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1.
Int J Oncol ; 54(6): 1907-1920, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31081062

RESUMO

The p53 protein is a tumour suppressor and transcription factor that regulates the expression of target genes involved in numerous stress responses systems. In this study, we designed a screening strategy using DNA damage­induced mouse and human transcriptome data to identify novel downstream targets of p53. Our method selected genes with an induced expression in multiple organs of X­ray­irradiated p53 wild­type mice. The expression of inka box actin regulator 2 gene, known as Inka2, was upregulated in 12 organs when p53 expression was induced. Similarly, INKA2 was induced in a p53­dependent manner at both the mRNA and protein level in human cells treated with adriamycin. Reporter assays confirmed that p53 directly regulated INKA2 through an intronic binding site. The overexpression of INKA2 produced a slight decrease in cancer cell growth in the colony formation assay. Moreover, the analysis of The Cancer Genome Atlas (TCGA) data revealed a decreased INKA2 expression in tumour samples carrying p53 mutations compared with p53 wild­type samples. In addition, significantly higher levels of DNA methylation were observed in the INKA2 promoter in tumour samples, concordant with the reduced INKA2 expression in tumour tissues. These results demonstrate the potential of INKA2 as a cancer cell growth inhibitor. Furthermore, INKA2 protein interacts with the serine/threonine­protein kinase, p21 (RAC1) activated kinase (PAK)4, which phosphorylates ß­catenin to prevent ubiquitin­proteasomal degradation. As ß­catenin was downregulated in a stable INKA2­expressing cell line, the findings of this study suggest that INKA2 is a novel, direct downstream target of p53 that potentially decreases cell growth by inhibiting the PAK4­ß­catenin pathway.


Assuntos
Perfilação da Expressão Gênica/métodos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias/genética , Proteína Supressora de Tumor p53/metabolismo , Quinases Ativadas por p21/metabolismo , Animais , Linhagem Celular Tumoral , Metilação de DNA , Doxorrubicina/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Células HCT116 , Humanos , Camundongos , Mutação , Neoplasias/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Análise de Sequência de RNA/métodos , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , beta Catenina/metabolismo
2.
J Am Soc Nephrol ; 30(5): 855-864, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30975718

RESUMO

BACKGROUND: A family history of urolithiasis is associated with a more than doubling of urolithiasis risk, and a twin study estimating 56% heritability of the condition suggests a pivotal role for host genetic factors. However, previous genome-wide association studies (GWAS) have identified only six risk-related loci. METHODS: To identify novel urolithiasis-related loci in the Japanese population, we performed a large-scale GWAS of 11,130 cases and 187,639 controls, followed by a replication analysis of 2289 cases and 3817 controls. Diagnosis of urolithiasis was confirmed either by a clinician or using medical records or self-report. We also assessed the association of urolithiasis loci with 16 quantitative traits, including metabolic, kidney-related, and electrolyte traits (such as body mass index, lipid storage, eGFR, serum uric acid, and serum calcium), using up to 160,000 samples from BioBank Japan. RESULTS: The analysis identified 14 significant loci, including nine novel loci. Ten regions showed a significant association with at least one quantitative trait, including metabolic, kidney-related, and electrolyte traits, suggesting a common genetic basis for urolithiasis and these quantitative traits. Four novel loci were related to metabolic traits, obesity, hypertriglyceridemia, or hyperuricemia. The remaining ten loci were associated with kidney- or electrolyte-related traits; these may affect crystallization. Weighted genetic risk score analysis indicated that the highest risk group (top 20%) showed an odds ratio of 1.71 (95% confidence interval, 1.42 to 2.06) - 2.13 (95% confidence interval, 2.00 to 2.27) compared with the reference group (bottom 20%). CONCLUSIONS: Our findings provide evidence that host genetic factors related to regulation of metabolic and crystallization pathways contribute to the development of urolithiasis.

