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1.
PNAS Nexus ; 2(7): pgad220, 2023 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-37448957

RESUMEN

Mammalian genomes encode large number of long noncoding RNAs (lncRNAs) that play key roles in various biological processes, including proliferation, differentiation, and stem cell pluripotency. Recent studies have addressed that some lncRNAs are dysregulated in human cancers and may play crucial roles in tumor development and progression. Here, we show that the lncRNA ZNNT1 is required for the proliferation and tumorigenicity of colon cancer cells with wild-type p53. ZNNT1 knockdown leads to decreased ubiquitination and stabilization of p53 protein. Moreover, we demonstrate that ZNNT1 needs to interact with SART3 to destabilize p53 and to promote the proliferation and tumorigenicity of colon cancer cells. We further show that SART3 is associated with the ubiquitin-specific peptidase USP15 and that ZNNT1 may induce p53 destabilization by inhibiting this interaction. These results suggest that ZNNT1 interferes with the SART3-USP15 complex-mediated stabilization of p53 protein and thereby plays important roles in the proliferation and tumorigenicity of colon cancer cells. Our findings suggest that ZNNT1 may be a promising molecular target for the therapy of colon cancer.

2.
Curr Issues Mol Biol ; 44(4): 1587-1596, 2022 Apr 07.
Artículo en Inglés | MEDLINE | ID: mdl-35723366

RESUMEN

Epithelial ovarian cancer is classified into four major histological subtypes: serous, clear cell, endometrioid and mucinous. Ovarian clear cell carcinoma (OCCC) responds poorly to conventional chemotherapies and shows poor prognosis. Thus, there is a need to develop new drugs for the treatment of OCCC. In this study, we performed CRISPR/Cas9 screens against OCCC cell lines and identified candidate genes important for their proliferation. We found that quite different genes are required for the growth of ARID1A and PIK3CA mutant and wild-type OCCC cell lines, respectively. Furthermore, we found that the epigenetic regulator KDM2A and the translation regulator PAIP1 may play important roles in the growth of ARID1A and PIK3CA mutant, but not wild-type, OCCC cells. The results of our CRISPR/Cas9 screening may be useful in elucidating the molecular mechanism of OCCC tumorigenesis and in developing OCCC-targeted drugs.

3.
Cancer Sci ; 113(6): 2034-2043, 2022 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-35377528

RESUMEN

Ovarian cancer is the fifth most common cause of cancer-related death in women. Ovarian clear cell carcinoma (OCCC) is a chemotherapy-resistant epithelial ovarian cancer with poor prognosis. As a basis for the development of therapeutic agents that could improve the prognosis of OCCC, we performed a screen for proteins critical for the tumorigenicity of OCCC using the CRISPR/Cas9 system. Here we show that knockdown of the phosphate exporter XPR1/SLC53A1 induces the growth arrest and apoptosis of OCCC cells in vitro. Moreover, we show that knockdown of XPR1/SLC53A1 inhibits the proliferation of OCCC cells xenografted into immunocompromised mice. These results suggest that XPR1/SLC53A1 plays a critical role in the tumorigenesis of OCCC cells. We speculate that XPR1/SLC53A1 might be a promising molecular target for the therapeutic treatment of OCCC.


Asunto(s)
Adenocarcinoma de Células Claras , Neoplasias Ováricas , Adenocarcinoma de Células Claras/patología , Animales , Carcinoma Epitelial de Ovario/genética , Femenino , Humanos , Ratones , Neoplasias Ováricas/patología , Fosfatos/uso terapéutico , Pronóstico
4.
J Neurosci ; 42(12): 2448-2473, 2022 03 23.
Artículo en Inglés | MEDLINE | ID: mdl-35121636

