P53 Deficiency Research Articles

Unveiling RACK1: a key regulator of the PI3K/AKT pathway in prostate cancer development.

The dysregulated PI3K/AKT pathway is pivotal in the onset and progression of various cancers, including prostate cancer. However, targeting this pathway directly poses challenges due to compensatory upregulation of alternative oncogenic pathways. This study focuses on the novel regulatory activity of the Receptor for Activated Protein Kinase (RACK1), a scaffolding/adaptor protein, in governing the PI3K/AKT pathway within prostate cancer. Through a genetic mouse model, our research unveils RACK1's pivotal role in orchestrating AKT activation and the genesis of prostate cancer. RACK1 deficiency hampers AKT activation, effectively impeding prostate tumor formation induced by PTEN and p53 deficiency. Mechanistically, RACK1 facilitates AKT membrane translocation and fosters its interaction with mTORC2, thereby promoting AKT activation and subsequent tumor cell proliferation and tumor formation. Notably, inhibiting AKT activation via RACK1 deficiency does not trigger feedback upregulation of HER3 and androgen receptor (AR) expression and activation, distinguishing it from direct PI3K or AKT targeting. These findings position RACK1 as a critical regulator of the PI3K/AKT pathway and a promising target for curtailing prostate cancer development arising from pathway aberrations.

R-loop formation contributes to mTORC1 activation-dependent DNA replication stress induced by p53 deficiency

R-loop formation contributes to mTORC1 activation-dependent DNA replication stress induced by p53 deficiency

DDX41 dissolves G-quadruplexes to maintain erythroid genome integrity and prevent cGAS-mediated cell death.

Deleterious germline DDX41 variants constitute the most common inherited predisposition disorder linked to myeloid neoplasms (MNs). The role of DDX41 in hematopoiesis and how its germline and somatic mutations contribute to MNs remain unclear. Here we show that DDX41 is essential for erythropoiesis but dispensable for the development of other hematopoietic lineages. Using stage-specific Cre models for erythropoiesis, we reveal that Ddx41 knockout in early erythropoiesis is embryonically lethal, while knockout in late-stage terminal erythropoiesis allows mice to survive with normal blood counts. DDX41 deficiency induces a significant upregulation of G-quadruplexes (G4), noncanonical DNA structures that tend to accumulate in the early stages of erythroid precursors. We show that DDX41 co-localizes with G4 on the erythroid genome. DDX41 directly binds to and dissolves G4, which is significantly compromised in MN-associated DDX41 mutants. Accumulation of G4 by DDX41 deficiency induces erythroid genome instability, defects in ribosomal biogenesis, and upregulation of p53. However, p53 deficiency does not rescue the embryonic death of Ddx41 hematopoietic-specific knockout mice. In parallel, genome instability also activates the cGas-Sting pathway, which is detrimental to survival since cGas-deficient and hematopoietic-specific Ddx41 knockout mice are viable without detectable hematologic phenotypes, although these mice continue to show erythroid ribosomal defects and upregulation of p53. These findings are further supported by data from a DDX41 mutated MN patient and human iPSC-derived bone marrow organoids. Our study establishes DDX41 as a G4 dissolver, essential for erythroid genome stability and suppressing the cGAS-STING pathway.

Combined inhibition of CTPS1 and ATR is a metabolic vulnerability in p53-deficient myeloma cells.

In multiple myeloma, as in B-cell malignancies, mono- and especially bi-allelic TP53 gene inactivation is a high-risk factor for treatment resistance, and there are currently no therapies specifically targeting p53 deficiency. In this study, we evaluated if the loss of cell cycle control in p53-deficient myeloma cells would confer a metabolically actionable vulnerability. We show that CTP synthase 1 (CTPS1), which encodes a CTP synthesis rate-limiting enzyme essential for DNA and RNA synthesis in lymphoid cells, is overexpressed in samples from myeloma patients displaying a high proliferation rate (high MKI67 expression) or a low p53 score (synonymous with TP53 deletion and/or mutation). This overexpression of CTPS1 was associated with reduced survival in two cohorts. Using scRNA-seq analysis in 24 patient samples, we further demonstrate that myeloma cells in the S or G2/M phase display high CTPS1 expression. Pharmacological inhibition of CTPS1 by STP-B induced cell cycle arrest in early S phase in isogenic NCI-H929 or XG7 TP53 +/+, TP53 -/-, and TP53 R175H/R175H cells and in a TP53 -/R123STOP patient sample. The functional annotation of transcriptional changes in 10 STP-B-treated myeloma cell lines revealed a decrease in protein translation and confirmed the blockade of cells into the S phase. The pharmacological inhibition of ATR, which governs the intrinsic S/G2 checkpoint, in STP-B-induced S-phase arrested cells synergistically induced cell death in TP53 +/+, TP53 -/-, and TP53 R175H/R175H isogenic cell lines (Bliss score >15). This combination induced replicative stress and caspase-mediated cell death and was highly effective in resistant/refractory patient samples with TP53 deletion and/or mutation and in TP53 -/- NCI-H929 xenografted NOD-scid IL2Rgamma mice. Our in vitro, ex vivo, and in vivo data provide the rationale for combined CTPS1 and ATR inhibition for the treatment of p53-deficient patients.