3.
J Invest Dermatol ; 139(7): 1459-1469, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30684556

RESUMO

Identification of the specific genetic variants responsible for the increased susceptibility to familial or sporadic cancers is important. Using a forward genetics approach to map such loci in a mouse skin cancer model, we previously identified a strong genetic locus, Stmm3, conferring resistance to chemically induced skin papillomas on chromosome 4. Here, we report the cyclin-dependent kinase inhibitor gene Cdkn2a/p19Arf as a major responsible gene for the Stmm3 locus. We provide evidence that the function of Stmm3 is dependent on p53 and that p19ArfMSM confers stronger resistance to papillomas than p16Ink4aMSMin vivo. In addition, we found that genetic polymorphism in p19Arf between a resistant strain, MSM/Ms (Val), and a susceptible strain, FVB/N (Leu), alters the susceptibility to papilloma development, malignant conversion, and the epithelial-mesenchymal transition. Moreover, we demonstrated that the p19ArfMSM allele more efficiently activates the p53 pathway than the p19ArfFVB allele in vitro and in vivo. Furthermore, we found polymorphisms in CDKN2A in the vicinity of a polymorphism in mouse Cdkn2a associated with the risk of human cancers in the Japanese population. Genetic polymorphisms in Cdkn2a and CDKN2A may affect the cancer risk in both mice and humans.

4.
Gastroenterology ; 156(5): 1455-1466, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30529582

RESUMO

BACKGROUND & AIMS: Genome-wide association studies (GWASs) have associated approximately 50 loci with risk of colorectal cancer (CRC)-nearly one third of these loci were initially associated with CRC in studies conducted in East Asian populations. We conducted a GWAS of East Asians to identify CRC risk loci and evaluate the generalizability of findings from GWASs of European populations to Asian populations. METHODS: We analyzed genetic data from 22,775 patients with CRC (cases) and 47,731 individuals without cancer (controls) from 14 studies in the Asia Colorectal Cancer Consortium. First, we performed a meta-analysis of 7 GWASs (10,625 cases and 34,595 controls) and identified 46,554 promising risk variants for replication by adding them to the Multi-Ethnic Global Array (MEGA) for genotype analysis in 6445 cases and 7175 controls. These data were analyzed, along with data from an additional 5705 cases and 5961 controls genotyped using the OncoArray. We also obtained data from 57,976 cases and 67,242 controls of European descent. Variants at identified risk loci were functionally annotated and evaluated in correlation with gene expression levels. RESULTS: A meta-analyses of all samples from people of Asian descent identified 13 loci and 1 new variant at a known locus (10q24.2) associated with risk of CRC at the genome-wide significance level of P < 5 × 10-8. We did not perform experiments to replicate these associations in additional individuals of Asian ancestry. However, the lead risk variant in 6 of these loci was also significantly associated with risk of CRC in European descendants. A strong association (44%-75% increase in risk per allele) was found for 2 low-frequency variants: rs201395236 at 1q44 (minor allele frequency, 1.34%) and rs77969132 at 12p11.21 (minor allele frequency, 1.53%). For 8 of the 13 associated loci, the variants with the highest levels of significant association were located inside or near the protein-coding genes L1TD1, EFCAB2, PPP1R21, SLCO2A1, HLA-G, NOTCH4, DENND5B, and GNAS. For other intergenic loci, we provided evidence for the possible involvement of the genes ALDH7A1, PRICKLE1, KLF5, WWOX, and GLP2R. We replicated findings for 41 of 52 previously reported risk loci. CONCLUSIONS: We showed that most of the risk loci previously associated with CRC risk in individuals of European descent were also associated with CRC risk in East Asians. Furthermore, we identified 13 loci significantly associated with risk for CRC in Asians. Many of these loci contained genes that regulate the immune response, Wnt signaling to ß-catenin, prostaglandin E2 catabolism, and cell pluripotency and proliferation. Further analyses of these genes and their variants is warranted, particularly for the 8 loci for which the lead CRC risk variants were not replicated in persons of European descent.


