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1.
PLoS One ; 14(4): e0215009, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30958856

RESUMO

Interorganelle phospholipid transfer is critical for eukaryotic membrane biogenesis. In the yeast Saccharomyces cerevisiae, phosphatidylserine (PS) synthesized by PS synthase, Pss1, in the endoplasmic reticulum (ER) is decarboxylated to phosphatidylethanolamine (PE) by PS decarboxylase, Psd1, in the ER and mitochondria or by Psd2 in the endosome, Golgi, and/or vacuole, but the mechanism of interorganelle PS transport remains to be elucidated. Here we report that Sfh1, a member of Sec14 family proteins of S. cerevisiae, possesses the ability to enhance PE production by Psd2. Overexpression of SFH1 in the strain defective in Psd1 restored its growth on non-fermentable carbon sources and increased the intracellular and mitochondrial PE levels. Sfh1 was found to bind various phospholipids, including PS, in vivo. Bacterially expressed and purified Sfh1 was suggested to have the ability to transport fluorescently labeled PS between liposomes by fluorescence dequenching assay in vitro. Biochemical subcellular fractionation suggested that a fraction of Sfh1 localizes to the endosome, Golgi, and/or vacuole. We propose a model that Sfh1 promotes PE production by Psd2 by transferring phospholipids between the ER and endosome.


Assuntos
Carboxiliases/deficiência , Proteínas de Ciclo Celular/biossíntese , Proteínas Cromossômicas não Histona/biossíntese , Mitocôndrias/metabolismo , Modelos Biológicos , Consumo de Oxigênio , Proteínas de Saccharomyces cerevisiae/biossíntese , Saccharomyces cerevisiae/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Endossomos/genética , Endossomos/metabolismo , Complexo de Golgi/genética , Complexo de Golgi/metabolismo , Mitocôndrias/genética , Fosfatidiletanolaminas/metabolismo , Fosfatidilserinas/genética , Fosfatidilserinas/metabolismo , Proteínas de Transferência de Fosfolipídeos/genética , Proteínas de Transferência de Fosfolipídeos/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Vacúolos/genética , Vacúolos/metabolismo
2.
Pharmazie ; 74(3): 154-156, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30961681

RESUMO

MicroRNAs (miRs) are crucial regulators for tumorigenesis through negatively regulating their target genes expression in the manner of 3'-untranslated region (3'-UTR) binding. MiR-205-5p has been reported to function as a tumor suppressor in several cancer types. The aim of this study was to investigate the role of miR-205-5p/chromobox homolog 1 (CBX1) axis in human pituitary tumors. The expression of miR-205-5p was firstly examined by quantitative real-time PCR and the results revealed that miR-205-5p expression was declined in pituitary cell lines compared with normal cell line. Overexpression of miR-205-5p effectively decreased cell proliferation and cell migration. Based on the results of bioinformatic analysis, luciferase reporter assay, and western blot, we identified CBX1 as a direct target of miR-205-5p. Notably, overexpression of CBX1 promoted cell proliferation and migration. The effects of miR-205-5p overexpression on cell proliferation and migration can be reversed by CBX1 overexpression. Based on these findings, we deducted that miR-205-5p inhibits the cell proliferation and migration through directly targeting CBX1.


Assuntos
Proteínas Cromossômicas não Histona/genética , MicroRNAs/genética , Neoplasias Hipofisárias/genética , Apoptose/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Proteínas Cromossômicas não Histona/biossíntese , Proteínas Cromossômicas não Histona/metabolismo , Progressão da Doença , Regulação para Baixo , Células HEK293 , Humanos , MicroRNAs/biossíntese , MicroRNAs/metabolismo , Neoplasias Hipofisárias/metabolismo , Neoplasias Hipofisárias/patologia
3.
Dokl Biochem Biophys ; 484(1): 66-68, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31012017

RESUMO

The PBAF(SWI/SNF) multiprotein complex, which changes the chromatin structure, is widely involved in the regulation of eukaryotic gene expression. A specific component of this complex is the PHF10 protein, which is involved in recruiting this complex to chromatin. We showed that the PHF10 expression in cells of different lines is activated by the c-MYC oncogene. Since PHF10 stimulates cell proliferation, its c-MYC-dependent activation in cancer cells should lead to an increase in their proliferation rate.