IFN-γ induces acute graft-versus-host disease by promoting HMGB1-mediated nuclear-to-cytoplasm translocation and autophagic degradation of p53.

Acute graft-versus-host disease (aGVHD) poses a significant impediment to achieving a more favourable therapeutic outcome in allogeneic hematopoietic stem cell transplantation (allo-HSCT). Our prior investigations disclosed a correlation between p53 down-regulation in CD4+ T cells and the occurrence of aGVHD. Notably, the insufficiency of the CCCTC-binding factor (CTCF) emerged as a pivotal factor in repressing p53 expression. However, the existence of additional mechanisms contributing to the reduction in p53 expression remains unclear. Interferon (IFN)-γ, a pivotal proinflammatory cytokine, assumes a crucial role in regulating alloreactive T-cell responses and plays a complex part in aGVHD development. IFN-γ has the capacity to induce autophagy, a vital catabolic process facilitating protein degradation, in various cell types. Presently, whether IFN-γ participates in the development of aGVHD by instigating the autophagic degradation of p53 in CD4+ T cells remains an unresolved question. In the present study, we demonstrated that heightened levels of IFN-γ in the plasma during aGVHD promoted the activation, proliferation, and autophagic activity of CD4+ T cells. Furthermore, IFN-γ induced the nuclear-to-cytoplasm translocation and autophagy-dependent degradation of p53 in CD4+ T cells. The translocation and autophagic degradation of p53 were contingent upon HMGB1, which underwent up-regulation and translocation from the nucleus to the cytoplasm following IFN-γ stimulation. In conclusion, our data unveil a novel mechanism underlying p53 deficiency in CD4+ T cells among aGVHD patients. This deficiency is induced by IFN-γ and relies on autophagy, establishing a link between IFN-γ, HMGB1-mediated translocation, and the autophagic degradation of p53.

The totipotent 2C-like state safeguards genomic stability of mouse embryonic stem cells.

Mouse embryonic stem cells (mESCs) sporadically transition to a transient totipotent state that resembles blastomeres of the two-cell (2C) embryo stage, which has been proposed to contribute to exceptional genomic stability, one of the key features of mESCs. However, the biological significance of the rare population of 2C-like cells (2CLCs) in ESC cultures remains to be tested. Here we generated an inducible reporter cell system for specific elimination of 2CLCs from the ESC cultures to disrupt the equilibrium between ESCs and 2CLCs. We show that removing 2CLCs from the ESC cultures leads to dramatic accumulation of DNA damage, genomic mutations, and rearrangements, indicating impaired genomic instability. Furthermore, 2CLCs removal results in increased apoptosis and reduced proliferation of mESCs in both serum/LIF and 2i/LIF culture conditions. Unexpectedly, p53 deficiency results in defective response to DNA damage, leading to early accumulation of DNA damage, micronuclei, indicative of genomic instability, cell apoptosis, and reduced self-renewal capacity of ESCs when devoid of 2CLCs in cultures. Together, our data reveal that transition to the privileged 2C-like state is a major component of the intrinsic mechanisms that maintain the exceptional genomic stability of mESCs for long-term self-renewal.