Assuntos
Grupo com Ancestrais do Continente Asiático/genética , Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Loci Gênicos , Polimorfismo de Nucleotídeo Único , Ásia/epidemiologia , Estudos de Casos e Controles , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/etnologia , Neoplasias Colorretais/imunologia , Frequência do Gene , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Fenótipo , Medição de Risco , Fatores de Risco
5.
PLoS One ; 13(12): e0209096, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30557369

RESUMO

Genome-wide association studies (GWAS) have identified greater than 30 variants associated with ovarian cancer, but most of these variants were investigated in European populations. Here, we integrated GWAS and subsequent functional analyses to identify the genetic variants with potential regulatory effects. We conducted GWAS for ovarian cancer using 681 Japanese cases and 17,492 controls and found that rs137672 on 22q13.1 exhibited a strong association with a P-value of 1.05 × 10(-7) and an odds ratio of 0.573 with a 95% confidence interval of 0.466-0.703. In addition, three previously reported SNPs, i.e., rs10088218, rs9870207 and rs1400482, were validated in the Japanese population (P < 0.05) with the same risk allele as noted in previous studies. Functional studies including regulatory feature analysis and electrophoretic mobility shift assay (EMSA) revealed two regulatory SNPs in 22q13.1, rs2072872 and rs6509, that affect the binding affinity to some nuclear proteins in ovarian cancer cells. The plausible regulatory proteins whose motifs could be affected by the allele changes of these two SNPs were also proposed. Moreover, the protective G allele of rs6509 was associated with a decreased SYNGR1 expression level in normal ovarian tissues. Our findings elucidated the regulatory variants in 22q13.1 that are associated with ovarian cancer risk.


Assuntos
Cromossomos Humanos Par 22/genética , Variação Genética , Estudo de Associação Genômica Ampla , Neoplasias Ovarianas/genética , Idoso , Alelos , Estudos de Casos e Controles , Feminino , Técnicas de Genotipagem , Humanos , Japão , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único
6.
Cancer Sci ; 109(12): 4015-4024, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30281874

RESUMO

Gastric cancer is the third leading cause of cancer mortality in Japan and worldwide. Although previous studies identify various genetic variations associated with gastric cancer, host genetic factors are largely unidentified. To identify novel gastric cancer loci in the Japanese population, herein, we carried out a large-scale genome-wide association study using 6171 cases and 27 178 controls followed by three replication analyses. Analysis using a total of 11 507 cases and 38 904 controls identified two novel loci on 12q24.11-12 (rs6490061, P = 3.20 × 10-8 with an odds ratio [OR] of 0.905) and 20q11.21 (rs2376549, P = 8.11 × 10-10 with an OR of 1.109). rs6490061 is located at intron 19 of the CUX2 gene, and its expression was suppressed by Helicobacter pylori infection. rs2376549 is included within the gene cluster of DEFB families that encode antibacterial peptides. We also found a significant association of rs7849280 in the ABO gene locus on 9q34.2 (P = 2.64 × 10-13 with an OR of 1.148). CUX2 and ABO expression in gastric mucosal tissues was significantly associated with rs6490061 and rs7849280 (P = 0.0153 and 8.00 × 10-11 ), respectively. Our findings show the crucial roles of genetic variations in the pathogenesis of gastric cancer.

7.
J Urol ; 2018 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-30063927

RESUMO

PURPOSE: We evaluated the association of hypospadias with 17 susceptibility loci previously identified by a European genome-wide association study (GWAS) in Japanese patients. We also examined the expression of candidate genes in male mouse embryos to discuss the possible underlying mechanisms of this disease. MATERIALS AND METHODS: We enrolled 169 Japanese patients (mean age at surgery: 3.7 years) who underwent repair of hypospadias. Genotyping of 17 SNPs was performed by using a multiplex-polymerase chain reaction (PCR) invader assay. We also performed in situ hybridization to determine whether candidate genes were expressed in the male genital tubercle during embryonic development of the external genitalia in mice. RESULTS: SNP rs3816183 of HAAO was significantly associated with susceptibility to hypospadias in general (p=0.0019) and both anterior/middle hypospadias (p=0.0283) and posterior hypospadias (p=0.0226), while SNP rs6499755 of IRX6 showed an association with susceptibility to anterior/middle hypospadias (p=0.0472). In mouse embryos, there was no significant upregulation of Haao expression in the developing male external genitalia. Irx3 and Irx5, which are linked to Irx6 within the IrxB cluster, were expressed in the mesenchyme remote from the urethral plate epithelium during the critical embryonic period for masculinization. Irx6 was expressed in the ectodermal epithelium, showing prominent dorsal ectodermal expression without expression in ventral ectoderm adjacent to the urethral plate during the same period. CONCLUSIONS: Genetic variations of HAAO and IRX6 influence susceptibility to hypospadias in Japanese population, although further study will be needed to clarify the mechanism by which contributes to the pathogenesis of hypospadias.

