scholarly journals p53 as a hub in cellular redox regulation and therapeutic target in cancer

2019 ◽  
Vol 11 (4) ◽  
pp. 330-341 ◽  
Author(s):  
Sofi E Eriksson ◽  
Sophia Ceder ◽  
Vladimir J N Bykov ◽  
Klas G Wiman
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Redox Homeostasis ◽  
Single Amino Acid ◽  
Cysteine Residues ◽  
Mutant P53 ◽  
Efficient Treatment ◽  
Cellular Redox ◽  
Core Domain ◽  
Cellular Environment

Abstract The TP53 tumor suppressor gene encodes a DNA-binding transcription factor that regulates multiple cellular processes including cell growth and cell death. The ability of p53 to bind to DNA and activate transcription is tightly regulated by post-translational modifications and is dependent on a reducing cellular environment. Some p53 transcriptional target genes are involved in regulation of the cellular redox homeostasis, e.g. TIGAR and GLS2. A large fraction of human tumors carry TP53 mutations, most commonly missense mutations that lead to single amino acid substitutions in the core domain. Mutant p53 proteins can acquire so called gain-of-function activities and influence the cellular redox balance in various ways, for instance by binding of the Nrf2 transcription factor, a major regulator of cellular redox state. The DNA-binding core domain of p53 has 10 cysteine residues, three of which participate in holding a zinc atom that is critical for p53 structure and function. Several novel compounds that refold and reactivate missense mutant p53 bind to specific p53 cysteine residues. These compounds can also react with other thiols and target components of the cellular redox system, such as glutathione. Dual targeting of mutant p53 and redox homeostasis may allow more efficient treatment of cancer.

scholarly journals Activation of the DNA-binding ability of human heat shock transcription factor 1 may involve the transition from an intramolecular to an intermolecular triple-stranded coiled-coil structure.

1994 ◽  
Vol 14 (11) ◽  
pp. 7557-7568 ◽  
Author(s):  
J Zuo ◽  
R Baler ◽  
G Dahl ◽  
R Voellmy
Keyword(s):  
Transcription Factor ◽  
Heat Stress ◽  
Heat Shock ◽  
Dna Binding ◽  
Hydrophobic Interactions ◽  
Coiled Coil ◽  
Heat Shock Transcription Factor ◽  
Single Amino Acid ◽  
Binding Ability ◽  
Heat Shock Element

Heat stress regulation of human heat shock genes is mediated by human heat shock transcription factor hHSF1, which contains three 4-3 hydrophobic repeats (LZ1 to LZ3). In unstressed human cells (37 degrees C), hHSF1 appears to be in an inactive, monomeric state that may be maintained through intramolecular interactions stabilized by transient interaction with hsp70. Heat stress (39 to 42 degrees C) disrupts these interactions, and hHSF1 homotrimerizes and acquires heat shock element DNA-binding ability. hHSF1 expressed in Xenopus oocytes also assumes a monomeric, non-DNA-binding state and is converted to a trimeric, DNA-binding form upon exposure of the oocytes to heat shock (35 to 37 degrees C in this organism). Because endogenous HSF DNA-binding activity is low and anti-hHSF1 antibody does not recognize Xenopus HSF, we employed this system for mapping regions in hHSF1 that are required for the maintenance of the monomeric state. The results of mutagenesis analyses strongly suggest that the inactive hHSF1 monomer is stabilized by hydrophobic interactions involving all three leucine zippers which may form a triple-stranded coiled coil. Trimerization may enable the DNA-binding function of hHSF1 by facilitating cooperative binding of monomeric DNA-binding domains to the heat shock element motif. This view is supported by observations that several different LexA DNA-binding domain-hHSF1 chimeras bind to a LexA-binding site in a heat-regulated fashion, that single amino acid replacements disrupting the integrity of hydrophobic repeats render these chimeras constitutively trimeric and DNA binding, and that LexA itself binds stably to DNA only as a dimer but not as a monomer in our assays.

scholarly journals Molecular Insights into Function and Competitive Inhibition of Pseudomonas aeruginosa Multiple Virulence Factor Regulator