RESUMEN

Signal-induced proliferation-associated 1 (SIPA1)-like 1 (SIPA1L1; also known as SPAR1) has been proposed to regulate synaptic functions that are important in maintaining normal neuronal activities, such as regulating spine growth and synaptic scaling, as a component of the PSD-95/NMDA-R-complex. However, its physiological role remains poorly understood. Here, we performed expression analyses using super-resolution microscopy (SRM) in mouse brain and demonstrated that SIPA1L1 is mainly localized to general submembranous regions in neurons, but surprisingly, not to PSD. Our screening for physiological interactors of SIPA1L1 in mouse brain identified spinophilin and neurabin-1, regulators of G-protein-coupled receptor (GPCR) signaling, but rejected PSD-95/NMDA-R-complex components. Furthermore, Sipa1l1-/- mice showed normal spine size distribution and NMDA-R-dependent synaptic plasticity. Nevertheless, Sipa1l1-/- mice showed aberrant responses to α2-adrenergic receptor (a spinophilin target) or adenosine A1 receptor (a neurabin-1 target) agonist stimulation, and striking behavioral anomalies, such as hyperactivity, enhanced anxiety, learning impairments, social interaction deficits, and enhanced epileptic seizure susceptibility. Male mice were used for all experiments. Our findings revealed unexpected properties of SIPA1L1, suggesting a possible association of SIPA1L1 deficiency with neuropsychiatric disorders related to dysregulated GPCR signaling, such as epilepsy, attention deficit hyperactivity disorder (ADHD), autism, or fragile X syndrome (FXS).SIGNIFICANCE STATEMENT Signal-induced proliferation-associated 1 (SIPA1)-like 1 (SIPA1L1) is thought to regulate essential synaptic functions as a component of the PSD-95/NMDA-R-complex. In our screening for physiological SIPA1L1-interactors, we identified G-protein-coupled receptor (GPCR)-signaling regulators. Moreover, SIPA1L1 knock-out (KO) mice showed striking behavioral anomalies, which may be relevant to GPCR signaling. Our findings revealed an unexpected role of SIPA1L1, which may open new avenues for research on neuropsychiatric disorders that involve dysregulated GPCR signaling. Another important aspect of this paper is that we showed effective methods for checking PSD association and identifying native protein interactors that are difficult to solubilize. These results may serve as a caution for future claims about interacting proteins and PSD proteins, which could eventually save time and resources for researchers and avoid confusion in the field.


Asunto(s)
Proteínas Activadoras de GTPasa/metabolismo , N-Metilaspartato , Proteínas del Tejido Nervioso , Animales , Homólogo 4 de la Proteína Discs Large , Masculino , Ratones , Ratones Noqueados , Proteínas de Microfilamentos/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Receptor de Adenosina A1 , Receptores Acoplados a Proteínas G/metabolismo
5.
FEBS Open Bio ; 12(3): 582-593, 2022 03.
Artículo en Inglés | MEDLINE | ID: mdl-34965029

RESUMEN

Certain cancers, such as ovarian clear cell carcinoma (OCCC), display high levels of genetic variation between patients, making it difficult to develop effective therapies. In order to identify novel genes critical to OCCC growth, we carried out a comprehensive CRISPR-Cas9 knockout screen against cell growth using an OCCC cell line and a normal ovarian surface epithelium cell line. We identified the gene encoding DHX38/PRP16, an ATP-dependent RNA helicase involved in splicing, as critical for the growth and tumorigenesis of OCCC. DHX38/PRP16 knockdown in OCCC cells, but not normal cells, induces apoptosis and impairs OCCC tumorigenesis in a mouse model. Our results suggest that DHX38/PRP16 may play a role in OCCC tumorigenesis and could potentially be a promising therapeutic target.


Asunto(s)
Adenocarcinoma de Células Claras , Neoplasias Ováricas , Factores de Empalme de ARN , Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/metabolismo , Adenocarcinoma de Células Claras/patología , Animales , Sistemas CRISPR-Cas/genética , Carcinogénesis/genética , ARN Helicasas DEAD-box/genética , ARN Helicasas DEAD-box/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Neoplasias Ováricas/tratamiento farmacológico , Factores de Empalme de ARN/genética , Factores de Empalme de ARN/metabolismo , Factores de Empalme de ARN/uso terapéutico
6.
Nucleic Acids Res ; 49(18): e104, 2021 10 11.
Artículo en Inglés | MEDLINE | ID: mdl-34291282