8.
Oncotarget ; 9(3): 3936-3945, 2018 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-29423095

RESUMO

SNP rs2294008 in Prostate Stem Cell Antigen (PSCA) and decreased PSCA expression are associated with gastric cancer. The objective of this study is to investigate the role of rs2294008 and PSCA expression in the gastritis-gastric cancer carcinogenic pathway. We conducted a case-control association study of H. pylori-infected gastritis and gastric cancer. rs2294008 was associated with the progression to chronic active gastritis (P = 9.4 × 10-5; odds ratio = 3.88, TT + TC vs CC genotype), but not with H. pylori infection per se nor with the progression from active gastritis to gastric cancer. We also assessed the association of rs2294008 with PSCA mRNA expression in the gastric mucosa at various disease stages and found that rs2294008 was associated with PSCA expression (P = 1.3 × 10-12). H. pylori infection (P = 5.1 × 10-8) and eradication therapy (P < 1 × 10-11) resulted in the reduced and increased PSCA expression, respectively, indicating negative regulation of PSCA expression by H. pylori infection. PSCA expression was decreased in severe gastritis compared with mild gastritis only among T allele carriers. Our findings revealed the regulation of PSCA expression by host genetic variation and bacterial infection might contribute to gastritis progression after H. pylori infection.

9.
Carcinogenesis ; 39(5): 652-660, 2018 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-29471430

RESUMO

Colorectal cancer (CRC) is the fourth leading cause of cancer mortality worldwide. Genome-wide association studies (GWAS) identified more than 50 CRC loci. However, most of the previous studies were conducted in European population, and host genetic factors among Japanese population are largely remained to be identified. To identify novel loci in the Japanese population, here, we performed a large-scale GWAS using 6692 cases and 27 178 controls followed by a replication analysis using more than 11 000 case-control samples. We found the significant association of 10 loci (P < 5 × 10-8), including 2 novel loci on 16q24.1 (IRF8-FOXF1, rs847208, P = 3.15 × 10-9 and odds ratio = 1.107 with 95% confidence interval (CI) of 1.071-1.145) and 20q13.12 (TOX2, rs6065668, P = 4.47 × 10-11 and odds ratio = 0.897 with 95% CI of 0.868-0.926). Moreover, 35 previously reported single nucleotide polymorphisms (SNPs) in 24 regions were validated in the Japanese population (P < 0.05) with the same risk allele as in the previous studies. SNP rs6065668 was significantly associated with TOX2 expression in the sigmoid colon. In addition, nucleotide substitutions in the regulatory region of TOX2 were predicted to alter the binding of several transcription factors, including KLF5. Our findings elucidate the important role of genetic variations in the development of CRC in the Japanese population.

10.
Cell Rep ; 22(6): 1473-1483, 2018 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-29425503

RESUMO

Recent proteome analyses have provided a comprehensive overview of various posttranslational modifications (PTMs); however, PTMs involving protein citrullination remain unclear. We performed a proteomic analysis of citrullinated proteins, and we identified more than 100 PAD4 (peptidyl arginine deiminase 4) substrates. Approximately one-fifth of the PAD4 substrates contained an RG/RGG motif, and PAD4 competitively inhibited the methylation of the RGG motif in FET proteins (FUS, EWS, and TAF15) and hnRNPA1, which are causative genes for ALS (amyotrophic lateral sclerosis). PAD4-mediated citrullination significantly inhibited the aggregation of FET proteins, a frequently observed feature in neurodegenerative diseases. FUS protein levels in arsenic-induced stress granules were significantly increased in Padi4-/- mouse embryonic fibroblasts (MEFs). Moreover, rs2240335 was associated with low expression of PADI4 in the brain and a high risk of ALS (p = 0.0381 and odds ratio of 1.072). Our findings suggest that PAD4-mediated RGG citrullination plays a key role in protein solubility and ALS pathogenesis.