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Tomoe Kitao ◽  
Francois Lepine ◽  
Seda Babloudi ◽  
Frederick Walte ◽  
Stefan Steinbacher ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Transcription Factor ◽  
Drug Discovery ◽  
Dna Binding ◽  
Quorum Sensing ◽  
Binding Domain ◽  
Antimicrobial Drug ◽  
Single Amino Acid ◽  
Persistent Infections ◽  
New Approaches

ABSTRACT New approaches to antimicrobial drug discovery are urgently needed to combat intractable infections caused by multidrug-resistant (MDR) bacteria. Multiple virulence factor regulator (MvfR or PqsR), a Pseudomonas aeruginosa quorum sensing transcription factor, regulates functions important in both acute and persistent infections. Recently identified non-ligand-based benzamine-benzimidazole (BB) inhibitors of MvfR suppress both acute and persistent P. aeruginosa infections in mice without perturbing bacterial growth. Here, we elucidate the crystal structure of the MvfR ligand binding domain (LBD) in complex with one potent BB inhibitor, M64. Structural analysis indicated that M64 binds, like native ligands, to the MvfR hydrophobic cavity. A hydrogen bond and pi interaction were found to be important for MvfR-M64 affinity. Surface plasmon resonance analysis demonstrated that M64 is a competitive inhibitor of MvfR. Moreover, a protein engineering approach revealed that Gln194 and Tyr258 are critical for the interaction between MvfR and M64. Random mutagenesis of the full-length MvfR protein identified a single-amino-acid substitution, I68F, at a DNA binding linker domain that confers M64 insensitivity. In the presence of M64, I68F but not the wild-type (WT) MvfR protein retained DNA binding ability. Our findings strongly suggest that M64 promotes conformational change at the DNA binding domain of MvfR and that the I68F mutation may compensate for this change, indicating allosteric inhibition. This work provides critical new insights into the molecular mechanism of MvfR function and inhibition that could aid in the optimization of anti-MvfR compounds and improve our understanding of MvfR regulation. IMPORTANCE Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen that causes serious acute, persistent, and relapsing infections. New approaches to antimicrobial drug discovery are urgently needed to combat intractable infections caused by this pathogen. The Pseudomonas aeruginosa quorum sensing transcription factor MvfR regulates functions important in both acute and persistent infections. We used recently identified inhibitors of MvfR to perform structural studies and reveal important insights that would benefit the optimization of anti-MvfR compounds. Altogether, the results reported here provide critical detailed mechanistic insights into the function of MvfR domains that may benefit the optimization of the chemical, pharmacological, and safety properties of MvfR antagonist series.

Activation of the DNA-binding ability of human heat shock transcription factor 1 may involve the transition from an intramolecular to an intermolecular triple-stranded coiled-coil structure

1994 ◽  
Vol 14 (11) ◽  
pp. 7557-7568
Author(s):  
J Zuo ◽  
R Baler ◽  
G Dahl ◽  
R Voellmy
Keyword(s):  
Transcription Factor ◽  
Heat Stress ◽  
Heat Shock ◽  
Dna Binding ◽  
Hydrophobic Interactions ◽  
Coiled Coil ◽  
Heat Shock Transcription Factor ◽  
Single Amino Acid ◽  
Binding Ability ◽  
Heat Shock Element

