Proline 152 mutation in p53 abolishes cognate DNA binding, induces gain of function tumorigenesis
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE119654 | GEO | 2019/08/02
REPOSITORIES: GEO
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Project description:Proline 152 mutation in p53 abolishes cognate DNA binding, induces gain of function tumorigenesis
| PRJNA489860 | ENA
Project description:TP53 (p53) is the most commonly mutated gene in human cancers, and p53 missense mutations are present in more than 40% of all human tumors. Most p53 mutations are located within the DNA binding domain, including hotspot mutations R175H, R248W, and R273H. To elucidate the precise mechanism by which p53 missense mutants execute their gain-of-functions (GOFs) in vivo, we used a proteomic screen to identify any protein that recognizes the p53 DNA-binding domain (DBD) in a manner dependent on its mutation status. To do so, we first purified the potential binding proteins associated with the p53 DBD through multi-step affinity chromatography from a SFB-p53 H1299 stable cell line. The affinity-purified SFB-p53 interacting proteins were detected by liquid chromatography mass spectrometry/mass spectrometry (LC–MS/MS) and revealed a few cellular proteins that have been reported to interact with the DNA binding domain of p53, such as TP53BP1, USP28, and Sirt1. Interestingly, we also identified many new interacting proteins from the same complex.
2023-02-17 | PXD039886 | Pride
Project description:Mutations in the DNA binding domain (DBD) of TP53 are often associated with the gain-of-function (GOF) of mutp53, resulting in pervasive transcription of chromatin regulatory genes that promote genome-wide histone methylation and acetylation. Efforts to target this tumor-promoting function of mutp53 remain elusive. Here, we therapeutically exploit the GOF mechanisms of p53 codon 158 mutation, a DBD mutant found to be prevalent in lung squamous cell carcinoma. Using high throughput compound screening and combination analyses, we uncovered that mutp53R158G exhibits strong synergistic cytotoxicity to cisplatin-induced DNA stress and belinostat, a histone deacetylase inhibitor. This treatment regime acetylates mutp53, alters DNA binding motifs and upregulates TRAIP, a RING domain-containing E3 ubiquitin ligase which dephosphorylates IĸB and impedes nuclear translocation of RelA (p65), thus repressing oncogenic nuclear factor kappa-B (NF-ĸB) signalling and inducing apoptosis. This mechanistic intervention was validated with other acetylators of mutp53 and in several cancer models. Given that this transcriptional modulation and cytotoxic vulnerability appears inapt in p53 wild-type (WT) or null cells, our work provides a novel therapeutic opportunity in Arg158 15 -mutp53 tumors utilizing a regimen consisting of DNA-damaging agents and mutp53 acetylators, which is currently being pursued clinically
2020-02-26 | GSE129026 | GEO
Project description:Mutations in the DNA binding domain (DBD) of TP53 are often associated with the gain-of-function (GOF) of mutp53, resulting in pervasive transcription of chromatin regulatory genes that promote genome-wide histone methylation and acetylation. Efforts to target this tumor-promoting function of mutp53 remain elusive. Here, we therapeutically exploit the GOF mechanisms of p53 codon 158 mutation, a DBD mutant found to be prevalent in lung squamous cell carcinoma. Using high throughput compound screening and combination analyses, we uncovered that mutp53R158G exhibits strong synergistic cytotoxicity to cisplatin-induced DNA stress and belinostat, a histone deacetylase inhibitor. This treatment regime acetylates mutp53, alters DNA binding motifs and upregulates TRAIP, a RING domain-containing E3 ubiquitin ligase which dephosphorylates IĸB and impedes nuclear translocation of RelA (p65), thus repressing oncogenic nuclear factor kappa-B (NF-ĸB) signalling and inducing apoptosis. This mechanistic intervention was validated with other acetylators of mutp53 and in several cancer models. Given that this transcriptional modulation and cytotoxic vulnerability appears inapt in p53 wild-type (WT) or null cells, our work provides a novel therapeutic opportunity in Arg158 15 -mutp53 tumors utilizing a regimen consisting of DNA-damaging agents and mutp53 acetylators, which is currently being pursued clinically.