P53 deficiency mediates cisplatin resistance by upregulating RRM2 and crotonylation of RRM2K283 through the downregulation of SIRT7.

p53 deficiency plays a crucial role in chemotherapy resistance through various biological events, including posttranslational modifications (PTMs). Recently, lysine crotonylation (Kcr) has been shown to play a vital role in cancer progression. However, the global p53-regulated crotonylome and the function of these altered Kcr proteins after p53 deficiency remain unclear. In this study, we used a SILAC-based quantitative crotonylome to identify 3,520 Kcr in 1924 crotonylated proteins in response to p53 knockout. We found that increased crotonylation of RRM2 at K283 (RRM2K283Cr) in the presence of p53 deficiency promoted HCT116 cell resistance to cisplatin. We discovered that SIRT7 could be the decrotonylase of RRM2 and was downregulated after p53 knockout, resulting in increased RRM2K283Cr. Mechanistically, p53 deficiency inhibited cell apoptosis by upregulating RRM2 protein expression and RRM2K283Cr-mediated cleaved-PARP1 and cleaved-caspase3 expression, and SIRT7 was downregulated to upregulate crotonylation of RRM2 upon p53 deficiency. In conclusion, our results indicated that p53 deficiency plays a malignant role in colon cancer resistance to cisplatin therapy by regulating RRM2 protein and RRM2K283Cr expression. Our findings provide a novel therapeutic target against p53-deficient cancer.

Abstract NG02: Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers

Abstract NG02: Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers

Abstract 6947: Exploring the single-cell dynamics of FOXM1 under cell cycle perturbations

Abstract The cell cycle plays a critical role in maintaining normal cellular functions and preventing abnormal cell growth. The transcription factor FOXM1 has emerged as a crucial regulator of cell cycle progression and has been implicated in various physiological and pathological processes. Notably, FOXM1 overexpression has been observed in numerous cancer types, including liver, prostate, breast, lung, and colon cancer. Despite its significance, our understanding of how FOXM1 dynamic behaves at the single-cell level under different cell cycle perturbations and its link to cell behavior heterogeneity remains limited. Here, we investigated the dynamic behavior of FOXM1 in individual cells exposed to various cell cycle perturbations, focusing on understanding heterogeneity in cell behavior and improving disease treatment. At the single-cell level, our investigation reveals six distinct dynamic features of FOXM1 in response to various therapeutic interventions, each of which corresponds to a unique cell phenotype. Our result further demonstrates that for cells exhibiting a high FOXM1 pattern, targeting aurora kinase and PLK1 may hold promise as a therapeutic approach for cancer suggesting that FOXM1 is a strong biomarker, and its pattern can be predictive of drug response, especially in cases with G1 checkpoint defects like Rb1 loss or p53 deficiency. Citation Format: Tooba Jawwad, Maliwan Kamkaew, Kriengkrai Phongkitkarun, Porncheera Chusorn, Somponnat Sampattavanich. Exploring the single-cell dynamics of FOXM1 under cell cycle perturbations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6947.

Haploinsufficiency of Adenomatous Polyposis Coli Coupled with Kirsten Rat Sarcoma Viral Oncogene Homologue Activation and P53 Loss Provokes High-Grade Glioblastoma Formation in Mice.

Glioblastoma multiforme (GBM) is the most common and deadly type of brain tumor originating from glial cells. Despite decades of clinical trials and research, there has been limited success in improving survival rates. However, molecular pathology studies have provided a detailed understanding of the genetic alterations associated with the formation and progression of glioblastoma-such as Kirsten rat sarcoma viral oncogene homolog (KRAS) signaling activation (5%), P53 mutations (25%), and adenomatous polyposis coli (APC) alterations (2%)-laying the groundwork for further investigation into the biological and biochemical basis of this malignancy. These analyses have been crucial in revealing the sequential appearance of specific genetic lesions at distinct histopathological stages during the development of GBM. To further explore the pathogenesis and progression of glioblastoma, here, we developed the glial-fibrillary-acidic-protein (GFAP)-Cre-driven mouse model and demonstrated that activated KRAS and p53 deficiencies play distinct and cooperative roles in initiating glioma tumorigenesis. Additionally, the combination of APC haploinsufficiency with mutant Kras activation and p53 deletion resulted in the rapid progression of GBM, characterized by perivascular inflammation, large necrotic areas, and multinucleated giant cells. Consequently, our GBM models have proven to be invaluable resources for identifying early disease biomarkers in glioblastoma, as they closely mimic the human disease. The insights gained from these models may pave the way for potential advancements in the diagnosis and treatment of this challenging brain tumor.