Assuntos
Proteínas Cromossômicas não Histona/biossíntese , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/biossíntese , Proteínas de Neoplasias/biossíntese , Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição/biossíntese , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/genética , Proteínas de Homeodomínio/genética , Humanos , Proteínas de Neoplasias/genética , Neoplasias/genética , Neoplasias/patologia , Proteínas Proto-Oncogênicas c-myc/genética , Fatores de Transcrição/genética
4.
J Exp Clin Cancer Res ; 38(1): 108, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30823889

RESUMO

BACKGROUND: Cancer cells are characterized by chromosomal instability (CIN) and it is thought that errors in pathways involved in faithful chromosome segregation play a pivotal role in the genesis of CIN. Cohesin forms a large protein ring that binds DNA strands by encircling them. In addition to this central role in chromosome segregation, cohesin is also needed for DNA repair, gene transcription regulation and chromatin architecture. Though mutations in both cohesin and cohesin-regulator genes have been identified in many human cancers, the contribution of cohesin to cancer development is still under debate. METHODS: Normal mucosa, early adenoma, and carcinoma samples deriving from 16 subjects affected by colorectal cancer (CRC) were analyzed by OncoScan for scoring both chromosome gains and losses (CNVs) and loss of heterozygosity (LOH). Then the expression of SMC1A was analyzed by immunochemistry in 66 subjects affected by CRC. The effects of SMC1A overexpression and mutated SMC1A were analyzed in vivo using immunocompromised mouse models. Finally, we measured global gene expression profiles in induced-tumors by RNA-seq. RESULTS: Here we showed that SMC1A cohesin core gene was present as extra-copies, mutated, and overexpressed in human colorectal carcinomas. We then demonstrated that cohesin overexpression led to the development of aggressive cancers in immunocompromised mice through gene expression dysregulation. CONCLUSION: Collectively, these results support a role of defective cohesin in the development of human colorectal cancer.


Assuntos
Adenocarcinoma/genética , Adenocarcinoma/patologia , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Adenoma/genética , Adenoma/patologia , Adulto , Idoso , Animais , Proteínas de Ciclo Celular/biossíntese , Instabilidade Cromossômica , Proteínas Cromossômicas não Histona/biossíntese , Feminino , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Pessoa de Meia-Idade
5.
BMB Rep ; 52(2): 157-162, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30760381

RESUMO

Our previous study found that two novel cancer-related genes, PRR11 and SKA2, constituted a classic gene pair that was regulated by p53 and NF-Y in lung cancer. However, their role and regulatory mechanism in breast cancer remain elusive. In this study, we found that the expression levels of PRR11 and SKA2 were upregulated and have a negative prognotic value in breast cancer. Loss-of-function experiments showed that RNAi-mediated knockdown of PRR11 and/or SKA2 inhibited proliferation, migration, and invasion of breast cancer cells. Mechanistic experiments revealed that knockdown of PRR11 and/or SKA2 caused dysregulation of several downstream genes, including CDK6, TPM3, and USP12, etc. Luciferase reporter assays demonstrated that wild type p53 significantly repressed the PRR11-SKA2 bidirectional promoter activity, but not NF-Y. Interestingly, NF-Y was only essential for and correlated with the expression of PRR11, but not SKA2. Consistently, adriamycin-induced (ADR) activation of endogenous p53 also caused significant repression of the PRR11 and SKA2 gene pair expression. Notably, breast cancer patients with lower expression levels of either PRR11 or SKA2, along with wild type p53, exhibited better disease-free survival compared to others with p53 mutations and/or higher expression levels of either PRR11 or SKA2. Collectively, our study indicates that the PRR11 and SKA2 transcription unit might be an oncogenic contributor and might serve as a novel diagnostic and therapeutic target in breast cancer. [BMB Reports 2019; 52(2): 157-162].


