique, the 5'-flanking regions of the phenylalanine ammonia lyase (Pal) genes of a yam species, Dioscorea bulbifera, and the phosphoglucose isomerase (Pgi) gene of D. tokoro were successfully isolated. Two novel modifications of the TAIL-PCR procedure introduced here, namely (1) the use of a battery of random 10-mers (RAPD primers) as short arbitrary primers, and (2) the use of a total of five nested, gene-specific primers, allow the rapid isolation of the 5'-flanking region of any gene from organisms with large genomes. Isolated 5'-flanking regions were fused to the gus gene, and tested for transient expression in tobacco BY2 cells. All the isolated 5'-flanking regions were shown to drive reporter gene expression. Three Pal promoters responded to salicylic acid, presumably as a result of the binding of a MYB transcriptional activator to the multiple MREs (Myb Recognition Elements) present in these regions.
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PMID:Rapid isolation of promoter sequences by TAIL-PCR: the 5'-flanking regions of Pal and Pgi genes from yams (Dioscorea). 1082 Nov 91

We have identified four mutations in Xenopus TFIIIA that increase the stability of TFIIIA-5 S rRNA gene complexes. In each case, the mutation has a relatively modest effect on equilibrium binding affinity. In three cases, these equilibrium binding effects can be ascribed primarily to decreases in the rate constant for protein-DNA complex dissociation. In the fourth case, however, a substitution of phenylalanine for the wild-type leucine at position 148 in TFIIIA results in much larger compensating changes in the kinetics of complex assembly and dissociation. The data support a model in which a relatively unstable population of complexes with multi-component dissociation kinetics forms rapidly; complexes then undergo a slow conformational change that results in very stable, kinetically homogeneous TFIIIA-DNA complexes. The L148F mutant protein acts as a particularly potent transcriptional activator when it is fused to the VP16 activation domain and expressed in yeast cells. Substitution of L148 to tyrosine or tryptophan produces an equally strong transcriptional activator. Substitution to histidine results in genetic and biochemical effects that are more modest than, but similar to, those observed with the L148F mutation. We propose that an amino acid with a planar side chain at position 148 can intercalate between adjacent base pairs in the intermediate element of the 5 S rRNA gene. Intercalation occurs slowly but results in a very stable DNA-protein complex. These results suggest that transcriptional activation by a cis-acting sequence element is largely dependent on the kinetic, rather than the thermodynamic, stability of the complex formed with an activator protein. Thus, transcriptional activation is dependent in large part on the lifetime of the activator-DNA complex rather than on binding site occupancy at steady state. Introduction of intercalating amino acids into zinc finger proteins may be a useful tool for producing artificial transcription factors with particularly high in vivo activity.
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PMID:Mutations in TFIIIA that increase stability of the TFIIIA-5 S rRNA gene complex: unusual effects on the kinetics of complex assembly and dissociation. 1588 46

Aft1p is an iron-responsive transcriptional activator that plays a central role in maintaining iron homeostasis in Saccharomyces cerevisiae. Aft1p is regulated primarily by iron-induced shuttling of the protein between the nucleus and cytoplasm, but its nuclear import is not regulated by iron. Here, we have shown that the nuclear export of Aft1p is promoted in the presence of iron and that Msn5p is the nuclear export receptor (exportin) for Aft1p. Msn5p recognizes Aft1p in the iron-replete condition. Phosphorylation of S210 and S224 in Aft1p, which is not iron dependent, and the iron-induced intermolecular interaction of Aft1p are both essential for its recognition by Msn5p. Mutation of Cys291 of Aft1p to Phe, which causes Aft1p to be retained in the nucleus and results in constitutive activation of Aft1-target genes, disrupts the intermolecular interaction of Aft1p. Collectively, these results suggest that iron induces a conformational change in Aft1p, in which Aft1p Cys291 plays a critical role, and that, in turn, Aft1p is recognized by Msn5p and exported into the cytoplasm in an iron-dependent manner.
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PMID:Mechanism underlying the iron-dependent nuclear export of the iron-responsive transcription factor Aft1p in Saccharomyces cerevisiae. 1753 22