Abstract C056: The initiation and progression of pancreatic ductal adenocarcinoma through a p53 lens

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a 5-year survival rate of only 11%. Slated to be the 2nd most deadly cancer by 2026, pancreatic cancer survival rates have not improved substantially over the years. With a lack of early detection methods and a highly immunosuppressive tumor microenvironment (TME), PDAC tumors are highly resistant to treatment. Oncogenic mutations in KRAS are present in >90% of cases and mutations in the tumor suppressor gene TP53 are present in ~72% of cases. However, why p53 mutations are so prevalent and how they enhance PDAC development is not well understood. To elucidate how exactly p53 loss promotes PDAC, we are using genetically engineered mouse models combined with single-cell and spatial transcriptomic technologies. In this mouse model, PDAC is driven from adult acinar cells with a tamoxifen-inducible Ptf1aCreER allele by oncogenic Kras (KrasG12D) along with either p53 wild type (p53wt) or p53 deficiency (p53flox). In this model, acinar cells can give rise to PDAC likely through a transdifferentiation process called acinar-to-ductal metaplasia (ADM), which leads to pancreatic intraepithelial neoplasias (PanINs) and then PDAC. The initial cell fate transition, ADM, can be activated by growth factors, inflammation, and injury. We have conducted single-cell and spatial transcriptomics to compare early stages of tumor development in the presence and absence of p53, and we will present our latest analyses. Providing insight into the early events of PDAC will lead to new approaches for improving clinical intervention in human PDAC. Citation Format: Kathryn J. Hanson, Allegra C. Minor, Brittany M. Flowers, Alyssa M. Kaiser, Nicholas Hughes, Hannes Vogel, Le Cong, Laura D. Attardi. The initiation and progression of pancreatic ductal adenocarcinoma through a p53 lens [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C056.

P53 exerts anticancer effects by regulating enhancer formation and activity.

The abnormality of the p53 tumor suppressor is crucial in lung cancer development, because p53 regulates target gene promoters to combat cancer. Recent studies have shown extensive p53 binding to enhancer elements. However, whether p53 exerts a tumor suppressor role by shaping the enhancer landscape remains poorly understood. In the current study, we employed several functional genomics approaches to assess the enhancer activity at p53 binding sites throughout the genome based on our established TP53 knockout (KO) human bronchial epithelial cells (BEAS-2B). A total of 943 active regular enhancers and 370 super-enhancers (SEs) disappeared upon the deletion of p53, indicating that p53 modulates the activity of hundreds of enhancer elements. We found that one p53-dependent SE, located on chromosome 9 and designated as KLF4-SE, regulated the expression of the Krüppel-like factor 4 ( KLF4) gene. Furthermore, the deletion of p53 significantly decreased the KLF4-SE enhancer activity and the KLF4 expression, but increased colony formation ability in the nitrosamines 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced cell transformation model. Subsequently, in TP53 KO cells, the overexpression of KLF4 partially reversed the increased clonogenic capacity caused by p53 deficiency. Consistently, KLF4 expression also decreased in lung cancer tissues and cell lines. It appeared that overexpression of KLF4 significantly suppressed the proliferation and migration of lung cancer cells. Collectively, our results suggest that the regulation of enhancer formation and activity by p53 is an integral component of the p53 tumor suppressor function. Therefore, our findings offer some novel insights into the regulation mechanism of p53 in lung oncogenesis and introduce a new strategy for screening therapeutic targets.

Repeated radon exposure induced epithelial-mesenchymal transition-like transformation via disruption of p53-dependent mitochondrial function.

As a human carcinogen, radon and its progeny are the second most important risk factor for lung cancer after smoking. The tumor suppressor gene, p53, is reported to play an important role in the maintenance of mitochondrial function. In this work, we investigated the association between p53 and p53-responsive signaling pathways and radon-induced carcinogenesis. After repeated radon exposure, the malignant characteristics, cell cycle arrest, cell apoptotic rate, adenosine triphosphate (ATP) content, reactive oxygen species (ROS) level, mitochondrial DNA (mtDNA) copy number as well as indicative biomarkers involved in mitochondrial energy metabolism were evaluated in BEAS-2B cells or BALB-c mouse lung tissue. Radon exposure induced epithelial-mesenchymal transition (EMT)-like transformation in BEAS-2B cells, as indicated by increased cell proliferation and migration. Additional mitochondrial alterations, including decreased ATP content, increased ROS levels, mtDNA copy numbers, cell apoptosis, and G2/M cell cycle arrest were observed. Radon exposure caused an energy generation shift from aerobic respiration to glycolysis as reflected by increased expression of TIGAR and p53R2 proteins and decreased expression of SCO2 protein in BEAS-2B cells, and increased expression of p53, SCO2 and TIGAR proteins in mouse lung tissue, respectively. The effects of p53 deficiency on the prevention of mitochondrial dysfunction suggested a protective role of p53 in radon-induced malignant-like features in BEAS-2B cells. Repeated radon exposure induced EMT-like transformation in BEAS-2B cells via disruption of mitochondrial function. Activation of p53 and p53-responsive signaling pathways in BEAS-2B cells and BALB-c mice may confer a protective mechanism for radon-induced lung injury.

