scholarly journals Engineering Optogenetic Control of Endogenous p53 Protein Levels

2019 ◽  
Vol 9 (10) ◽  
pp. 2095 ◽  
Author(s):  
Pierre Wehler ◽  
Barbara Di Ventura
Keyword(s):  
Blue Light ◽  
Light Exposure ◽  
P53 Protein ◽  
P53 Gene ◽  
Protein Levels ◽  
Cycle Arrest ◽  
P53 Activation ◽  
Low Levels ◽  
Export System ◽  
Transcription Factor P53

The transcription factor p53 is a stress sensor that turns specific sets of genes on to allow the cell to respond to the stress depending on its severity and type. p53 is classified as tumor suppressor because its function is to maintain genome integrity promoting cell cycle arrest, apoptosis, or senescence to avoid proliferation of cells with damaged DNA. While in many human cancers the p53 gene is itself mutated, there are some in which the dysfunction of the p53 pathway is caused by the overexpression of negative regulators of p53, such as Mdm2, that keep it at low levels at all times. Here we develop an optogenetic approach to control endogenous p53 levels with blue light. Specifically, we control the nuclear localization of the Mmd2-binding PMI peptide using the light-inducible export system LEXY. In the dark, the PMI-LEXY fusion is nuclear and binds to Mdm2, consenting to p53 to accumulate and transcribe the target gene p21. Blue light exposure leads to the export of the PMI-LEXY fusion into the cytosol, thereby Mdm2 is able to degrade p53 as in the absence of the peptide. This approach may be useful to study the effect of localized p53 activation within a tissue or organ.

The role of the p53 protein in the apoptotic response

1994 ◽  
Vol 345 (1313) ◽  
pp. 277-280 ◽  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Mammalian Cells ◽  
P53 Protein ◽  
P53 Gene ◽  
Chemotherapeutic Agents ◽  
Apoptotic Pathway ◽  
Cycle Arrest ◽  
Cell Death Pathway ◽  
Binding Function

When mammalian cells or tissues are exposed to DNA damaging agents a programmed cell death pathway is induced as well as a cell cycle arrest. In mice in which the p53 gene has been inactivated by homologous recombination this response is profoundly diminished. These mice develop normally so that developmentally induced apoptotic events do not require p53. The p53 gene product is a 393 amino acid nuclear protein that binds specifically to DNA and can act as a positive transcription factor. High levels of p53 can induce the transcription of gene products involved in the cell cycle arrest and apoptotic pathway. The p53 proteins activity is very tightly controlled both by allosteric regulation of its DNA binding function and by regulation of the protein’s stability. These results are discussed in the context of the mutations in p53 found in human tumours and their implications for the treatment of the disease by the use of radiation and chemotherapeutic agents that target DNA.

scholarly journals High-expression of ROCK1 modulates the apoptosis of lens epithelial cells in age-related cataracts by targeting p53 gene

2020 ◽  
Author(s):  
Shanshan Hu ◽  
Dongmei Su ◽  
Lei Sun ◽  
Zhongying Wang ◽  
Lina Guan ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Protein Level ◽  
P53 Protein ◽  
P53 Gene ◽  
Lens Epithelial Cells ◽  
Protein Levels ◽  
P53 Signaling ◽  
Age Related ◽  
Gene And Protein Expression

Abstract Background: Age-related cataract (ARC) is a serious visual impairment disease, and its pathogenesis is unclear. This article aims to investigate the role of ROCK1 in the apoptosis of lens epithelial cells (LECs) in age-related cataracts. Methods: We collect anterior capsule samples from normal people, patients with age-related cataracts, young mice and naturally aging cataract mice. The Oxidative stress-induced apoptosis model was constructed by cultivating HLE-B3 cells with H2O2. MTT, Hoechst 33342, and TUNEL assay were performed to explore proliferation and apoptosis. HE assay was used to observe cell morphology. The gene and protein expression were assessed by quantitative real-time PCR, western blot, immunofluorescence, and immunohistochemical staining.Result: The results from the clinic and mice experiments showed that the numbers of lens epithelial cells from cataract individuals were less than the control individuals. In vitro, the apoptotic cells were increased in lens epithelial cells under H2O2 treatment. The ROCK1 protein level increased in the lens epithelial cells from age-related cataract patients and the old mice, respectively. Meanwhile, the up-regulation of the ROCK1 gene was associated with H2O2-induced HLE-B3 cells apoptosis. MTT and apoptosis assay showed ROCK1 was necessary in mediating H2O2-induced lens epithelial cells apoptosis through ROCK1 over-expression and knockdown experiment, respectively. Further investigation showed that p53 protein levels had been increased during ROCK1-mediated apoptosis in response to H2O2. Besides, ROCK1 phosphorylated p53 at ser15 to up-regulate its protein level. Conclusions: This study established the novel association of ROCK1/p53 signaling with lens epithelial cells apoptosis and age-related cataract genesis.