Heat stress regulation of human heat shock genes is mediated by human heat shock transcription factor hHSF1, which contains three 4-3 hydrophobic repeats (LZ1 to LZ3). In unstressed human cells (37 degrees C), hHSF1 appears to be in an inactive, monomeric state that may be maintained through intramolecular interactions stabilized by transient interaction with hsp70. Heat stress (39 to 42 degrees C) disrupts these interactions, and hHSF1 homotrimerizes and acquires heat shock element DNA-binding ability. hHSF1 expressed in Xenopus oocytes also assumes a monomeric, non-DNA-binding state and is converted to a trimeric, DNA-binding form upon exposure of the oocytes to heat shock (35 to 37 degrees C in this organism). Because endogenous HSF DNA-binding activity is low and anti-hHSF1 antibody does not recognize Xenopus HSF, we employed this system for mapping regions in hHSF1 that are required for the maintenance of the monomeric state. The results of mutagenesis analyses strongly suggest that the inactive hHSF1 monomer is stabilized by hydrophobic interactions involving all three leucine zippers which may form a triple-stranded coiled coil. Trimerization may enable the DNA-binding function of hHSF1 by facilitating cooperative binding of monomeric DNA-binding domains to the heat shock element motif. This view is supported by observations that several different LexA DNA-binding domain-hHSF1 chimeras bind to a LexA-binding site in a heat-regulated fashion, that single amino acid replacements disrupting the integrity of hydrophobic repeats render these chimeras constitutively trimeric and DNA binding, and that LexA itself binds stably to DNA only as a dimer but not as a monomer in our assays.

scholarly journals Two monomers of yeast transcription factor ADR1 bind a palindromic sequence symmetrically to activate ADH2 expression.

1991 ◽  
Vol 11 (3) ◽  
pp. 1566-1577 ◽  
Author(s):  
S K Thukral ◽  
A Eisen ◽  
E T Young
Keyword(s):  
Transcription Factor ◽  
Amino Acid ◽  
Dna Binding ◽  
Dimethyl Sulfate ◽  
Binding Activity ◽  
Single Amino Acid ◽  
Minor Effect ◽  
Palindromic Sequence ◽  
Amino Terminal ◽  
Dna Binding Activity

ADR1 is a transcription factor from Saccharomyces cerevisiae that regulates ADH2 expression through a 22-bp palindromic sequence (UAS1). Size fractionation studies revealed that full-length ADR1 and a truncated ADR1 protein containing the first 229 amino acids, which has the complete DNA-binding domain, ADR1:17-229, exist as monomers in solution. However, two complexes were formed with target DNA-binding sites. UV-cross-linking studies suggested that these two complexes represent one and two molecules of ADR1 bound to DNA. Studies of ADR1 complexes formed with wild-type UAS1, asymmetrically altered UAS1, and one half of UAS1 showed that ADR1 can bind to one half of UAS1 and gives rise to a complex containing one molecule of ADR1. Dimethyl sulfate interference studies were consistent with this interpretation and in addition indicated that purine contact sites in each half of UAS1 were identical. Increasing the distance between the two halves of UAS1 had at most a minor effect of the thermodynamics of formation of the two complexes. These data are more consistent with ADR1 binding as two independent monomers, one to each half of UAS1. However, binding of two ADR1 monomers at UAS1 is apparently essential for transactivation in vivo. Further, we have identified a stretch of 18 amino acid residues amino terminal to the zinc two-finger domains of ADR1 which is essential for DNA-binding activity. Single amino acid substitutions of residues in this region resulted in severely reduced DNA-binding activity.

scholarly journals Preferences in a trait decision determined by transcription factor variants

2018 ◽  
Vol 115 (34) ◽  
pp. E7997-E8006 ◽  
Author(s):  
Michael W. Dorrity ◽  
Josh T. Cuperus ◽  
Jolie A. Carlisle ◽  
Stanley Fields ◽  
Christine Queitsch
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Fungal Pathogens ◽  
Binding Domain ◽  
Single Amino Acid ◽  
Environmental Cues ◽  
Dna Binding Domain ◽  
Independent Manner ◽  
Dna Binding Specificity ◽  
The Many

Few mechanisms are known that explain how transcription factors can adjust phenotypic outputs to accommodate differing environments. In Saccharomyces cerevisiae, the decision to mate or invade relies on environmental cues that converge on a shared transcription factor, Ste12. Specificity toward invasion occurs via Ste12 binding cooperatively with the cofactor Tec1. Here, we determine the range of phenotypic outputs (mating vs. invasion) of thousands of DNA-binding domain variants in Ste12 to understand how preference for invasion may arise. We find that single amino acid changes in the DNA-binding domain can shift the preference of yeast toward either mating or invasion. These mutations define two distinct regions of this domain, suggesting alternative modes of DNA binding for each trait. We characterize the DNA-binding specificity of wild-type Ste12 to identify a strong preference for spacing and orientation of both homodimeric and heterodimeric sites. Ste12 mutants that promote hyperinvasion in a Tec1-independent manner fail to bind cooperative sites with Tec1 and bind to unusual dimeric Ste12 sites composed of one near-perfect and one highly degenerate site. We propose a model in which Ste12 alone may have evolved to activate invasion genes, which could explain how preference for invasion arose in the many fungal pathogens that lack Tec1.