Functional and transcriptional sequelae of cardiomyocyte telomere shortening and their pathologic relevance in dilated cardiomyopathy and ischemic heart failure

Abstract Background Heart failure (HF) is associated with cardiomyocyte (CM) telomere shortening. Sequelae have been described with the telomere-p53-mitochondrion model. P53 deficiency only partially ameliorates mitochondrial defects in telomerase deficient mice hinting at additional pathways. Purpose We assessed the cardiac phenotype and transcriptome of early/advanced telomere shortening and evaluated the pathological relevance by correlation to published HF datasets. Methods Male (6-8w) telomerase RNA-component deficient mice, early (generation 2, mTRG2) and advanced (mTRG5) were phenotyped vs age-matched C57BL/6 mice (B6). Adult CMs from mTRG2/G5 vs B6 mice were characterized (live imaging, quantitative fluorescence in situ hybridization (Q-FISH) and transcriptome analysis (microarray Mouse MTA 1.0)). Gene expression analysis was performed (Ingenuity Pathway Analysis, Qiagen) and correlated to comparable datasets from > 120 000 curated datasets. Results CMs exhibited significant telomere shortening in mTR(-/-) mice increasing with every generation (mTRG2/G5 p<0.0001 vs B6). In parallel, echocardiography revealed increasing defects of left ventricular (LV) systolic function (LVEF, mTRG5 <0.01 vs B6), LV global longitudinal strain mTRG2/G5 p<0.001 vs B6) and LV contractile reserve (mTRG2/G5 p<0.05 vs B6). Live imaging showed increasing impairment of mitochondrial polarization with all energy substrates (mTRG2/G5 p<0.0001 vs B6). Transcriptome analysis revealed 357/2906 differentially expressed genes (DEGs) in mTRG2/G5-CMs vs B6-CMs (fold change <-1.5 or >1.5 and p-value <0.05). Upstream Regulator (UR) analysis revealed significant overrepresentation of numerous regulators including angiotensinogen (z-score 7.5, p-value 1.1E-32), transforming growth factor (TGF)-b1 (z-score 7.5, p-value 2.2E-60), interleukin (IL)-1 (z-score 5.6, p-value 1.1E-26), IL-6 (expression fold change 11.3, z-score 6.9 p-value 2.65E-32), IL-17A (z-score 4.8, p-value 4.4E-27) and NFkB (z-score 6.0 p-value 4.9E-19). Activation of TP53 (z-score 5.8, p-value 5.3E-31) and inhibition of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PPARGC1A) (z-score -4.4, p-value 6.3E-19) confirmed the relevance of the p53-mitochondrion-axis. Analysis matching revealed a significant overlap of gene expression patterns in the same direction (positive z-score) between mTR-/--mice and all comparable datasets of dilated (DCM) and ischemic cardiomyopathy (ICM) (human/mouse), the overlap increasing with increasing telomere shortening (e.g. DCM vs mTRG2: z-score 17.7, UR p-value 1.5E-16; DCM vs mTRG5: z-score 56.8, UR-value 1.5E-62). Matching to datasets after cardiac regeneration (e.g. after left ventricular assist device implantation) revealed a reversal of gene expression patterns (negative z-score). Conclusions Our results reveal the large extent of transcriptional alterations upon cardiac telomere shortening and support their central role in the pathogenesis of HF.

A genetic mosaic mouse model illuminates the pre-malignant progression of basal-like breast cancer.