RESUMEN

Single-cell RNA-seq (scRNA-seq) can be used to characterize cellular heterogeneity in thousands of cells. The reconstruction of a gene network based on coexpression patterns is a fundamental task in scRNA-seq analyses, and the mutual exclusivity of gene expression can be critical for understanding such heterogeneity. Here, we propose an approach for detecting communities from a genetic network constructed on the basis of coexpression properties. The community-based comparison of multiple coexpression networks enables the identification of functionally related gene clusters that cannot be fully captured through differential gene expression-based analysis. We also developed a novel metric referred to as the exclusively expressed index (EEI) that identifies mutually exclusive gene pairs from sparse scRNA-seq data. EEI quantifies and ranks the exclusive expression levels of all gene pairs from binary expression patterns while maintaining robustness against a low sequencing depth. We applied our methods to glioblastoma scRNA-seq data and found that gene communities were partially conserved after serum stimulation despite a considerable number of differentially expressed genes. We also demonstrate that the identification of mutually exclusive gene sets with EEI can improve the sensitivity of capturing cellular heterogeneity. Our methods complement existing approaches and provide new biological insights, even for a large, sparse dataset, in the single-cell analysis field.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Redes Reguladoras de Genes , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Análisis de la Célula Individual/métodos , Transcriptoma/genética , Humanos
7.
Oncogene ; 39(5): 1018-1030, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31582837

RESUMEN

The epigenetic factor UHRF1 regulates transcription by modulating DNA methylation and histone modification, and plays critical roles in proliferation, development, and tumorigenesis. Here, we show that Wnt/c-Myc signaling upregulates UHRF1, which in turn downregulates TUSC3, a candidate tumor suppressor gene that is frequently deleted or downregulated in several cancers. We also show that UHRF1-mediated downregulation of TUSC3 is required for the proliferation of colon cancer cells. Furthermore, we demonstrate that UHRF1 suppresses TUSC3 expression by interacting with methylated H3K14 and thereby suppressing the acetylation of H3K14 by the histone acetyltransferase KAT7. Our study provides evidence for the significance of UHRF1-KAT7-mediated regulation of histone methylation/acetylation in the proliferation of tumor cells and in a diverse set of biological processes controlled by Wnt/c-Myc signaling.


Asunto(s)
Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Neoplasias del Colon/patología , Regulación Neoplásica de la Expresión Génica , Histona Acetiltransferasas/metabolismo , Histonas/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Acetilación , Línea Celular Tumoral , Proliferación Celular , Humanos , Metilación , Proteínas Proto-Oncogénicas c-myc/metabolismo , Vía de Señalización Wnt
8.
EMBO Rep ; 19(11)2018 11.
Artículo en Inglés | MEDLINE | ID: mdl-30213795

RESUMEN

Glioblastoma is one of the most aggressive forms of cancers and has a poor prognosis. Genomewide analyses have revealed that a set of core signaling pathways, the p53, RB, and RTK pathways, are commonly deregulated in glioblastomas. However, the molecular mechanisms underlying the tumorigenicity of glioblastoma are not fully understood. Here, we show that the lysine deacetylase SIRT2 is required for the proliferation and tumorigenicity of glioblastoma cells, including glioblastoma stem cells. Furthermore, we demonstrate that SIRT2 regulates p73 transcriptional activity by deacetylation of its C-terminal lysine residues. Our results suggest that SIRT2-mediated inactivation of p73 is critical for the proliferation and tumorigenicity of glioblastoma cells and that SIRT2 may be a promising molecular target for the therapy of glioblastoma.


Asunto(s)
Neoplasias Encefálicas/patología , Glioblastoma/patología , Sirtuina 2/metabolismo , Proteína Tumoral p73/metabolismo , Acetilación , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Neoplasias Encefálicas/metabolismo , Proliferación Celular , Furanos/farmacología , Técnicas de Silenciamiento del Gen , Glioblastoma/metabolismo , Humanos , Lisina/metabolismo , Ratones Desnudos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Quinolinas/farmacología , Sirtuina 2/antagonistas & inhibidores , Sirtuina 2/genética , Células Tumorales Cultivadas , Proteína Tumoral p73/genética , Ensayos Antitumor por Modelo de Xenoinjerto
9.
Oncol Rep ; 40(1): 309-318, 2018 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-29749539