scholarly journals Structural basis of reactivation of oncogenic p53 mutants by a small molecule: methylene quinuclidinone (MQ)

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Oksana Degtjarik ◽  
Dmitrij Golovenko ◽  
Yael Diskin-Posner ◽  
Lars Abrahmsén ◽  
Haim Rozenberg ◽  
...  
Keyword(s):  
Genotoxic Stress ◽  
Biologically Active ◽  
Structural Basis ◽  
Cysteine Residues ◽  
Mutant P53 ◽  
Tumor Suppressor P53 ◽  
Gene Encoding ◽  
Core Domain ◽  
Expression Of Genes ◽  
P53 Reactivation

AbstractIn response to genotoxic stress, the tumor suppressor p53 acts as a transcription factor by regulating the expression of genes critical for cancer prevention. Mutations in the gene encoding p53 are associated with cancer development. PRIMA-1 and eprenetapopt (APR-246/PRIMA-1MET) are small molecules that are converted into the biologically active compound, methylene quinuclidinone (MQ), shown to reactivate mutant p53 by binding covalently to cysteine residues. Here, we investigate the structural basis of mutant p53 reactivation by MQ based on a series of high-resolution crystal structures of cancer-related and wild-type p53 core domains bound to MQ in their free state and in complexes with their DNA response elements. Our data demonstrate that MQ binds to several cysteine residues located at the surface of the core domain. The structures reveal a large diversity in MQ interaction modes that stabilize p53 and its complexes with DNA, leading to a common global effect that is pertinent to the restoration of non-functional p53 proteins.

scholarly journals Balance between mutually exclusive traits shifted by variants of a yeast transcription factor

2017 ◽  
Author(s):  
Michael W. Dorrity ◽  
Josh T. Cuperus ◽  
Jolie A. Carlisle ◽  
Stanley Fields ◽  
Christine Queitsch
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factor ◽  
Amino Acid ◽  
Dna Binding ◽  
Single Amino Acid ◽  
Environmental Cues ◽  
Dna Binding Domain ◽  
Independent Manner ◽  
Half Site

AbstractIn Saccharomyces cerevisiae, the decision to mate or invade relies on environmental cues that converge on a shared transcription factor, Ste12. Specificity toward invasion occurs via Ste12 binding cooperatively with the co-factor Tec1. Here, we characterize the in vitro binding preferences of Ste12 to identify a defined spacing and orientation of dimeric sites, one that is common in pheromone-regulated genes. We find that single amino acid changes in the DNA-binding domain of Ste12 can shift the preference of yeast toward either mating or invasion. These mutations define two distinct regions of this domain, suggesting alternative modes of DNA binding for each trait. Some exceptional Ste12 mutants promote hyperinvasion in a Tec1-independent manner; these fail to bind cooperative sites with Tec1 and bind to unusual dimeric Ste12 sites that contain one highly degenerate half site. We propose a model for how activation of invasion genes could have evolved with Ste12 alone.

Two monomers of yeast transcription factor ADR1 bind a palindromic sequence symmetrically to activate ADH2 expression

1991 ◽  
Vol 11 (3) ◽  
pp. 1566-1577
Author(s):  
S K Thukral ◽  
A Eisen ◽  
E T Young
Keyword(s):  
Transcription Factor ◽  
Amino Acid ◽  
Dna Binding ◽  
Dimethyl Sulfate ◽  
Binding Activity ◽  
Single Amino Acid ◽  
Minor Effect ◽  
Palindromic Sequence ◽  
Amino Terminal ◽  
Dna Binding Activity