Basal-like breast cancer (BLBC) is highly aggressive, and often characterized by BRCA1 and p53 deficiency. Although conventional mouse models enabled the investigation of BLBC at malignant stages, its initiation and pre-malignant progression remain understudied. Here, we leveraged a mouse genetic system known as mosaic analysis with double markers (MADM) to study BLBC initiation by generating rare GFP+Brca1, p53-deficient mammary cells alongside RFP+ wild-type sibling cells. After confirming the close resemblance of mammary tumors arising in this model to human BLBC at both transcriptomic and genomic levels, we focused our studies on the pre-malignant progression of BLBC. Initiated GFP+ mutant cells showed a stepwise pre-malignant progression trajectory from focal expansion to hyper-alveolarization and then to micro-invasion. Furthermore, despite morphological similarities to alveoli, hyper-alveolarized structures actually originate from ductal cells based on twin-spot analysis of GFP-RFP sibling cells. Finally, luminal-to-basal transition occurred exclusively in cells that have progressed to micro-invasive lesions. Our MADM model provides excellent spatiotemporal resolution to illuminate the pre-malignant progression of BLBC, and should enable future studies on early detection and prevention for this cancer.

Arsenite-induced Radiosensitization of Glioma Cells Is Dependent on p53 Deficiency.

Arsenite is a radiosensitizer of glioma cells both in vitro and in vivo; however, the underlying mechanism of action is unclear. Radiosensitizers specific for p53-deficient tumors are a promising adjunct to radiotherapy because, unlike normal cells, many tumor cells lack p53. Previously, we demonstrated that arsenite sensitizes the p53-deficient glioma cell line U87MG-E6 to X-rays. Using flowcytometry, we expand these findings to p53-proficient U87MG cells exposed to heavy ion beams, including carbon and iron ions. Arsenite sensitized U87MG-E6, but not U87MG, cells to heavy ion beams and X-rays. Cell cycle analysis indicated that sensitization of U87MG-E6 was related to an increase in the percentage of cells in the late S/G2/M phases after combined treatment with arsenite, especially when carbon ion beams were used. Induction of γH2AX was significant in U87MG-E6, but not in U87MG, cells after irradiation with carbon ion beams plus arsenite. Arsenite sensitizes cells by increasing the percentage of cells in the late S/G2/M phases after irradiation, possibly via inhibition of DNA repair in the context of p53 deficiency. The findings provide information that may be useful for the development of advanced radiotherapy protocols.

Role of p53/circRNA0085439/Ku70 axis in DNA damage response in lung cells exposed to ZnO nanoparticles: Involvement of epigenetic regulation

BackgroundNano-Zinc oxide (Nano-ZnO) has been increasingly applied in agriculture, industry and biomedicine. However, the genotoxic effects of Nano-ZnO and the underlying mechanisms remain incompletely clear.MethodsHuman bronchial epithelial cell line (HBE) was used to observe the effects of Nano-ZnO on DNA damage repair-related proteins and epithelial mesenchymal transition (EMT) by Western blotting. Then, CRISPR/cas9-based technique was used to create p53 knockout (p53-KO) cell line. RNA-seq analysis was performed to uncover the circular RNA (circRNA) profile after Nano-ZnO treatment in p53-KO cells compared with p53 wild-type (p53-wt) cells. LC–MS/MS was used to discover the potential binding proteins of circRNA_0085439 in the p53 deficiency background after Nano-ZnO treatment. Nano-ZnO-induced DNA damage and EMT were also investigated in vivo by instillation of Nano-ZnO (50 µg/mouse).ResultsNano-ZnO exposure caused DNA damage and EMT at both in vitro and in vivo background, which was reflected by increased DNA damage associated proteins such as ATM and ATR and γ H2AX. p53 expression increased at the early stage post Nano-ZnO treatment decreased later. RNA-seq assay showed a highest increase of circRNA_0085439 expression in p53-KO cells compared with the p53-wt cells after Nano-ZnO exposure. Silencing of p53 expression promoted its translocation of circRNA_0085439 from cytoplasm to nucleus leading to the formation of circRNA_0085439/Ku70 complex resulting in the decreased expression of Ku70 protein. In addition, increased EMT markers, N-cadherin and Vimentin, was observed in lung epithelial cells and in mouse lungs at day 7 after Nano-ZnO exposure.ConclusionsThis study unraveled the epigenetic mechanisms underlying Nano-ZnO-induced DNA damage and EMT. The effect of Nano-ZnO-induced DNA damage through p53/circRNA_0085439/Ku70 pathway likely contribute to Nano-ZnO-induced cell cytotoxicity and apoptosis. Our findings will provide information to further elucidate the molecular mechanisms of Nano-ZnO-induced cytotoxicity and genotoxicity.