11.
Sci Rep ; 7(1): 10739, 2017 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-28878391

RESUMO

p53 encodes a transcription factor that transactivates downstream target genes involved in tumour suppression. Although osteosarcoma frequently has p53 mutations, the role of p53 in osteosarcomagenesis is not fully understood. To explore p53-target genes comprehensively in calvarial bone and find out novel druggable p53 target genes for osteosarcoma, we performed RNA sequencing using the calvarial bone and 23 other tissues from p53 +/+ and p53 -/- mice after radiation exposure. Of 23,813 genes, 69 genes were induced more than two-fold in irradiated p53 +/+ calvarial bone, and 127 genes were repressed. Pathway analysis of the p53-induced genes showed that genes associated with cytokine-cytokine receptor interactions were enriched. Three genes, CD137L, CDC42 binding protein kinase gamma and Follistatin, were identified as novel direct p53 target genes that exhibited growth-suppressive effects on osteosarcoma cell lines. Of the three genes, costimulatory molecule Cd137l was induced only in calvarial bone among the 24 tissues tested. CD137L-expressing cells exhibited growth-suppressive effects in vivo. In addition, recombinant Fc-fusion Cd137l protein activated the immune response in vitro and suppressed osteosarcoma cell growth in vivo. We clarified the role of CD137L in osteosarcomagenesis and its potential therapeutic application. Our transcriptome analysis also indicated the regulation of the immune response through p53.

12.
Oncotarget ; 8(34): 55790-55803, 2017 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-28915553

RESUMO

p53 mutation is a marker of poor prognosis in breast cancers. To identify downstream targets of p53, we screened two transcriptome datasets, including cDNA microarrays of MCF10A breast epithelial cells with wild-type p53 or p53-null background, and RNA sequence analysis of breast invasive carcinoma. Here, we unveil ten novel p53 target candidates that are up-regulated after the induction of p53 in wild-type cells. Their expressions are also high in breast invasive carcinoma tissues with wild-type p53. The GO analysis identified epidermis development and ectoderm development, which COL17A1 participates, as significantly up-regulated by wild-type p53. The COL17A1 expressions increased in a p53-dependent manner in human breast cells and mouse mammary tissues. Reporter assay and ChIP assay identified intronic p53-binding sequences in the COL17A1 gene. The MDA-MB-231 cells that genetically over-express COL17A1 gene product exhibited reduced migration and invasion in vitro. Similarly, COL17A1 expression was decreased in metastatic tumors compared to primary tumors and normal tissues, even from the same patients. Moreover, high COL17A1 expression was associated with longer survival of patients with invasive breast carcinoma. In conclusion, we revealed that COL17A1 is a novel p53 transcriptional target in breast tissues that inhibits cell migration and invasion and is associated with better prognosis.

13.
Oncotarget ; 8(34): 55821-55836, 2017 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-28915555

RESUMO

The p53 protein is a sophisticated transcription factor that regulates dozens of target genes simultaneously in accordance with the cellular circumstances. Although considerable efforts have been made to elucidate the functions of p53-induced genes, a holistic understanding of the orchestrated signaling network repressed by p53 remains elusive. Here, we performed a systematic analysis to identify simultaneously regulated p53-repressed genes in breast cancer cells. Consequently, 28 genes were designated as the p53-repressed gene module, whose gene components were simultaneously suppressed in breast cancer cells treated with Adriamycin. A ChIP-seq database showed that p53 does not preferably bind to the region around the transcription start site of the p53-repressed gene module elements compared with that of p53-induced genes. Furthermore, we demonstrated that p21/CDKN1A plays a pivotal role in the suppression of the p53-repressed gene module in breast cancer cells. Finally, we showed that appropriate suppression of some genes belonging to the p53-repressed gene module contributed to a better prognosis of breast cancer patients. Taken together, these findings disentangle the gene regulatory network underlying the built-in p53-mediated tumor suppression system.