Uteroplacental insufficiency increases p53 phosphorylation without triggering the p53-MDM2 functional circuit response in the IUGR rat kidney

2006 ◽  
Vol 291 (2) ◽  
pp. R412-R418 ◽  
Author(s):  
Mariana Baserga ◽  
Merica A. Hale ◽  
Xingrao Ke ◽  
Zeng Ming Wang ◽  
Xing Yu ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Posttranslational Modifications ◽  
Rat Kidney ◽  
P53 Protein ◽  
P53 Gene ◽  
Artery Ligation ◽  
Pregnant Rat ◽  
Protein Levels ◽  
Amino Terminal ◽  
Induced Apoptosis

Uteroplacental insufficiency (UPI) leads to intrauterine growth restriction (IUGR), which predisposes infants toward renal insufficiency early in life and increases the risk of kidney-related adult morbidities, such as hypertension. This compromised in utero environment has been demonstrated to impair nephrogenesis, as evidenced by a reduced nephron endowment in humans and in rats rendered IUGR by UPI. Concordantly, we have observed that IUGR rats have increased kidney p53 protein levels associated with increased apoptosis. Several factors can regulate p53 gene expression and activity, including posttranslational modifications and protein-protein interactions in the cell. Among these, two important mechanisms are 1) phosphorylation of the amino terminal serine 15 [phospho-p53 (Ser15)], which increases p53 stability and apoptotic activity, and 2) the murine double-minute (MDM2) functional circuit that limits further p53-induced apoptosis by promoting proteosomal degradation of p53. We hypothesize that UPI induces an increase in phospho-p53 (Ser15) in association with an absent MDM2 response, predisposing the kidney to increased apoptosis. To test our hypothesis, we induced IUGR through bilateral uterine artery ligation of the pregnant rat. UPI significantly increased phospho-p53 (Ser15), as well as ataxia teleangiectasia-mutated kinase/A-T-related kinase and dsDNA-activated protein kinase kinase levels, which induce phosphorylation of p53. In contrast, UPI induced no increase in kidney MDM2 mRNA and protein levels in IUGR pups. We conclude that among multiple mechanisms that affect nephrogenesis, UPI induces an increase in p53 phosphorylation without a corresponding increase in MDM2 expression, and we speculate that this response may contribute to the increased apoptosis previously described in the IUGR kidney.

scholarly journals Mdm2 Is Required for Inhibition of Cdk2 Activity by p21, Thereby Contributing to p53-Dependent Cell Cycle Arrest

2007 ◽  
Vol 27 (11) ◽  
pp. 4166-4178 ◽  
Author(s):  
Luciana E. Giono ◽  
James J. Manfredi
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Cycle Progression ◽  
Target Genes ◽  
P53 Protein ◽  
Protein Levels ◽  
Cycle Arrest ◽  
Cdk2 Activity ◽  
Dependent Cell ◽  
Interfering Rna

ABSTRACT p53 is extensively posttranslationally modified in response to various types of cellular stress. Such modifications have been implicated in the regulation of p53 protein levels as well as its DNA binding and transcriptional activities. Treatment of cells with doxorubicin causes phosphorylation and acetylation of p53, transcriptional upregulation of p21 and other target genes, and growth arrest. In contrast, downregulation of Mdm2 by a small interfering RNA (siRNA) approach led to increased levels of p53 lacking phosphorylation at serine 15 and acetylation at lysine 382. Levels of binding of p53 to the p21 promoter were comparable following treatment with doxorubicin or Mdm2 siRNA. Moreover, p53 was transcriptionally active and capable of inducing or repressing a variety of its target genes. Surprisingly, p53 upregulated by Mdm2 siRNA had no effect on cell cycle progression. Although comparable in level to that achieved by treatment with the p53 activators actinomycin D and nutlin-3, the increases in p53 and p21 after downregulation of Mdm2 were not sufficient to trigger cell cycle arrest. This version of p21 was capable of interacting with cyclin-dependent kinase 2 (Cdk2) but failed to inhibit its activity. Taken together, these results argue that Mdm2 is needed for full inhibition of Cdk2 activity by p21, thereby positively contributing to p53-dependent cell cycle arrest.

scholarly journals MDM2 antagonists induce p53-dependent apoptosis in AML: implications for leukemia therapy

Blood ◽  
2005 ◽  
Vol 106 (9) ◽  
pp. 3150-3159 ◽  
Author(s):  
Kensuke Kojima ◽  
Marina Konopleva ◽  
Ismael J. Samudio ◽  
Masato Shikami ◽  
Maria Cabreira-Hansen ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Transcriptional Activation ◽  
P53 Protein ◽  
Negative Regulator ◽  
Wild Type ◽  
Protein Levels ◽  
P53 Signaling ◽  
P53 Activation ◽  
Induction Of Apoptosis ◽  
Wild Type P53