P53 Deficiency-Dependent Oncogenicity of Runx3.

The RUNX transcription factors are frequently dysregulated in human cancers, suggesting their potential as attractive targets for drug treatment. However, all three transcription factors have been described as both tumor suppressors and oncogenes, indicating the need to determine their molecular mechanisms of action. Although RUNX3 has long been considered a tumor suppressor in human cancers, several recent studies have shown that RUNX3 is upregulated during the development or progression of various malignant tumors, suggesting it may act as a "conditional" oncogene. Resolving this paradox and understanding how a single gene can exhibit both oncogenic and tumor-suppressive properties is essential for successful drug targeting of RUNX. This review describes the evidence for the activities of RUNX3 in human cancer and proposes an explanation for the duality of RUNX3 involving the status of p53. In this model, p53 deficiency causes RUNX3 to become oncogenic, leading to aberrant upregulation of MYC.

Features of the immune and metabolomic profile in overweight children associated with polymorphic variants of the IL-1B and TP53 genes

Background. The growing number of overweight children is an important problem of modern healthcare. Excess body weight causes the risk of severe metabolic disorders developing. Analysis of changes in the immune, metabolic, and genetic profile of overweight children as markers of the early development of metabolic disorders is an effective tool for the prevention, diagnosis, and treatment of these pathologies.
 Aim. To identify the features of the immune and metabolomic profile in overweight children aged 711 years associated with polymorphic variants of the IL-1B (rs1143634) and TP53 (rs1042522) genes.
 Material and methods. 132 children aged 711 years were examined: 54 children with overweight and 78 children with normal body weight. The lymphocytes subpopulation composition was analyzed by flow cytometry; the content of leptin, cortisol, superoxide dismutase, lipid hydroperoxide and interleukin-1 using enzyme immunoassay, immunoglobulin G using the method of radial immunodiffusion method by Mancini. Polymorphic variants of the IL-1B (rs1143634) and TP53 (rs1042522) genes were identified by real-time polymerase chain reaction. Statistical data processing was implemented in the Statistica 10, SNPStat and Gen-Expert programs.
 Results. The immune profile of the examined children was characterized by an increased content of CD3+CD8+-, CD19+-lymphocytes and immunoglobulin G against the background of CD3+CD4+-lymphocytes, p53 deficiency and a decrease in the number of CD4+/CD8+. The metabolomic profile of children was characterized by an excess content of leptin, cortisol, lipid hydroperoxides, and superoxide dismutase. The identified changes in immune and metabolomic regulation in overweight children were significantly associated with the G allele and GG genotype of the IL-1B gene (rs1143634), as well as with the C allele and CC genotype of the TP53 gene (rs1042522), which cause the initiation of inflammation and inhibition of apoptosis.
 Conclusion. The established features of immune and metabolomic regulation associated with the G allele and GG genotype of the IL-1B gene (rs1143634) and with the C allele and CC genotype of TP53 gene (rs1042522) represent a complex of markers of overweight formation in children aged 711 years.

P53 Deficiency Accelerate Esophageal Epithelium Intestinal Metaplasia Malignancy

Barrett's esophagus (BE) is a precancerous lesion of esophageal adenocarcinoma (EAC). It is a pathological change in which the squamous epithelium distal esophagus is replaced by columnar epithelium. Loss of P53 is involved in the development of BE and is taken as a risk factor for the progression. We established a HET1A cell line with P53 stably knockdown by adenovirus vector infection, followed by 30 days of successive acidic bile salt treatment. MTT, transwell assay, and wound closure assay were applied to assess cell proliferation and migration ability. The expression of key factors was analyzed by RT-qPCR, western blotting and immunohistochemical staining. Our data show that the protein expression level of P53 reduced after exposure to acidic bile salt treatment, and the P53 deficiency favors the survival of esophageal epithelial cells to accommodate the stimulation of acidic bile salts. Furthermore, exposure to acidic bile salt decreases cell adhesions by repressing the JAK/STAT signaling pathway and activating VEGFR/AKT in P53-deficient esophageal cells. In EAC clinical samples, P53 protein expression is positively correlated with that of ICAM1 and STAT3 and negatively correlated with VEGFR protein expression levels. These findings elucidate the role of P53 in the formation of BE, explain the mechanism of P53 deficiency as a higher risk of progression for BE formation, and provide potential therapeutic targets for EAC.