Assuntos
Neoplasias da Mama/genética , Proteínas Cromossômicas não Histona/genética , Proteínas/genética , Proteína Supressora de Tumor p53/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Proteínas Cromossômicas não Histona/biossíntese , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Células MCF-7 , Invasividade Neoplásica , Prognóstico , Regiões Promotoras Genéticas , Proteínas/metabolismo , Ativação Transcricional , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima
6.
Mol Cell ; 71(1): 129-141.e8, 2018 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-29979962

RESUMO

The enhancer regions of the myogenic master regulator MyoD give rise to at least two enhancer RNAs. Core enhancer eRNA (CEeRNA) regulates transcription of the adjacent MyoD gene, whereas DRReRNA affects expression of Myogenin in trans. We found that DRReRNA is recruited at the Myogenin locus, where it colocalizes with Myogenin nascent transcripts. DRReRNA associates with the cohesin complex, and this association correlates with its transactivating properties. Despite being expressed in undifferentiated cells, cohesin is not loaded on Myogenin until the cells start expressing DRReRNA, which is then required for cohesin chromatin recruitment and maintenance. Functionally, depletion of either cohesin or DRReRNA reduces chromatin accessibility, prevents Myogenin activation, and hinders muscle cell differentiation. Thus, DRReRNA ensures spatially appropriate cohesin loading in trans to regulate gene expression.


Assuntos
Proteínas de Ciclo Celular/biossíntese , Proteínas Cromossômicas não Histona/biossíntese , Elementos Facilitadores Genéticos , Músculo Esquelético/metabolismo , Miogenina/biossíntese , RNA não Traduzido/metabolismo , Transcrição Genética , Animais , Proteínas de Ciclo Celular/genética , Diferenciação Celular , Cromatina/genética , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/genética , Células HEK293 , Humanos , Camundongos , Músculo Esquelético/citologia , Proteína MyoD/biossíntese , Proteína MyoD/genética , Miogenina/genética , RNA não Traduzido/genética
7.
J Cell Sci ; 131(8)2018 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-29507117

RESUMO

Cullin-RING-type E3 ligases (CRLs) control a broad range of biological processes by ubiquitylating numerous cellular substrates. However, the role of CRL E3 ligases in chromatid cohesion is unknown. In this study, we identified a new CRL-type E3 ligase (designated as CRL7SMU1 complex) that has an essential role in the maintenance of chromatid cohesion. We demonstrate that SMU1, DDB1, CUL7 and RNF40 are integral components of this complex. SMU1, by acting as a substrate recognition module, binds to H2B and mediates monoubiquitylation at the lysine (K) residue K120 through CRL7SMU1 E3 ligase complex. Depletion of CRL7SMU1 leads to loss of H2B ubiquitylation at the SMC1a locus and, thus, subsequently compromised SMC1a expression in cells. Knockdown of CRL7SMU1 components or loss of H2B ubiquitylation leads to defective sister chromatid cohesion, which is rescued by restoration of SMC1a expression. Together, our results unveil an important role of CRL7SMU1 E3 ligase in promoting H2B ubiquitylation for maintenance of sister chromatid cohesion during mitosis.This article has an associated First Person interview with the first author of the paper.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Cromátides/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Histonas/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteínas de Ciclo Celular/biossíntese , Proteínas de Ciclo Celular/genética , Cromátides/genética , Proteínas Cromossômicas não Histona/biossíntese , Proteínas Cromossômicas não Histona/genética , Segregação de Cromossomos , Histonas/genética , Humanos , Transdução de Sinais , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
8.
BMC Cancer ; 18(1): 144, 2018 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-29409457