RESUMEN

Ovarian clear cell carcinoma (OCCC) is a chemotherapy­resistant epithelial ovarian cancer with poor prognosis. To identify genomic alterations involved in the development of OCCC, we analyzed somatic copy number alterations in OCCC using comparative genomic hybridization (CGH). Here we showed that the chromosomal regions 8p11.21, 8p11.22, 12p13.31 and 20q13.2 were amplified in OCCC. We also demonstrated that small segments in the chromosomal regions 3q26.1, 4q13.2 and 22q11.23 were deleted. Kaplan­Meier survival analyses revealed that patients with amplification within 8p11.21 or a deletion within 3q26.1 had a shorter progression­free survival (PFS) time than those without such alterations. In addition, patients with amplification in three of the four chromosomal regions 8p11.21, 8p11.22, 12p13.31 and 20q13.2 had shorter overall survival (OS). We also demonstrated that amplification of 12p13.3 or three of the four chromosomal regions 8p11.21, 8p11.22, 12p13.31 and 20q13.2, or a deletion in the chromosomal region 3q26.1 was associated with chemotherapy resistance. Our findings suggest that copy number alterations in 8p11.21­22, 12p13.31, 20q13.2, 3q26.1, 4q13.2 and 22q11.23 are critical for the development and survival of OCCC.


Asunto(s)
Adenocarcinoma de Células Claras/genética , Variaciones en el Número de Copia de ADN/genética , Neoplasias Ováricas/genética , Pronóstico , Adenocarcinoma de Células Claras/patología , Adulto , Anciano , Anciano de 80 o más Años , Hibridación Genómica Comparativa , Supervivencia sin Enfermedad , Femenino , Amplificación de Genes/genética , Dosificación de Gen , Humanos , Estimación de Kaplan-Meier , Persona de Mediana Edad , Neoplasias Ováricas/patología , Ovario/patología
10.
Oncotarget ; 9(20): 15266-15274, 2018 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-29632642

RESUMEN

Ovarian clear cell carcinoma (OCCC) exhibits distinct phenotypes, such as resistance to chemotherapy, poor prognosis and an association with endometriosis. Biomarkers and imaging techniques currently in use are not sufficient for reliable diagnosis of this tumor or prediction of therapeutic response. It has recently been reported that analysis of somatic mutations in cell-free circulating DNA (cfDNA) released from tumor tissues can be useful for tumor diagnosis. In the present study, we attempted to detect mutations in PIK3CA and KRAS in cfDNA from OCCC patients using droplet digital PCR (ddPCR). Here we show that we were able to specifically detect PIK3CA-H1047R and KRAS-G12D in cfDNA from OCCC patients and monitor their response to therapy. Furthermore, we found that by cleaving wild-type PIK3CA using the CRISPR/Cas9 system, we were able to improve the sensitivity of the ddPCR method and detect cfDNA harboring PIK3CA-H1047R. Our results suggest that detection of mutations in cfDNA by ddPCR would be useful for the diagnosis of OCCC, and for predicting its recurrence.

11.
Oncol Lett ; 15(3): 4005-4009, 2018 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-29456745

RESUMEN

The Wnt signaling pathway is aberrantly activated in the majority of human colorectal tumors. ß-catenin, a key component of the Wnt signaling pathway, interacts with the T-cell factor/lymphoid enhancer-binding factor family of transcription factors and activates transcription of Wnt target genes. Sp5 is one of the Wnt target genes, and its expression is commonly upregulated in colon cancer cells. The present study demonstrates that the expression of Sp5 is not upregulated in the colon cancer cell line HCT116, in which Wnt signaling is constitutively activated. Furthermore, the results demonstrate that Sp5 has the potential to inhibit cell proliferation through upregulation of the cell cycle inhibitor p27. These findings suggest that HCT116 cells downregulate Sp5 to avoid p27-mediated growth arrest.