ADR1 is a transcription factor from Saccharomyces cerevisiae that regulates ADH2 expression through a 22-bp palindromic sequence (UAS1). Size fractionation studies revealed that full-length ADR1 and a truncated ADR1 protein containing the first 229 amino acids, which has the complete DNA-binding domain, ADR1:17-229, exist as monomers in solution. However, two complexes were formed with target DNA-binding sites. UV-cross-linking studies suggested that these two complexes represent one and two molecules of ADR1 bound to DNA. Studies of ADR1 complexes formed with wild-type UAS1, asymmetrically altered UAS1, and one half of UAS1 showed that ADR1 can bind to one half of UAS1 and gives rise to a complex containing one molecule of ADR1. Dimethyl sulfate interference studies were consistent with this interpretation and in addition indicated that purine contact sites in each half of UAS1 were identical. Increasing the distance between the two halves of UAS1 had at most a minor effect of the thermodynamics of formation of the two complexes. These data are more consistent with ADR1 binding as two independent monomers, one to each half of UAS1. However, binding of two ADR1 monomers at UAS1 is apparently essential for transactivation in vivo. Further, we have identified a stretch of 18 amino acid residues amino terminal to the zinc two-finger domains of ADR1 which is essential for DNA-binding activity. Single amino acid substitutions of residues in this region resulted in severely reduced DNA-binding activity.

scholarly journals Progressive Rotavirus Infection Downregulates Redox-Sensitive Transcription Factor Nrf2 and Nrf2-Driven Transcription Units

2020 ◽  
Vol 2020 ◽  
pp. 1-48
Author(s):  
Upayan Patra ◽  
Urbi Mukhopadhyay ◽  
Arpita Mukherjee ◽  
Rakesh Sarkar ◽  
Mamta Chawla-Sarkar
Keyword(s):  
Transcription Factor ◽  
Stress Response ◽  
Antioxidant Defense ◽  
Cellular Stress Response ◽  
Erythroid Cell ◽  
Heme Oxygenase 1 ◽  
Related Factor ◽  
Defense System ◽  
Cellular Redox ◽  
Cellular Environment

Eukaryotic cells adopt highly tuned stress response physiology under threats of exogenous stressors including viruses to maintain cellular homeostasis. Not surprisingly, avoidance of cellular stress response pathways is an essential facet of virus-induced obligatory host reprogramming to invoke a cellular environment conducive to viral perpetuation. Adaptive cellular responses to oxidative and electrophilic stress are usually taken care of by an antioxidant defense system, core to which lies the redox-responsive transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2-driven transcriptional cascade. Deregulation of host redox balance and redox stress-sensitive Nrf2 antioxidant defense have been reported for many viruses. In the current study, we aimed to study the modulation of the Nrf2-based host cellular redox defense system in response to Rotavirus (RV) infection in vitro. Interestingly, we found that Nrf2 protein levels decline sharply with progression of RV infection beyond an initial upsurge. Moreover, Nrf2 decrease as a whole was found to be accompanied by active nuclear vacuity of Nrf2, resulting in lowered expression of stress-responsive Nrf2 target genes heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1, and superoxide dismutase 1 both in the presence and absence of Nrf2-driven transcriptional inducers. Initial induction of Nrf2 concurred with RV-induced early burst of oxidative stress and therefore was sensitive to treatments with antioxidants. Reduction of Nrf2 levels beyond initial hours, however, was found to be independent of the cellular redox status. Furthermore, increasing the half-life of Nrf2 through inhibition of the Kelch-like erythroid cell-derived protein with CNC homology- (ECH-) associated protein 1/Cullin3-RING Box1-based canonical Nrf2 turnover pathway could not restore Nrf2 levels post RV-SA11 infection. Depletion of the Nrf2/HO-1 axis was subsequently found to be sensitive to proteasome inhibition with concurrent observation of increased K48-linked ubiquitination associated with Nrf2. Together, the present study describes robust downregulation of Nrf2-dependent cellular redox defense beyond initial hours of RV infection, justifying our previous observation of potent antirotaviral implications of Nrf2 agonists.

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