RESUMO

BACKGROUND: Limited data are available regarding the ability of biomarkers to predict complete pathological response to neoadjuvant chemoradiotherapy in locally advanced rectal cancer. Complete response translates to better patient survival. DEK is a transcription factor involved not only in development and progression of different types of cancer, but is also associated with treatment response. This study aims to analyze the role of DEK in complete pathological response following chemoradiotherapy for locally advanced rectal cancer. METHODS: Pre-treated tumour samples from 74 locally advanced rectal-cancer patients who received chemoradiation therapy prior to total mesorectal excision were recruited for construction of a tissue microarray. DEK immunoreactivity from all samples was quantified by immunohistochemistry. Then, association between positive stained tumour cells and pathologic response to neoadjuvant treatment was measured to determine optimal predictive power. RESULTS: DEK expression was limited to tumour cells located in the rectum. Interestingly, high percentage of tumour cells with DEK positiveness was statistically associated with complete pathological response to neoadjuvant treatment based on radiotherapy and fluoropyrimidine-based chemotherapy and a marked trend toward significance between DEK positiveness and absence of treatment toxicity. Further analysis revealed an association between DEK and the pro-apoptotic factor P38 in the pre-treated rectal cancer biopsies. CONCLUSIONS: These data suggest DEK as a potential biomarker of complete pathological response to treatment in locally advanced rectal cancer.


Assuntos
Biomarcadores Tumorais/biossíntese , Proteínas Cromossômicas não Histona/biossíntese , Proteínas Oncogênicas/biossíntese , Proteínas de Ligação a Poli-ADP-Ribose/biossíntese , Neoplasias Retais/metabolismo , Neoplasias Retais/terapia , Idoso , Idoso de 80 Anos ou mais , Quimiorradioterapia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Terapia Neoadjuvante , Valor Preditivo dos Testes , Prognóstico , Neoplasias Retais/patologia , Resultado do Tratamento
9.
Asia Pac J Clin Oncol ; 14(5): e283-e288, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29124886

RESUMO

AIM: Lung cancer is typically categorized into small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). NSCLC comprises of the majority of lung cancer with a poor prognosis in advanced cases. Transcriptional profiling studies, including microarrays and RNA-sequencing studies, have significantly enriched our knowledge of gene expression patterns in NSCLC. A recent transcriptional profiling study identified high prevalence of CBX3/HP1-gamma upregulation in human NSCLC samples. CBX3/HP1-gamma is an isoform of the heterochromatin protein 1 family, which plays a role in heterochromatin formation and is linked to cancer. METHODS: We examined lung cancer samples from our hospital using immunohistochemistry for CBX3/HP1-gamma staining. We also analyzed publicly available databases of NSCLC transcriptional profiling to validate our results. RESULTS: We identified a high prevalence (77.2%) of samples with positive CBX3/HP1-gamma staining by immunohistochemistry in NSCLC patient samples. Independently, we queried a publicly available dataset (GSE40419) containing RNA-seq data from 77 patients. Upregulation of CBX3/HP1-gamma in tumor samples was present in 60.2% of the patients. A similar correlation was also observed in the The Cancer Genome Atlas (TCGA) database. Interestingly, we discovered a highly significant association between positive CBX3/HP1-gamma staining and EGFR mutation in our patient samples (40 of 42 patients, P < 0.001). Treatment of EGFR mutant NSCLC cell lines with the EGFR inhibitor gefitinib failed to yield a change in CBX/HP1-gamma expression, suggesting that CBX/HP1-gamma expression may be independent of EGFR downstream signaling. CONCLUSION: We report a significant upregulation of CBX3/HP1-gamma in NSCLC patients, and also a possible relationship between CBX3/HP1-gamma expression and EGFR mutation.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Proteínas Cromossômicas não Histona/biossíntese , Neoplasias Pulmonares/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Receptores ErbB/genética , Feminino , Humanos , Imuno-Histoquímica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Mutação , Regulação para Cima
10.
Cancer Res ; 77(20): 5479-5490, 2017 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-28916652