2020-02-26 | GSE128673 | GEO
Project description:The Forkhead transcription factor, FOXM1, is a key regulator of the cell cycle and is over-expressed in most types of cancer. FOXM1, similar to other Forkhead (FKH) factors binds to a canonical FKH motif in vitro. However, genome-wide mapping studies in different cell lines have shown a lack of enrichment of the FKH motif at FOXM1 binding sites suggesting an alternative mode of chromatin recruitment. We have investigated the role of direct versus indirect DNA binding in FOXM1 recruitment by performing ChIP-seq with WT and DNA binding deficient FOXM1. An in vitro fluorescence polarization (FP) assay was used to identify point mutations in the DNA binding domain (DBD) of FOXM1 that inhibit binding to a FKH consensus sequence. Stable cell lines expressing either WT or DBD mutant green fluorescence protein (GFP)-tagged FOXM1 were utilized for genome-wide mapping studies comparing the distribution of the DBD mutant protein to the WT. This showed that interaction of the DBD of FOXM1 with target DNA is essential for recruitment. Moreover, analysis of protein interactome of the WT versus DBD mutant FOXM1 showed that the reduced recruitment is not due to the mutation inhibiting protein-protein interactions. This study showed that a functional DBD domain is essential for FOXM1 chromatin recruitment. Even in FOXM1 mutants that show almost complete loss of binding the protein-protein interactome and pattern of phosphorylation are largely unaffected. These results strongly support a model whereby FOXM1 is specifically recruited to chromatin through co-factor interactions by binding directly to both canonical and non-canonical DNA sequences.
2015-06-11 | GSE60032 | GEO
Project description:Our lab generated the p53 K316P mouse, which mimicks a common amino acid change found in bats. The K316P mutation, found in the nuclear localization signal of p53, results in increased cytoplasmic localization of p53. We found that K316P mutant mice develop several metabolic phenotypes, including increased body fat percentage, and increased liver lipid levels. In order to determine the mechanism through which K316P mutation increases liver lipid levels, we performed metabolomic analysis of mouse livers from WT and K316P mutant mice. Mouse livers were isolated from four wild type (WT) and four p53 K316P (M) mice for lipidomic analysis. Samples were isolated and flash frozen in liquid nitrogen. Lipids were then extracted from each liver sample and analyzed using mass spectrometry.
2022-12-07 | ST002396 | MetabolomicsWorkbench
Project description:In parallel to monomeric epigenetic regulators, sequence-specific epigenetic regulators represent versatile synthetic dual-target ligands that achieve regulatory control over multi-gene networks. Development of DNA-binding domain (DBD)-HDAC inhibitors and DBD-HAT activators, which result in increased histone acetylation, has become one promising research field. However, there is no report regarding the gene regulatory pattern by sequence-specific epigenetic repressor. We report here for the first time, the synthesis of DBD-HAT inhibitors and demonstrate that these conjugates could retain their dual-target activity using predicted working model of thermal stability assay and in vitro HAT activity assay. Evaluation of antiproliferative activity in cancer cells showed that 2 (with a medium linker length of 13-atom) exhibited the highest antiproliferative activity in p53 wild-type cancer cell lines (IC50 of 1.8-2.6 uM in A549 and MV4-11 cells) and not in p53 mutant cancer cell lines. A mechanistic investigation using microarray analysis and an apoptotic assay showed that the antiproliferative effect of 2 occurred via the up-regulation of p53 target genes, and the subsequent initiation of p53-dependent apoptosis. Our research on sequence-specific dual-target epigenetic repressor offers us an alternative way to modulate HAT-governed therapeutically important genes and contributes to offer a fresh insight into antitumor therapeutics.