14.
Int J Oncol ; 51(2): 724-736, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28714518

RESUMO

p53, one of the most frequently mutated genes in colon cancer, suppresses cancer development through transactivation of its targets. Herein, we conducted a comprehensive analysis of the p53 downstream pathway in colorectal cancer by using multi-omics analysis. Mass spectrometric analysis of HCT116 p53+/+ and HCT116 p53-/- cells treated with adriamycin identified 124 proteins increased by DNA damage in a p53-dependent manner. Further screening using a cDNA microarray and the TCGA database revealed MICALL1 as a novel p53 target, and we identified functional p53 binding motifs located approximately 3000 base pairs upstream of the MICALL1 gene. MICALL1 expression was significantly decreased in colorectal cancer tissues with p53 mutation compared with those without p53 mutation. In response to DNA damage, MICALL1 co-localized with RAB8A and CD2AP at tubular recycling endosomes, whereas these proteins hardly localized at tubular recycling endosomes when p53 or MICALL1 expression was inhibited by siRNA. Our findings show that p53 regulates tubular recycling endosome biogenesis via transcriptional regulation of MICALL1, whose expression is frequently suppressed in colorectal cancer tissues.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias Colorretais/metabolismo , Proteínas do Citoesqueleto/genética , Doxorrubicina/farmacologia , Proteínas com Domínio LIM/genética , Proteína Supressora de Tumor p53/genética , Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sítios de Ligação , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HCT116 , Humanos , Proteínas com Domínio LIM/química , Proteínas com Domínio LIM/metabolismo , Espectrometria de Massas , Mutação , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Proteômica/métodos , Transdução de Sinais/efeitos dos fármacos , Proteínas rab de Ligação ao GTP/metabolismo
15.
EBioMedicine ; 20: 109-119, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28558959

RESUMO

Although recent cancer genomics studies have identified a large number of genes that were mutated in human cancers, p53 remains as the most frequently mutated gene. To further elucidate the p53-signalling network, we performed transcriptome analysis on 24 tissues in p53+/+ or p53-/- mice after whole-body X-ray irradiation. Here we found transactivation of a total of 3551 genes in one or more of the 24 tissues only in p53+/+ mice, while 2576 genes were downregulated. p53 mRNA expression level in each tissue was significantly associated with the number of genes upregulated by irradiation. Annotation using TCGA (The Cancer Genome Atlas) database revealed that p53 negatively regulated mRNA expression of several cancer therapeutic targets or pathways such as BTK, SYK, and CTLA4 in breast cancer tissues. In addition, stomach exhibited the induction of Krt6, Krt16, and Krt17 as well as loricrin, an epidermal differentiation marker, after the X-ray irradiation only in p53+/+ mice, implying a mechanism to protect damaged tissues by rapid induction of differentiation. Our comprehensive transcriptome analysis elucidated tissue specific roles of p53 and its signalling networks in DNA-damage response that will enhance our understanding of cancer biology.


Assuntos
Regulação da Expressão Gênica , Transdução de Sinais , Transcrição Genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Animais , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Biologia Computacional , Dano ao DNA , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Camundongos , Camundongos Knockout , Processamento de RNA , Transcriptoma
16.
Sci Adv ; 3(5): e1603204, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28560349

RESUMO

The transcription factor p53 is at the core of a built-in tumor suppression system that responds to varying degrees of stress input and is deregulated in most human cancers. Befitting its role in maintaining cellular fitness and fidelity, p53 regulates an appropriate set of target genes in response to cellular stresses. However, a comprehensive understanding of this scheme has not been accomplished. We show that argininosuccinate synthase 1 (ASS1), a citrulline-aspartate ligase in de novo arginine synthesis pathway, was directly transactivated by p53 in response to genotoxic stress, resulting in the rearrangement of arginine metabolism. Furthermore, we found that x-ray irradiation promoted the systemic induction of Ass1 and concomitantly increased plasma arginine levels in p53+/+ mice but not in p53-/- mice. Notably, Ass1+/- mice exhibited hypersensitivity to whole-body irradiation owing to increased apoptosis in the small intestinal crypts. Analyses of ASS1-deficient cells generated using the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 (CRISPR-associated 9) system revealed that ASS1 plays a pivotal role in limiting Akt phosphorylation. In addition, aberrant activation of Akt resulting from ASS1 loss disrupted Akt-mediated cell survival signaling activity under genotoxic stress. Building on these results, we demonstrated that p53 induced an intrinsic Akt repressor, ASS1, and the perturbation of ASS1 expression rendered cells susceptible to genotoxic stress. Our findings uncover a new function of p53 in the regulation of Akt signaling and reveal how p53, ASS1, and Akt are interrelated to each other.