12.
Gastroenterology ; 150(5): 1171-1182, 2016 05.
Artículo en Inglés | MEDLINE | ID: mdl-26873401

RESUMEN

BACKGROUND & AIMS: Esophageal squamous cell carcinoma (ESCC) is the predominant form of esophageal cancer in Japan. Smoking and drinking alcohol are environmental risk factors for ESCC, whereas single nucleotide polymorphisms in ADH1B and ALDH2, which increase harmful intermediates produced by drinking alcohol, are genetic risk factors. We conducted a large-scale genomic analysis of ESCCs from patients in Japan to determine the mutational landscape of this cancer. METHODS: We performed whole-exome sequence analysis of tumor and nontumor esophageal tissues collected from 144 patients with ESCC who underwent surgery at 5 hospitals in Japan. We also performed single-nucleotide polymorphism array-based copy number profile and germline genotype analyses of polymorphisms in ADH1B and ALDH2. Polymorphisms in CYP2A6, which increase harmful effects of smoking, were analyzed. Functions of TET2 mutants were evaluated in KYSE410 and HEK293FT cells. RESULTS: A high proportion of mutations in the 144 tumor samples were C to T substitution in CpG dinucleotides (called the CpG signature) and C to G/T substitutions with a flanking 5' thymine (called the APOBEC signature). Based on mutational signatures, patients were assigned to 3 groups, which associated with environmental (drinking and smoking) and genetic (polymorphisms in ALDH2 and CYP2A6) factors. Many tumors contained mutations in genes that regulate the cell cycle (TP53, CCND1, CDKN2A, FBXW7); epigenetic processes (MLL2, EP300, CREBBP, TET2); and the NOTCH (NOTCH1, NOTCH3), WNT (FAT1, YAP1, AJUBA) and receptor-tyrosine kinase-phosphoinositide 3-kinase signaling pathways (PIK3CA, EGFR, ERBB2). Mutations in EP300 and TET2 correlated with shorter survival times, and mutations in ZNF750 associated with an increased number of mutations of the APOBEC signature. Expression of mutant forms of TET2 did not increase cellular levels of 5-hydroxymethylcytosine in HEK293FT cells, whereas knockdown of TET2 increased the invasive activity of KYSE410 ESCC cells. Computational analyses associated the mutations in NFE2L2 we identified with transcriptional activation of its target genes. CONCLUSIONS: We associated environmental and genetic factors with base substitution patterns of somatic mutations and provide a registry of genes and pathways that are disrupted in ESCCs. These findings might be used to design specific treatments for patients with esophageal squamous cancers.


Asunto(s)
Biomarcadores de Tumor/genética , Carcinoma de Células Escamosas/genética , Neoplasias Esofágicas/genética , Genómica , Mutación , Polimorfismo de Nucleótido Simple , Alcohol Deshidrogenasa/genética , Aldehído Deshidrogenasa Mitocondrial/genética , Pueblo Asiatico/genética , Biomarcadores de Tumor/metabolismo , Carcinoma de Células Escamosas/etnología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Islas de CpG , Citocromo P-450 CYP2A6/genética , Análisis Mutacional de ADN , Neoplasias Esofágicas/etnología , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patología , Carcinoma de Células Escamosas de Esófago , Exoma , Dosificación de Gen , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Interacción Gen-Ambiente , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Genómica/métodos , Células HEK293 , Humanos , Japón/epidemiología , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Factores de Riesgo , Transfección
13.
Biochem Biophys Res Commun ; 460(2): 216-21, 2015 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-25770425

RESUMEN

LGR5 plays an important role in the self-renewal of stem cells and is used as a marker identifying self-renewing stem cells in small intestine and hair follicles. Moreover, LGR5 has been reported to be overexpressed in several cancers. SOX9 is a transcription factor that plays a key role in development, differentiation and lineage commitment in various tissues. It has also been reported that SOX9 is overexpressed in a variety of cancers and contributes to their malignant phenotype. Here we show that LGR5 is required for the tumorigenicity of glioblastoma cells. We further show that SOX9 is upregulated in glioblastoma cells and directly enhances the expression of LGR5. We also demonstrate that knockdown of SOX9 suppresses the proliferation and tumorigenicity of glioblastoma cells. These results suggest that SOX9-mediated transcriptional regulation of LGR5 is critical for the tumorigenicity of glioblastoma cells. We speculate that the SOX9-LGR5 pathway could be a potentially promising target for the therapy of glioblastoma.