2017-06-28 | GSE100466 | GEO
Project description:The TP53 gene is frequently mutated in human cancer. Research has focused predominantly on six major “hotspot” codons, accounting for only ~30% of cancer-associated p53 mutations. To comprehensively characterize the consequences of the p53 mutation spectrum, we created a synthetically designed library and measured the functional impact of ~10,000 DNA-binding domain (DBD) p53 variants in human cells in culture and in vivo. Our results highlight the differential outcome of distinct p53 mutations in human patients and elucidate the selective pressure driving p53 conservation throughout evolution. Furthermore, while loss of anti-proliferative functionality largely correlates with the occurrence of cancer-associated p53 mutations, we observe that selective gain-of-function may further favor particular mutants in vivo. Finally, when combined with additional acquired p53 mutations, seemingly neutral TP53 SNPs may modulate phenotypic outcome and presumably tumor progression.
2018-06-21 | GSE115072 | GEO
Project description:Mis-sense mutations affecting TP53 promote carcinogenesis both by inactivating its tumor suppressive functions, and by conferring aberrant pro-carcinogenic activities. We report here that mis-sense mutants in the p53 DNA-binding domain (DBD) and the transactivation domain (TAD) unexpectedly activate pro-carcinogenic epidermal growth factor receptor (EGFR) signaling via distinct, previously unrecognized molecular mechanisms. DBD- and TAD-specific TP53 mutants exhibited different cellular localization patterns and induced distinct gene expression profiles. Combining mass spectrometry with drug compound screens, we identified EGFR as a major signaling factor that is stabilized by TAD and DBD mutants in the cytosolic and nuclear compartments respectively, in a tissue-independent manner. Mechanistically, TAD mutants promote EGFR-mediated signaling by enhancing EGFR interaction with AKT via DDX31 in the cytosol. Conversely, DBD mutants maintain EGFR activity in the nucleus, by blocking EGFR interaction with the phosphatase SHP1, triggering upregulation of c-Myc and Cyclin D1 levels. Therapeutically, the sensitivity of DBD mutants to EGFR inhibition is enhanced by increasing the affinity of EGFR for SHP1, while that of TAD mutants can be induced by concurrent inhibition of AKT, mTOR or PI3K signaling. Thus, our findings suggest that gain-of-function, mis-sense mutations affecting two different p53 domains promote carcinogenesis by enhancing EGFR signaling via distinctive mechanisms. Our findings imply that cancer cells bearing domain-specific mutations may have distinct and exploitable therapeutic vulnerabilities.
2023-02-17 | PXD031725 | JPOST Repository
Project description:p63, like its homologue, the tumor suppressor p53, is also able to induce apoptosis in several cancer cell types. p53 family proteins are composed of three characteristic domains which are: 1) an N-terminal transactivation domain (TAD); 2) a central DNA-binding domain (DBD); and 3) an oligomerization domain (OD). In this study, we constructed recombinant adenoviruses containing hybrid genes composed of fragments of p53 and TAp63γ genes by connecting coding sequences of their three functional domains. The potency of tumor growth suppression of these hybrid molecules was evaluated using in vitro and in vivo models. One of the p53-p63 hybrid molecules, p63-53O, was observed to be the most potent activator of human cancer cells to apoptosis when compared to the p53, TAp63γ or several alternative p53-p63 hybrid molecules. p63-53O hybrid is composed of TAD and DBD of TAp63γ and OD of p53. In an effort to identify specific targets regulated by pro-apoptotic hybrid p63-53O, we next performed Affymetrix Genechip analysis and compared expression patterns in a human osteosarcoma cell line Saos-2 transfected separately with Ad-p53, Ad-TAp63γ and Ad-p63-53O. Saos-2 osteosarcoma cells were either transduced with adenoviral vectors expressing p53, TAp63gamma, p53-p63 hybrid (p63-53O), or GFP. After 24 hours, mRNA was isolated and analyzed by hybridization to Affymetrix HG-U133 plus 2 arrays.
2012-02-04 | E-GEOD-35512 | biostudies-arrayexpress