RESUMO

Androgen receptor (AR) signaling is a key driver of prostate cancer, and androgen-deprivation therapy (ADT) is a standard treatment for patients with advanced and metastatic disease. However, patients receiving ADT eventually develop incurable castration-resistant prostate cancer (CRPC). Here, we report that the chromatin modifier LSD1, an important regulator of AR transcriptional activity, undergoes epigenetic reprogramming in CRPC. LSD1 reprogramming in this setting activated a subset of cell-cycle genes, including CENPE, a centromere binding protein and mitotic kinesin. CENPE was regulated by the co-binding of LSD1 and AR to its promoter, which was associated with loss of RB1 in CRPC. Notably, genetic deletion or pharmacological inhibition of CENPE significantly decreases tumor growth. Our findings show how LSD1-mediated epigenetic reprogramming drives CRPC, and they offer a mechanistic rationale for its therapeutic targeting in this disease. Cancer Res; 77(20); 5479-90. ©2017 AACR.


Assuntos
Proteínas Cromossômicas não Histona/metabolismo , Histona Desmetilases/genética , Neoplasias de Próstata Resistentes à Castração/enzimologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias da Próstata/embriologia , Neoplasias da Próstata/genética , Androgênios/metabolismo , Animais , Linhagem Celular Tumoral , Reprogramação Celular/genética , Proteínas Cromossômicas não Histona/biossíntese , Proteínas Cromossômicas não Histona/genética , Progressão da Doença , Epigênese Genética , Xenoenxertos , Histona Desmetilases/metabolismo , Humanos , Masculino , Camundongos , Neoplasias da Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/metabolismo , Transdução de Sinais , Transfecção
11.
Tumour Biol ; 39(7): 1010428317716248, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28670979

RESUMO

Astrocytic tumors are the most common neuroepithelial neoplasms with high relapse rate after surgery. Understanding the molecular mechanisms for astrocytic tumorigenesis and progression will lead to early diagnosis and effective treatment of astrocytic tumors. The DEK mRNA and protein expression in normal brain tissues and astrocytic tumors was quantified. To investigate DEK functions in tumor cells, DEK gene was silenced with siRNA in U251 glioblastoma cells. Cell proliferation, cell cycle and apoptosis were then measured. The expression and activity of key genes that regulate cell proliferation and apoptosis were also measured. We identified DEK as a high expressed gene in astrocytic tumor tissues. DEK expression level was positively correlated with the pathological grade of astrocytic tumors. Gene silencing of DEK in U251 glioblastomas inhibited cell proliferation and blocked cells at G0/G1 phase of cell cycle. DEK depletion also induced cell apoptosis, with up-regulated expression of P53 and P21 and down-regulated expression of Bcl-2 and C-myc. The Caspase-3 activity in U251 cells was also significantly increased after knockdown. Our results provided evidences that DEK regulates proliferation and apoptosis of glioblastomas. DEK gene silencing may induce apoptosis through P53-dependent pathway. Our data indicated DEK plays multiple roles to facilitate tumor growth and maintenance. It can be used as a potential target for astrocytic tumor diagnosis and gene therapy.


Assuntos
Astrocitoma/genética , Proliferação de Células/genética , Transformação Celular Neoplásica/genética , Proteínas Cromossômicas não Histona/biossíntese , Glioblastoma/genética , Proteínas Oncogênicas/biossíntese , Apoptose/genética , Astrocitoma/patologia , Caspase 3/biossíntese , Ciclo Celular/genética , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/antagonistas & inibidores , Proteínas Cromossômicas não Histona/genética , Inibidor de Quinase Dependente de Ciclina p21/biossíntese , Regulação Neoplásica da Expressão Gênica , Glioblastoma/patologia , Humanos , Proteínas Oncogênicas/antagonistas & inibidores , Proteínas Oncogênicas/genética , Proteínas de Ligação a Poli-ADP-Ribose , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Proteínas Proto-Oncogênicas c-myc/biossíntese , RNA Interferente Pequeno , Proteína Supressora de Tumor p53/biossíntese
12.
Cancer Biol Ther ; 18(3): 177-185, 2017 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-28278080