Assuntos
Argininossuccinato Sintase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos da radiação , Ativação Transcricional/efeitos da radiação , Proteína Supressora de Tumor p53/metabolismo , Raios X/efeitos adversos , Animais , Apoptose/genética , Apoptose/efeitos da radiação , Argininossuccinato Sintase/genética , Sistemas CRISPR-Cas , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Sobrevivência Celular/efeitos da radiação , Humanos , Intestino Delgado/metabolismo , Intestino Delgado/patologia , Camundongos , Camundongos Knockout , Proteínas Proto-Oncogênicas c-akt/genética , Transdução de Sinais/genética , Ativação Transcricional/genética , Proteína Supressora de Tumor p53/genética
17.
Neoplasia ; 19(3): 185-195, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28152424

RESUMO

BACKGROUND & AIM: p53 activation by cellular stresses induces the transcription of hundreds of its target genes. To elucidate the entire picture of its downstream pathway, we screened a cDNA microarray dataset of adriamycin-treated HCT116 p53-/- or p53+/+ cells and identified EPSIN 3 as a novel p53 target. METHODS: Potential p53 binding sequences in the EPSIN 3 locus were evaluated by reporter and CHIP assays. To investigate the role of EPSIN 3 in the p53 downstream pathway, we assessed DNA damage-induced apoptosis in EPSIN 3-knockdown HCT116 cells or Epsin 3-deficient mice. In addition, we evaluated EPSIN 3 expression levels in various tissues, including gastric adenocarcinoma, human gastric mucosa with or without Helicobacter pylori infection, and mouse acute gastritis tissues induced by indomethacin. RESULTS: In response to DNA damage, p53 induced the expression of EPSIN 3 through the p53 binding elements in the EPSIN 3 promoter and the first intron. Knockdown of EPSIN 3 resulted in resistance to DNA damage-induced apoptosis both in vitro and in vivo. EPSIN 3 expression was down-regulated in gastric cancer tissues compared with normal tissues. In addition, Helicobacter pylori infection and indomethacin-induced acute gastritis repressed EPSIN 3 expression in gastric mucosa. CONCLUSIONS: EPSIN 3 is a novel p53 target and a key mediator of apoptosis. Chronic or acute mucosal inflammation as well as p53 inactivation induced down-regulation of EPSIN 3 and subsequently caused apoptosis resistance, which is a hallmark of cancer cells.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/genética , Apoptose/genética , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transdução de Sinais , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Linhagem Celular , Proliferação de Células , Gastrite/genética , Gastrite/metabolismo , Gastrite/microbiologia , Gastrite/patologia , Regulação Neoplásica da Expressão Gênica , Técnicas de Inativação de Genes , Células HCT116 , Humanos , Ligação Proteica , Proteólise , Neoplasias Gástricas/patologia , Estresse Fisiológico , Proteína Supressora de Tumor p53/genética
18.
Int J Oncol ; 48(6): 2415-24, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27035231

RESUMO

In response to various cellular stresses, p53 exerts its tumor suppressive effects such as apoptosis, cell cycle arrest, and senescence through the induction of its target genes. Recently, p53 was shown to control cellular homeostasis by regulating energy metabolism, glycolysis, antioxidant effect, and autophagy. However, its function in inositol synthesis was not reported. Through a microarray screening, we found that five genes related with myo-inositol metabolism were induced by p53. DNA damage enhanced intracellular myo-inositol content in HCT116 p53+/+ cells, but not in HCT116 p53-/- cells. We also indicated that inositol 3-phosphate synthase (ISYNA1) which encodes an enzyme essential for myo-inositol biosynthesis as a direct target of p53. Activated p53 regulated ISYNA1 expression through p53 response element in the seventh exon. Ectopic ISYNA1 expression increased myo-inositol levels in the cells and suppressed tumor cell growth. Knockdown of ISYNA1 caused resistance to adriamycin treatment, demonstrating the role of ISYNA1 in p53-mediated growth suppression. Furthermore, ISYNA1 expression was significantly associated with p53 mutation in bladder, breast cancer, head and neck squamous cell carcinoma, lung squamous cell carcinoma, and pancreatic adenocarcinoma. Our findings revealed a novel role of p53 in myo-inositol biosynthesis which could be a potential therapeutic target.