AbstractAlthough TP53 mutations are rare in acute myeloid leukemia (AML), inactivation of wild-type p53 protein frequently occurs through overexpression of its negative regulator MDM2 (murine double minute 2). Recently, small-molecule antagonists of MDM2, Nutlins, have been developed that inhibit the p53-MDM2 interaction and activate p53 signaling. Here, we study the effects of p53 activation by Nutlin-3 in AML cells. Treatment with MDM2 inhibitor triggered several molecular events consistent with induction of apoptosis: loss of mitochondrial membrane potential, caspase activation, phosphatidylserine externalization, and DNA fragmentation. There was a positive correlation in primary AML samples with wild-type p53 between baseline MDM2 protein levels and apoptosis induced by MDM2 inhibition. No induction of apoptosis was observed in AML samples harboring mutant p53. Colony formation of AML progenitors was inhibited in a dose-dependent fashion, whereas normal CD34+ progenitor cells were less affected. Mechanistic studies suggested that Nutlin-induced apoptosis was mediated by both transcriptional activation of proapoptotic Bcl-2 family proteins, and transcription-independent mitochondrial permeabilization resulting from mitochondrial p53 translocation. MDM2 inhibition synergistically enhanced cytotoxicity of cytosine arabinoside and doxorubicin in AML blasts but not in normal hematopoietic progenitor cells. p53 activation by targeting the p53-MDM2 interaction might offer a novel therapeutic strategy for AML that retain wild-type p53.

scholarly journals High-expression of ROCK1 modulates the apoptosis of lens epithelial cells in age-related cataracts by targeting p53 gene

2020 ◽  
Vol 26 (1) ◽  
Author(s):  
Shanshan Hu ◽  
Dongmei Su ◽  
Lei Sun ◽  
Zhongying Wang ◽  
Lina Guan ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Protein Level ◽  
P53 Protein ◽  
P53 Gene ◽  
Lens Epithelial Cells ◽  
Protein Levels ◽  
P53 Signaling ◽  
Age Related ◽  
Gene And Protein Expression

Abstract Background Age-related cataract (ARC) is a serious visual impairment disease, and its pathogenesis is unclear. This article aims to investigate the role of ROCK1 in the apoptosis of lens epithelial cells (LECs) in age-related cataracts. Methods We collect anterior capsule samples from normal people, patients with age-related cataracts, young mice and naturally aging cataract mice. The oxidative stress-induced apoptosis model was constructed by cultivating HLE-B3 cells with H2O2. MTT, Hoechst 33342, and TUNEL assay were performed to explore proliferation and apoptosis. HE assay was used to observe cell morphology. The gene and protein expression were assessed by quantitative real-time PCR, western blot, immunofluorescence, and immunohistochemical staining. Result The results from the clinic and mice experiments showed that the numbers of lens epithelial cells from cataract individuals were less than the control individuals. In vitro, the apoptotic cells were increased in lens epithelial cells under H2O2 treatment. The ROCK1 protein level increased in the lens epithelial cells from age-related cataract patients and the old mice, respectively. Meanwhile, the up-regulation of the ROCK1 gene was associated with H2O2-induced HLE-B3 cells apoptosis. MTT and apoptosis assay showed ROCK1 was necessary in mediating H2O2-induced lens epithelial cells apoptosis through ROCK1 over-expression and knockdown experiment, respectively. Further investigation showed that p53 protein levels had been increased during ROCK1-mediated apoptosis in response to H2O2. Besides, ROCK1 phosphorylated p53 at ser15 to up-regulate its protein level. Conclusions This study established the novel association of ROCK1/p53 signaling with lens epithelial cells apoptosis and age-related cataract genesis.

scholarly journals Interferon-Inducible Protein IFIXα1 Functions as a Negative Regulator of HDM2

2006 ◽  
Vol 26 (5) ◽  
pp. 1979-1996 ◽  
Author(s):  
Yi Ding ◽  
Jin-Fong Lee ◽  
Hua Lu ◽  
Mong-Hong Lee ◽  
Duen-Hwa Yan
Keyword(s):  
Amino Acid ◽  
Gene Family ◽  
Tumor Suppression ◽  
P53 Protein ◽  
Negative Regulator ◽  
Breast Cancer Cell Lines ◽  
Homologous Proteins ◽  
Protein Levels ◽  
P53 Activation ◽  
Inducible Protein