Asunto(s)
Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Receptores Acoplados a Proteínas G/fisiología , Factor de Transcripción SOX9/fisiología , Regulación hacia Arriba/fisiología , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Técnicas de Silenciamiento del Gen , Glioblastoma/patología , Humanos , Receptores Acoplados a Proteínas G/genética , Factor de Transcripción SOX9/genética
14.
Biochem Biophys Res Commun ; 459(3): 411-5, 2015 Apr 10.
Artículo en Inglés | MEDLINE | ID: mdl-25735981

RESUMEN

Upon binding of a Wnt ligand to the frizzled (FZD)-low density lipoprotein receptor related protein 5/6 (LRP5/6) receptor complex, the ß-catenin destruction complex, composed of Axin1, adenomatous polyposis coli (APC), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1), is immediately inactivated, which causes ß-catenin stabilization. However, the molecular mechanism of signal transduction from the receptor complex to the ß-catenin destruction complex is controversial. Here we show that Wnt3a treatment promotes the dissociation of the Axin1-APC complex in glioblastoma cells cultured in serum-free medium. Experiments with the GSK3 inhibitor BIO suggest that Axin1-APC dissociation was controlled by phosphorylation. Introduction of a phosphomimetic mutation into Thr160 of Axin1, located in the APC-binding region RGS, abrogated the interaction of Axin1 with APC. Consistent with these observations, the Axin1 phosphomimetic mutant lost the ability to reduce ß-catenin stability and to repress ß-catenin/TCF-dependent transcription. Taken together, our results suggest a novel mechanism of Wnt signaling through the dissociation of the ß-catenin destruction complex by Axin1 Thr160 modification.


Asunto(s)
Proteína Axina/química , Proteína Axina/metabolismo , Complejo de Señalización de la Axina/química , Complejo de Señalización de la Axina/metabolismo , Proteína de la Poliposis Adenomatosa del Colon/química , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Proteína Axina/genética , Sitios de Unión , Materiales Biomiméticos/química , Materiales Biomiméticos/metabolismo , Quinasa de la Caseína I/química , Quinasa de la Caseína I/metabolismo , Línea Celular Tumoral , Glioblastoma/metabolismo , Glucógeno Sintasa Quinasa 3/química , Glucógeno Sintasa Quinasa 3/metabolismo , Células HEK293 , Humanos , Proteína-5 Relacionada con Receptor de Lipoproteína de Baja Densidad/antagonistas & inhibidores , Proteína-5 Relacionada con Receptor de Lipoproteína de Baja Densidad/genética , Proteína-5 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismo , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/antagonistas & inhibidores , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/genética , Proteína-6 Relacionada a Receptor de Lipoproteína de Baja Densidad/metabolismo , Mutagénesis Sitio-Dirigida , Fosforilación , Estabilidad Proteica , ARN Interferente Pequeño/genética , Treonina/química , Vía de Señalización Wnt , beta Catenina/química , beta Catenina/metabolismo
15.
Bioorg Med Chem ; 23(2): 328-39, 2015 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-25515955

RESUMEN

Human sirtuin 2 (SIRT2) is an attractive target molecule for development of drugs to treat neurodegenerative diseases and cancer, because SIRT2 inhibitors have a protective effect against neurodegeneration and an anti-proliferative effect on cancer stem cells. We designed and synthesized a series of benzamide derivatives as SIRT2 inhibitor candidates. Among them, compound 17k showed the most potent SIRT2-inhibitory activity (IC50=0.60µM), with more than 150-fold selectivity over SIRT1 and SIRT3 isoforms (IC50 >100µM).