Assuntos
Inositol/biossíntese , Mio-Inositol-1-Fosfato Sintase/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Células HCT116 , Células HEK293 , Células Hep G2 , Humanos , Camundongos , Camundongos Knockout , Mio-Inositol-1-Fosfato Sintase/genética , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteína Supressora de Tumor p53/genética
19.
Cancer Sci ; 107(3): 298-306, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26757339

RESUMO

In response to various cellular stresses, p53 is activated and inhibits malignant transformation through the transcriptional regulation of its target genes. However, the full picture of the p53 downstream pathway still remains to be elucidated. Here we identified cystatin C, a major inhibitor of cathepsins, as a novel p53 target. In response to DNA damage, activated p53 induced cystatin C expression through p53 binding sequence in the first intron. We showed that cathepsin L activity was decreased in HCT116 p53(+/+) cells after adriamycin treatment, but not in HCT116 p53(-/-) cells. We also found that knockdown of cystatin C reduced adriamycin-induced caspase-3 activation. Cystatin C expression was significantly downregulated in breast cancer cells with p53 mutations, and decreased cystatin C expression was associated with poor prognosis of breast cancer. Our findings revealed an important role of the p53-cystatin C pathway in human carcinogenesis.


Assuntos
Adenocarcinoma/metabolismo , Catepsina L/metabolismo , Cistatina C/fisiologia , Proteína Supressora de Tumor p53/fisiologia , Adenocarcinoma/genética , Adenocarcinoma/mortalidade , Apoptose , Sequência de Bases , Sítios de Ligação , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/mortalidade , Sequência Consenso , Dano ao DNA , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Modelos de Riscos Proporcionais , Ativação Transcricional
20.
Sci Rep ; 5: 16497, 2015 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-26560363

RESUMO

Accumulation of iron in tissues increases the risk of cancer, but iron regulatory mechanisms in cancer tissues are largely unknown. Here, we report that p53 regulates iron metabolism through the transcriptional regulation of ISCU (iron-sulfur cluster assembly enzyme), which encodes a scaffold protein that plays a critical role in Fe-S cluster biogenesis. p53 activation induced ISCU expression through binding to an intronic p53-binding site. Knockdown of ISCU enhanced the binding of iron regulatory protein 1 (IRP1), a cytosolic Fe-S protein, to an iron-responsive element in the 5' UTR of ferritin heavy polypeptide 1 (FTH1) mRNA and subsequently reduced the translation of FTH1, a major iron storage protein. In addition, in response to DNA damage, p53 induced FTH1 and suppressed transferrin receptor, which regulates iron entry into cells. HCT116 p53(+/+) cells were resistant to iron accumulation, but HCT116 p53(-/-) cells accumulated intracellular iron after DNA damage. Moreover, excess dietary iron caused significant elevation of serum iron levels in p53(-/-) mice. ISCU expression was decreased in the majority of human liver cancer tissues, and its reduced expression was significantly associated with p53 mutation. Our finding revealed a novel role of the p53-ISCU pathway in the maintenance of iron homeostasis in hepatocellular carcinogenesis.


Assuntos
Homeostase , Proteínas com Ferro-Enxofre/metabolismo , Ferro/metabolismo , Transdução de Sinais , Proteína Supressora de Tumor p53/metabolismo , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Dano ao DNA , Ferritinas/genética , Ferritinas/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Técnicas de Inativação de Genes , Humanos , Espaço Intracelular/metabolismo , Proteína 1 Reguladora do Ferro/genética , Proteína 1 Reguladora do Ferro/metabolismo , Proteínas com Ferro-Enxofre/genética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Camundongos , Camundongos Knockout , Modelos Biológicos , Ligação Proteica , Isoformas de RNA , Proteína Supressora de Tumor p53/genética
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