UNIPROT:P51532 - FACTA Search

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

ComK, a key transcriptional regulator in the development of competence in Bacillus subtilis, is required for its own transcription as well as that of the late competence genes encoding proteins involved in DNA uptake. ComK is sequestered in a complex with ClpC and MecA until a peptide, ComS, accumulates in cells. ComS releases ComK from the inhibitory complex, thus allowing ComK to carry out its function as a transcriptional activator. Spx (formerly YjbD), a negative effector of competence, accumulates in clpP mutants. High concentrations of Spx may be responsible for the inability of clpP mutants to become competent because spx mutations are able to restore competence in the clpP mutant. In this paper, we showed, based on in vitro experiments, that Spx forms a quaternary complex with ClpC, MecA and ComK and enhances ComK binding to ClpC-MecA. Two ComS alanine substitution mutants (I13A and W43A), previously shown to be defective in vivo, were less efficient in releasing ComK from ClpC-MecA. The I13A mutant with a weaker binding affinity to MecA was inefficient in releasing ComK regardless of whether Spx was present. In contrast, the defect of the W43A mutant in dissociating ComK was more readily observed in the presence of Spx. Spx is a highly conserved protein among Gram-positive bacteria, in which it may function closely with the protease adaptor protein, MecA.
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PMID:Spx (YjbD), a negative effector of competence in Bacillus subtilis, enhances ClpC-MecA-ComK interaction. 1202 82

Protein kinase GCN2 regulates translation initiation by phosphorylating eukaryotic initiation factor 2alpha (eIF2alpha), impeding general protein synthesis but specifically inducing translation of GCN4, a transcriptional activator of amino acid biosynthetic genes in Saccharomyces cerevisiae. GCN2 activity is stimulated in amino acid-deprived cells through binding of uncharged tRNA to a domain related to histidyl tRNA synthetase. We show that GCN2 is phosphorylated by another kinase on serine 577, located N-terminal to the kinase domain. Mutation of Ser-577 to alanine produced partial activation of GCN2 in nonstarved cells, increasing the level of phosphorylated eIF2alpha, derepressing GCN4 expression, and elevating the cellular levels of tryptophan and histidine. The Ala-577 mutation also increased the tRNA binding affinity of purified GCN2, which can account for the elevated kinase activity of GCN2-S577A in nonstarved cells where uncharged tRNA levels are low. Whereas Ser-577 remains phosphorylated in amino acid-starved cells, its dephosphorylation could mediate GCN2 activation in other stress or starvation conditions by lowering the threshold of uncharged tRNA required to activate the protein.
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PMID:Serine 577 is phosphorylated and negatively affects the tRNA binding and eIF2alpha kinase activities of GCN2. 1207 Jan 58

Activation of class II major histocompatibility complex (MHC) gene expression is regulated by a master regulator, class II transcriptional activator (CIITA). Transactivation by CIITA requires its nuclear import. This study will address a mechanistic role for the leucine-rich repeats (LRR) of CIITA in regulating nuclear translocation by mutating 12 individual consensus-motif "leucine" residues in both its alpha-motifs and beta-motifs. While some leucine mutations in the LRR motif of CIITA cause congruent loss of transactivation function and nuclear import, other alanine substitutions in both the alpha-helices and the beta-sheets have normal transactivation function but a loss of nuclear accumulation (i.e., functional mutants). This seeming paradox is resolved by the observations that nuclear accumulation of these functional mutants does occur but is significantly less than wild-type. This difference is revealed only in the presence of leptomycin B and actinomycin D, which permit examination of nuclear accumulation unencumbered by nuclear export and new CIITA synthesis. Further analysis of these mutants reveals that at limiting concentrations of CIITA, a dramatic difference in transactivation function between mutants and wild-type CIITA is easily detected, in agreement with their lowered nuclear accumulation. These experiments reveal an interesting aspect of LRR in controlling the amount of nuclear accumulation.
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PMID:Leucine-rich repeats of the class II transactivator control its rate of nuclear accumulation. 1207 94