Asunto(s)
Benzamidas/química , Diseño de Fármacos , Inhibidores de Histona Desacetilasas/síntesis química , Sirtuina 2/antagonistas & inhibidores , Benzamidas/síntesis química , Benzamidas/metabolismo , Sitios de Unión , Inhibidores de Histona Desacetilasas/química , Inhibidores de Histona Desacetilasas/metabolismo , Humanos , Simulación del Acoplamiento Molecular , Unión Proteica , Estructura Terciaria de Proteína , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Sirtuina 1/antagonistas & inhibidores , Sirtuina 1/metabolismo , Sirtuina 2/genética , Sirtuina 2/metabolismo , Sirtuina 3/antagonistas & inhibidores , Sirtuina 3/metabolismo , Relación Estructura-Actividad
16.
Nat Commun ; 5: 3150, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24452072

RESUMEN

Aberrant activation of Wnt signalling results in colorectal tumours. Lgr5 is specifically expressed in stem cells of the intestine and has an essential role in maintaining tissue homeostasis. Lgr5-positive stem cells are responsible for the intestinal adenoma initiated by mutations in adenomatous polyposis coli. Furthermore, Lgr5 interacts with R-spondins and thereby activates Wnt signalling. However, the function of Lgr5 in colorectal tumourigenesis is unclear. Here we show that LGR5 is required for the tumourigenicity of colorectal cancer cells. We show that the transcription factor GATA6 directly enhances the expression of LGR5. We further demonstrate that GATA6 is upregulated in colorectal cancer cells due to the downregulation of miR-363, which directly targets GATA6. Moreover, we show that overexpression of miR-363 suppresses the tumourigenicity of colorectal cancer cells. These results suggest that the miR-363-GATA6-LGR5 pathway is critical for colorectal tumourigenesis and would be a promising target for the treatment of colorectal cancer.


Asunto(s)
Carcinogénesis , Factor de Transcripción GATA6/metabolismo , MicroARNs/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Humanos
17.
Biochem Biophys Res Commun ; 444(1): 13-8, 2014 Jan 31.
Artículo en Inglés | MEDLINE | ID: mdl-24406169

RESUMEN

Protocadherin10 (PCDH10)/OL-protocadherin is a cadherin-related transmembrane protein that has multiple roles in the brain, including facilitating specific cell-cell connections, cell migration and axon guidance. It has recently been reported that PCDH10 functions as a tumor suppressor and that its overexpression inhibits proliferation or invasion of multiple tumor cells. However, the function of PCDH10 in glioblastoma cells has not been elucidated. In contrast to previous reports on other tumors, we show here that suppression of the expression of PCDH10 by RNA interference (RNAi) induces the growth arrest and apoptosis of glioblastoma cells in vitro. Furthermore, we demonstrate that knockdown of PCDH10 inhibits the growth of glioblastoma cells xenografted into immunocompromised mice. These results suggest that PCDH10 is required for the proliferation and tumorigenicity of glioblastoma cells. We speculate that PCDH10 may be a promising target for the therapy of glioblastoma.


Asunto(s)
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Cadherinas/metabolismo , Glioblastoma/metabolismo , Glioblastoma/patología , Animales , Apoptosis , Neoplasias Encefálicas/genética , Cadherinas/antagonistas & inhibidores , Cadherinas/genética , Línea Celular Tumoral , Proliferación Celular , Técnicas de Silenciamiento del Gen , Glioblastoma/genética , Xenoinjertos , Humanos , Ratones , Ratones Desnudos , Invasividad Neoplásica , Proteínas Oncogénicas/antagonistas & inhibidores , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Protocadherinas , Interferencia de ARN
18.
Structure ; 19(10): 1496-508, 2011 Oct 12.
Artículo en Inglés | MEDLINE | ID: mdl-22000517

RESUMEN

Adenomatous polyposis coli (APC) is a tumor suppressor protein commonly mutated in colorectal tumors. APC plays important roles in Wnt signaling and other cellular processes. Here, we present the crystal structure of the armadillo repeat (Arm) domain of APC, which facilitates the binding of APC to various proteins. APC-Arm forms a superhelix with a positively charged groove. We also determined the structure of the complex of APC-Arm with the tyrosine-rich (YY) domain of the Src-associated in mitosis, 68 kDa protein (Sam68), which regulates TCF-1 alternative splicing. Sam68-YY forms numerous interactions with the residues on the groove and is thereby fixed in a bent conformation. We assessed the effects of mutations and phosphorylation on complex formation between APC-Arm and Sam68-YY. Structural comparisons revealed different modes of ligand recognition between the Arm domains of APC and other Arm-containing proteins.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/química , Proteína de la Poliposis Adenomatosa del Colon/química , Proteínas de Unión al ADN/química , Complejos Multiproteicos/química , Proteínas de Unión al ARN/química , Empalme Alternativo , Clonación Molecular , Simulación por Computador , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Mutación Missense , Fosforilación , Unión Proteica , Mapeo de Interacción de Proteínas , Estructura Secundaria de Proteína , Selenometionina/química , Difracción de Rayos X
19.
EMBO Rep ; 12(7): 682-9, 2011 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-21617703