ABSTRACT The 200-amino-acid repeat (HIN-200) gene family with the hematopoietic interferon (IFN)-inducible nuclear protein encodes highly homologous proteins involved in cell growth, differentiation, autoimmunity, and tumor suppression. IFIX is the newest member of the human HIN-200 family and is often downregulated in breast tumors and breast cancer cell lines. The expression of the longest isoform of IFIX gene products, IFIXα1, is associated with growth inhibition, suppression of transformation, and tumorigenesis. However, the mechanism underlying the tumor suppression activity of IFIXα1 is not well understood. Here, we show that IFIXα1 downregulates HDM2, a principal negative regulator of p53, at the posttranslational level. IFIXα1 destabilizes HDM2 protein and promotes its ubiquitination. The E3 ligase activity of HDM2 appears to be required for this IFIXα1 effect. Importantly, HDM2 downregulation is required for the IFIXα1-mediated increase of p53 protein levels, transcriptional activity, and nuclear localization, suggesting that IFIXα1 positively regulates p53 by acting as a negative regulator of HDM2. We found that IFIXα1 interacts with HDM2. Interestingly, the signature motif of the HIN-200 gene family, i.e., the 200-amino-acid HIN domain of IFIXα1, is sufficient not only for binding HDM2 but also for downregulating it, leading to p53 activation. Finally, we show that IFIX mediates HDM2 downregulation in an IFN-inducible system. Together, these results suggest that IFIXα1 functions as a tumor suppressor by repressing HDM2 function.

scholarly journals Transcriptional Activation of p53 during Cold Induced Torpor in the 13-Lined Ground SquirrelIctidomys tridecemlineatus

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Joshua Hefler ◽  
Cheng-Wei Wu ◽  
Kenneth B. Storey
Keyword(s):  
Skeletal Muscle ◽  
Transcriptional Activation ◽  
Ground Squirrel ◽  
Cellular Response ◽  
P53 Protein ◽  
Binding Activity ◽  
Protein Levels ◽  
Dna Binding Activity ◽  
Study Results ◽  
Transcription Factor P53

The transcription factor p53 is located at the centre of multiple pathways relating the cellular response to stress. Commonly known as a tumor suppressor, it is responsible for initiating diverse actions to protect the integrity of the genome, ranging from cell cycle arrest to apoptosis. This study investigated the regulation of p53 protein in hibernating 13-lined ground squirrelIctidomys tridecemlineatusduring multiple stages of the torpor-arousal cycle. Transcript and protein levels of p53 were both elevated in the skeletal muscle during early and late torpor stages of the hibernation cycle. Nuclear localization of p53 was also increased during late torpor, and this is associated with an increase in its DNA binding activity and expression of p53 transcriptional targetsp21CIP,gadd45α, and14-3-3σ. The increase in p53 transcriptional activity appears to be independent of its phosphorylation at Ser-15, Ser-46, and Ser-392, consistent with an absence of checkpoint kinase activation during torpor. Sequence analysis revealed unique amino acid substitutions in the ground squirrel p53 protein, which may contribute to an increase in protein stability compared to nonhibernators. Overall, the study results provided evidences for a potential role of p53 in the protection of the skeletal muscle during torpor.

scholarly journals The correlation between the use of personal protective equipment and level wild-type p53 of dental technicians in Surabaya

2017 ◽  
Vol 50 (1) ◽  
pp. 19 ◽  
Author(s):  
Puspa Dila Rohmaniar ◽  
Titiek Berniyanti ◽  
Retno Pudji Rahayu
Keyword(s):  
Personal Protective Equipment ◽  
P53 Protein ◽  
Pearson Correlation ◽  
P53 Gene ◽  
Working Environment ◽  
Protective Equipment ◽  
Wild Type ◽  
Cross Sectional ◽  
Protein Levels ◽  
Wild Type P53

Background: Exposure of metals among dental technicians that come from the working environment can lead to the formation reactive oxygen species (ROS). ROS can cause mutations in the p53 gene (p53). The mutation is transversion mutation GuanineThymine. p53 mutations can lead to low expression of the wild-type p53 protein (p53). Wild-type p53 involved in many biological processes such as regulation of genes involved in cell cycle, cell growth after DNA damage, and apoptosis. However, exposure to metals among dental technicians can be prevented through the use of personal protective equipment (PPE) during work. Purpose: The purpose of this study was to analyze the correlation between the use of personal protective equipment to wild-type p53 protein levels among dental technicians in Surabaya. Method: This study was observational analytic with cross sectional approach. 40 samples were taken by random sampling. Data were retrieved through interviews and observations. Wild-type p53 was analyzed from saliva with indirect ELISA method. Analysis of data used Kolmogorov Smirnov normality test and a Pearson correlation test. Value significance was p<0.05 (95% confidence level). Result: There was a significant association between the use of personal protective equipment with wild-type p53 levels with p=0.002 Conclusion: The use PPE properly is positively correlated with the wild-type p53 protein levels of dental technicians in Surabaya.

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