Identification of the environmental triggers involved in the expression of virulence genes is a fundamental objective in studies of bacterial pathogens. For uropathogens, urea, found in the urinary tract at concentrations of up to 500 mm, functions as an environmental signal. Urea freely diffuses into the bacterium Providencia stuartii and activates UreR, a member of the AraC family of transcriptional activators. Active UreR promotes transcription of virulence-associated urease genes and alerts the organisms of its immediate milieu. Thus, the UreR.urea complex has a dual role, acting as both a transcriptional activator as well as an environmental sensor. Here, we describe the molecular events associated with activation of gene expression by urea-bound UreR. The K(d) of the urea.UreR binding reaction was measured as 0.2 mm by fluorescence quenching assays, and the shape of the binding curve indicated a single specific urea-binding site on UreR. Histidine residues are critical for urea binding in urease, and therefore to identify the urea-binding site in UreR, five mutant UreR forms were generated with histidine to alanine substitutions. Two of the mutants (UreR(c)) exhibited a constitutive phenotype by both activating transcription and binding to DNA with an increased affinity in the absence of urea. The UreR(c) bound urea with an affinity similar to that of wild-type UreR. We concluded, therefore, that the mutations resulting in constitutive activity were not involved in the UreR.urea interaction. UreR was activated, then, either by binding urea or by histidine to alanine substitutions at one of two positions. Circular dichroism indicated little change in the structure of UreR when activated, and size-exclusion chromatography demonstrated that both rUreR and rUreR(c) were dimers in both the presence and absence of urea. Thus, the structural changes associated with activation are subtle.
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PMID:Urea-dependent signal transduction by the virulence regulator UreR. 1214 87

The phosphorylation state of transcription factors is a critical determinant of their function. C/EBPbeta occurs in cells as the transcriptional activator liver-enriched activating protein (LAP) and in the truncated form liver-enriched inhibitory protein (LIP) that inhibits transcription. Analysis of C/EBPbeta phosphorylation by isoelectric focusing (IEF) shows that LAP is present in multiple forms, each with a different degree of phosphorylation in 3T3-F442A fibroblasts. Growth hormone (GH) treatment induces a new band near the negative pole, consistent with GH-promoted dephosphorylation of LAP. In addition, bands near the positive pole are rapidly and transiently induced, suggesting that GH also stimulates phosphorylation at some site(s) on LAP. C/EBPbeta contains a highly conserved MAPK consensus site that corresponds to Thr(188) in murine (m) LAP and Thr(37) in mLIP. Immunoblotting with antiphosphopeptide antibodies specific for Thr(188/37) of C/EBPbeta (anti-P-C/EBPbeta) shows that GH rapidly and transiently promotes phosphorylation of mLAP and mLIP on the MAPK site. MEK inhibitors prevent this GH-promoted phosphorylation of LAP and LIP, suggesting that such phosphorylation depends on GH-activated MAPK signaling. Mutation of Thr(235) to Ala in the homologous MAPK site of human (h) LAP (hLAPT235A) inhibits transcription mediated by the c-fos promoter in response to GH, indicating that phosphorylation at the MAPK site is required for LAP to be transcriptionally active in the context of GH-stimulated activation of the c-fos promoter. Complexes bound to the c-fos C/EBP site transiently contain C/EBPbeta phosphorylated at the MAPK site. As phosphorylation subsides, the binding of less transcriptionally active forms of LAP increases, consistent with the transient nature of c-fos stimulation by GH and other growth factors. Thus, both phosphorylation and dephosphorylation of C/EBPbeta, in response to a single physiological stimulus such as GH, coordinately modulate the ability of C/EBPbeta to activate transcription by modulating its DNA binding activity and its transactivation capacity.
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PMID:Dual regulation of phosphorylation and dephosphorylation of C/EBPbeta modulate its transcriptional activation and DNA binding in response to growth hormone. 1221 25