RESUMEN

Tumour cells are known to be dependent on, or 'addicted to', not only oncogenes, but also some non-oncogenes. However, the mechanisms by which tumour cells are addicted to these genes have not been fully explained. Here, we show that overexpression of a member of the ETS family, EHF, is required for the survival of colon tumour cells that contain wild-type p53. We found that EHF directly activates the transcription of RUVBL1, an ATPase associated with chromatin-remodelling complexes. RUVBL1 blocks p53-mediated apoptosis by repressing the expression of p53 and its target genes. Moreover, we found that RUVBL1 represses p53 transcription by binding to the p53 promoter, interfering with RNF20/hBRE1-mediated histone H2B monoubiquitination and promoting PAF1-mediated histone H3K9 trimethylation. These results indicate that EHF-mediated RUVBL1 expression allows colon tumour cells to avoid p53-mediated apoptosis. Thus, EHF and RUVBL1 might be promising molecular targets for the treatment of colon tumours.


Asunto(s)
Apoptosis , Proteínas Portadoras/metabolismo , ADN Helicasas/metabolismo , Factores de Transcripción/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , ATPasas Asociadas con Actividades Celulares Diversas , Apoptosis/genética , Proteínas Portadoras/genética , Línea Celular Tumoral , Supervivencia Celular/genética , ADN Helicasas/genética , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HCT116 , Histonas/metabolismo , Humanos , Metilación , Transducción de Señal/genética , Factores de Transcripción/genética , Proteína p53 Supresora de Tumor/genética , Ubiquitinación , Regulación hacia Arriba
20.
J Biol Chem ; 285(21): 16145-54, 2010 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-20308060

RESUMEN

We have recently shown that beta-catenin-facilitated export of cadherins from the endoplasmic reticulum requires PX-RICS, a beta-catenin-interacting GTPase-activating protein for Cdc42. Here we show that PX-RICS interacts with isoforms of 14-3-3 and couples the N-cadherin-beta-catenin complex to the microtubule-based molecular motor dynein-dynactin. Similar to knockdown of PX-RICS, knockdown of either 14-3-3zeta or - resulted in the disappearance of N-cadherin and beta-catenin from the cell-cell boundaries. Furthermore, we found that PX-RICS and 14-3-3zeta/ are present in a large multiprotein complex that contains dynein-dynactin components as well as N-cadherin and beta-catenin. Both RNAi- and dynamitin-mediated inhibition of dynein-dynactin function also led to the absence of N-cadherin and beta-catenin at the cell-cell contact sites. Our results suggest that the PX-RICS-14-3-3zeta/ complex links the N-cadherin-beta-catenin cargo with the dynein-dynactin motor and thereby mediates its endoplasmic reticulum export.


Asunto(s)
Proteínas 14-3-3/metabolismo , Antígenos CD/metabolismo , Cadherinas/metabolismo , Dineínas/metabolismo , Retículo Endoplásmico/metabolismo , Proteínas Activadoras de GTPasa/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Complejos Multiproteicos/metabolismo , beta Catenina/metabolismo , Proteínas 14-3-3/genética , Animales , Antígenos CD/genética , Células COS , Cadherinas/genética , Chlorocebus aethiops , Complejo Dinactina , Dineínas/genética , Retículo Endoplásmico/genética , Proteínas Activadoras de GTPasa/genética , Técnicas de Silenciamiento del Gen , Células HeLa , Humanos , Proteínas Asociadas a Microtúbulos/genética , Microtúbulos/genética , Microtúbulos/metabolismo , Complejos Multiproteicos/genética , Transporte de Proteínas/fisiología , beta Catenina/genética , Proteína de Unión al GTP cdc42/genética , Proteína de Unión al GTP cdc42/metabolismo
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