In an effort to identify novel components of the PHO regulon in Saccharomyces cerevisiae, we have isolated and characterized suppressors of the Pho(-) phenotype associated with deletion of the Pho4 transcriptional activator. Here we report that either a defective form of the Rsp5 E3 ubiquitin ligase or deletion of the End3 component of the endocytic pathway restores growth of the pho4 Delta mutant in the presence of limiting inorganic phosphate (P i). The spa1-1 suppressor allele of RSP5 encodes a phenylalanine-to-valine replacement at position 748 (F748V) within the catalytic HECT domain of Rsp5. Consistent with suppression due to impaired ubiquitin ligase activity, the heat-sensitive growth defect of the spa1-1 mutant is suppressed either by overexpression of ubiquitin or by osmotic stabilization. Western blot analyses revealed that the cellular levels of the Pho87 and Pho91 low affinity P i are markedly increased in the spa1-1 mutant, yet Pho84 high affinity P i transporter levels are unaffected. Furthermore, Pho87 and Pho91 are ubiquitinated in vivo in an Rsp5-dependent manner, and the Pho+ phenotype of the spa1-1 suppressor is dependent upon Pho87 and Pho91. We conclude that turnover of the low affinity P i transporters is initiated by Rsp5-mediated ubiquitination followed by internalization and degradation by the endocytic pathway.
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PMID:The Rsp5 E3 ligase mediates turnover of low affinity phosphate transporters in Saccharomyces cerevisiae. 1816 38

A series of transcriptional activator (TAT)-protein transduction domains (PTDs) modified with hydrophobic amino acids were used as model cationic amphiphilic peptides to study the effect of hydrophobicity on interaction of such peptides with plasmid DNA. The peptide-DNA complexes were analyzed by dynamic light scattering and gel electrophoresis to determine their size and electrokinetic properties at various +/- charge ratios. Peptides in solution were found to have a tendency to aggregate and the hydrodynamic size of the aggregates depends on the structure of peptide. Peptides with smaller hydrophobic residues at the N-terminal formed smaller complexes with DNA compared to the ones with larger hydrophobic tails. DNA complexes having peptides with more than one hydrophobic moiety at the N-terminal had a tendency to aggregate. Among the peptides having single hydrophobic amino acid at the N-terminal, DNA complexes of Tyr-TAT and Phe-TAT were found to be stable in solution. The size of the hydrophobic domain and the type of hydrophobic amino acid at the N-terminal of cationic amphiphilic peptides play an important role not only in the complex formation but also in stabilizing the system. The studies presented here indicate that there is a potential for strategic development of these peptides into potential non-viral gene delivery vectors.
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PMID:The effect of the structure of small cationic peptides on the characteristics of peptide-DNA complexes. 1903 20

Protein-protein interactions play an essential role in cellular function, and methods to discover and characterize them in their native context are of paramount importance for gaining a deeper understanding of biological networks. In this study, an enhanced nonsense suppression system was utilized to incorporate the nonnatural amino acid p-benzoyl-L-phenylalanine (pBpa) throughout the transcriptional activation domain of the prototypical eukaryotic transcriptional activator Gal4 in vivo (S. cerevisiae). Functional studies of the pBpa-containing Gal4 mutants suggest that this essential binding interface of Gal4 is minimally impacted by these substitutions, with both transcriptional activity and sensitivity to growth conditions maintained. Further supporting this are in vivo cross-linking studies, including the detection of a key binding partner of Gal4, the inhibitor protein Gal80. Cross-linking with a range of pBpa-containing mutants revealed a Gal4 x Gal80 binding interface that extends beyond that previously predicted by conventional strategies. Thus, this approach can be broadened to the discovery of novel binding partners of transcription factors, information that will be critical for the development of therapeutically useful small molecule modulators of these protein-protein interactions.
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PMID:Impact of nonnatural amino acid mutagenesis on the in vivo function and binding modes of a transcriptional activator. 1976 47

Expression of the sulfur assimilation pathway in Aspergillus nidulans is under control of sulfur metabolite repression, which is composed of scon genes encoding subunits of ubiquitin ligase and the metR gene coding for a transcriptional activator. In this paper we report three dominant suppressors of methionine requirement isolated from a metB3 diploid strain. All three mutations lead to the substitution of phenylalanine 48 by serine or leucine in the conserved N-terminal region of the MetR protein. Strains carrying the dominant suppressor mutations exhibit increased activities of homocysteine synthase and sulfur assimilation enzymes as well as elevated levels of the corresponding transcripts. These changes are observed even under conditions of methionine repression, which suggests that the mutated MetR protein may be resistant to inactivation or degradation mediated by sulfur metabolite repression. We also found that a mutant impaired in sulfite reductase activity, known until now as sG8, has a frameshift which changes 41 C-terminal amino acids. Therefore, it is now designated metR18. This mutant has elevated levels of MetR-regulated transcripts and of activities of sulfur assimilation enzymes (except sulfite reductase), which can be repressed to the wild type level by exogenous methionine. Thus, metR18 and the three dominant suppressors represent new types of mutations affecting different parts of the A. nidulans MetR protein.
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PMID:Novel mutations reveal two important regions in Aspergillus nidulans transcriptional activator MetR. 2095 10


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