SoxS is the direct transcriptional activator of the superoxide regulon. SoxS recognizes a highly degenerate "soxbox" DNA sequence, and activates transcription from class I and class II promoters. SoxS is the smallest member of the AraC/XylS family of transcription regulators whose hallmark is dual helix-turn-helix (HTH) DNA-binding motifs. Evidence suggests that the N-terminal HTH motif of SoxS interacts with a highly conserved region of the soxbox termed recognition element 1 (RE1), while the C-terminal HTH motif interacts with the less conserved recognition element 2 (RE2). In the work described here, we prepared a complete library of 101 SoxS mutants containing single alanine substitutions of SoxS, and we characterized the mutant proteins in vivo and in vitro. With SoxS being closely related to MarA, we analyzed the effects of the SoxS mutations in the context of the MarA-mar crystal structure and with respect to the NMR study of MarA-DNA complexes in solution. From the properties of the alanine substitutions, we conclude the following. (1) Surface-exposed residues of helix 3 and helix 6, the recognition helices of the dual HTH motifs, are important to DNA binding and transcription activation; however, substitutions of residues predicted from the MarA-mar crystal structure to make contact with the sugar-phosphate backbone are more detrimental to DNA binding than mutations predicted to make base-specific contacts. (2) Substitution of several residues within the recognition helix predicted to make base-specific contacts with RE2 have relatively little effect on DNA-binding, suggesting the possibility of alternative protein-DNA interactions than those inferred from the MarA-mar crystal structure. (3) DNA binding and transcription activation were reduced by substitution of conserved amino acid residues comprising the hydrophobic core, presumably because they disrupt the structural integrity of SoxS. (4) Mutant K30A appears to be a positive control mutant defective in a protein-protein interaction with RNA polymerase that is required for transcription activation at all SoxS-dependent promoters because it binds and bends DNA normally but fails to activate transcription from both classes of promoters. Alanine substitutions of surface-exposed residues H3, K5, D9, S31, and V45 confer a similar phenotype. Since these residues are near K30 on the surface of the protein, the surface formed by the six residues may be used to make protein-protein interactions with RNA polymerase that are required for transcription activation at both class I and class II SoxS-dependent promoters. (5) Mutants F74A, D75A, M78A, D79A and Q85A appear to define a surface required for protein-protein interaction with RNA polymerase specifically at class II promoters because these positive control mutants bind and bend DNA normally but are defective in activation of class II promoters but not class I promoters. These SoxS mutants that bind and bend DNA normally but are defective in transcription activation represent the first positive control mutants with putative defects in protein-protein interactions with RNA polymerase among the SoxS/MarA/Rob subset of the AraC/XylS family of transcription regulators.
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PMID:A comprehensive alanine scanning mutagenesis of the Escherichia coli transcriptional activator SoxS: identifying amino acids important for DNA binding and transcription activation. 1221 88

Yeast protein kinase GCN2 stimulates the translation of transcriptional activator GCN4 by phosphorylating eIF2alpha in response to amino acid starvation. Kinase activation requires binding of uncharged tRNA to a histidyl tRNA synthetase-related domain in GCN2. Phosphorylation of serine 577 (Ser 577) in GCN2 by another kinase in vivo inhibits GCN2 function in rich medium by reducing tRNA binding activity. We show that rapamycin stimulates eIF2alpha phosphorylation by GCN2, with attendant induction of GCN4 translation, while reducing Ser 577 phosphorylation in nonstarved cells. The alanine 577 (Ala 577) mutation in GCN2 (S577A) dampened the effects of rapamycin on eIF2alpha phosphorylation and GCN4 translation, suggesting that GCN2 activation by rapamycin involves Ser 577 dephosphorylation. Rapamycin regulates the phosphorylation of Ser 577 and eIF2alpha by inhibiting the TOR pathway. Rapamycin-induced dephosphorylation of Ser 577, eIF2alpha phosphorylation, and induction of GCN4 all involve TAP42, a regulator of type 2A-related protein phosphatases. Our results add a new dimension to the regulation of protein synthesis by TOR proteins and demonstrate cross-talk between two major pathways for nutrient control of gene expression in yeast.
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PMID:Translational control by TOR and TAP42 through dephosphorylation of eIF2alpha kinase GCN2. 1265 28

Quorum sensing-dependent activation of the luminescence (lux) genes of Vibrio fischeri relies on the formation of a complex between the autoinducer molecule, N-(3-oxohexanoyl)-L-homoserine lactone, and the autoinducer-dependent transcriptional activator LuxR. In its active conformation, LuxR binds to a site known as the lux box centered at position -42.5 relative to the luxI transcriptional start site and is thought to function as an ambidextrous activator capable of making multiple contacts with RNA polymerase (RNAP). The specific role of region 4 of the Escherichia coli sigma70 subunit of RNAP in LuxR-dependent activation of the luxI promoter has been investigated. Single-round transcription assays were performed in the presence of purified LuxRDeltaN, the autoinducer-independent C-terminal domain of LuxR, and a variant RNAP which contained a C-terminally truncated sigma70 subunit devoid of region 4. Results indicated that region 4 is essential for LuxRDeltaN-dependent luxI transcription, therefore 16 single and two triple alanine substitutions in region 4.2 of sigma70 between amino acid residues 590 and 613 were examined for their effects on LuxR- and LuxRDeltaN-dependent transcription at the luxI promoter. Taken together, the analyses performed on these variants of RpoD suggest that some individual residues in region 4.2 are important to the mechanism of activator-dependent transcription initiation under investigation.
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PMID:Involvement of region 4 of the sigma70 subunit of RNA polymerase in transcriptional activation of the lux operon during quorum sensing. 1463 24

The DNA recognition sequence for the transcriptional activator, CII protein, which is critical for lysogenization by bacteriophage lambda, overlaps the -35 region of the P(RE) promoter. Data presented here show that activation by CII does not change the pattern of cleavage of the -35 region of P(RE) by iron (S)-1-(p-bromoacetamidobenzyl)-EDTA (Fe-BABE) conjugated to the sigma subunit of RNA polymerase (RNAP). Thus, the overall interaction between sigma and the -35 region of P(RE) is not significantly altered by CII. Therefore, the effects of the activator on RNAP binding to the promoter and formation of open complexes do not reflect a large-scale qualitative change in the nature of the interaction between RNAP and promoter DNA. The ability of CII to stimulate lysogenization is reduced in the presence of plasmid-borne rpoA variants encoding alanine substitutions at several positions in the C-terminal domain of the alpha subunit. However, it has not been possible to identify residues that directly affect the interaction between the activator and RNA polymerase.
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PMID:Interactions among CII protein, RNA polymerase and the lambda PRE promoter: contacts between RNA polymerase and the -35 region of PRE are identical in the presence and absence of CII protein. 1487 63

Utilization of nonfermentable carbon sources by Kluyveromyces lactis and Saccharomyces cerevisiae requires the Snf1p kinase and the Cat8p transcriptional activator, which binds to carbon source-responsive elements of target genes. We demonstrate that KlSnf1p and KlCat8p from K. lactis interact in a two-hybrid system and that the interaction is stronger with a kinase-dead mutant form of KlSnf1p. Of two putative phosphorylation sites in the KlCat8p sequence, serine 661 was identified as a key residue governing KlCat8p regulation. Serine 661 is located in the middle homology region, a regulatory domain conserved among zinc cluster transcription factors, and is part of an Snf1p consensus phosphorylation site. Single mutations at this site are sufficient to completely change the carbon source regulation of the KlCat8p transactivation activity observed. A serine-to-glutamate mutant form mimicking constitutive phosphorylation results in a nearly constitutively active form of KlCat8p, while a serine-to-alanine mutation has the reverse effect. Furthermore, it is shown that KlCat8p phosphorylation depends on KlSNF1. The Snf1-Cat8 connection is evolutionarily conserved: mutation of corresponding serine 562 of ScCat8p gave similar results in S. cerevisiae. The enhanced capacity of ScCat8S562E to suppress the phenotype caused by snf1 strengthens the hypothesis of direct phosphorylation of Cat8p by Snf1p. Unlike that of S. cerevisiae ScCAT8, KlCAT8 transcription is not carbon source regulated, illustrating the prominent role of posttranscriptional regulation of Cat8p in K. lactis.
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PMID:Key role of Ser562/661 in Snf1-dependent regulation of Cat8p in Saccharomyces cerevisiae and Kluyveromyces lactis. 1512 31

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