RESUMO

Drug resistance has always been the main problem in osteosarcoma treatment, and hypoxia seems to be one of the many causes for drug resistance. Therefore, in this study, we investigated how hypoxia triggers chemotherapy resistance in osteosarcoma. We first screened hypoxia- and normoxia- cultured osteosarcoma cells in silico to identify the differentially expressed genes specifically related to drug resistance. This led to the identification of spindle and kinetochore associated complex subunit 1 (SKA1) as a probable gene of interest. SKA1 was further overexpressed by a lentiviral vector into an osteosarcoma cell line to study its role in chemoresistance. Our data revealed that SKA1 overexpression reduced the expression of some multidrug resistance genes, and enhanced the sensitivity of two common chemotherapeutic drugs used in osteosarcoma patients, epirubicin (EPI) and ifosfamide (IFO). In addition, we also confirmed the role of SKA1 in EPI drug sensitivity in vivo. Taken together, our study indicated that hypoxia mediated downregulation of SKA1 expression increased the chemotherapy resistance in human osteosarcoma cells.


Assuntos
Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/genética , Hipóxia Celular/genética , Proteínas Cromossômicas não Histona/biossíntese , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Adolescente , Animais , Neoplasias Ósseas/metabolismo , Linhagem Celular Tumoral , Criança , Proteínas Cromossômicas não Histona/genética , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Camundongos Nus , Análise em Microsséries/métodos , Osteossarcoma/metabolismo , Adulto Jovem
13.
Micron ; 94: 37-45, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28027486

RESUMO

Chromatoid body (CB) is a cytoplasmic structure of male germ cells that has been indicated as having a role in the RNA and protein storage for the final differentiation of spermatozoa. Recent studies have indicated that some of these macromolecular complex components have nucleolar origin. The aims of the present study were to monitor the expression of fibrillarin nucleolar protein in mammalian seminiferous tubules at different stages of the spermatogenic cycle; to check fibrillarin distribution during the CB assembly; and also its interaction with two well-known CB markers (MIWI and HSP70). Seminiferous tubules were isolated by transilluminating microscope from testis of adult mice. Fibrillarin expression and also co-localization between fibrillarin and MIWI/HSP70 were performed by Western blot (WB) and by immunofluorescence (IF), respectively. Total proteins from testis of adult mice were also used to perform co-immunoprecipitation (Co-IP) experiments. Our results demonstrated higher fibrillarin expression in seminiferous tubules in stages IV-VI, and a close localization of fibrillarin with MIWI (a protein that plays a role in RNA metabolism in the CB), as well as with HSP70 (a protein that plays a role in the proteasome folding in the CB). We also performed Co-IP between fibrillarin/MIWI and between fibrillarin/HSP70 in order to determine whether MIWI or HSP70 interacts with this nucleolar protein. We found MIWI in the Co-IP precipitate, but not HSP70. In conclusion, our results show that fibrillarin may participate in the physiological activities performed by the CB by interacting with CB components that play a role in RNA metabolism.


Assuntos
Proteínas Cromossômicas não Histona/biossíntese , Substâncias Macromoleculares/metabolismo , RNA/metabolismo , Espermátides/metabolismo , Animais , Proteínas de Choque Térmico HSP70/metabolismo , Masculino , Camundongos , Túbulos Seminíferos/metabolismo , Espermatogênese/fisiologia
14.
PLoS One ; 11(8): e0156701, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27580131

RESUMO

Lung cancer is one of the most malignant tumors and the leading cause of cancer-related deaths worldwide. Among lung cancers, 40% are diagnosed as adenocarcinoma. Bromodomain containing 7 (BRD7) is a member of bromodomain-containing protein family. It was proved to be downregulated in various cancers. However, the role of BRD7 in lung adenocarcinoma is still unknown. Western blot and qRT-PCR was performed to measure the BRD7 expression in lung adenocarcinoma tissues and cells. CCK8 and migration assay was done to detect the functional role of BRD7 in lung adenocarcinoma. In this study, we showed that the expression of BRD7 was downregulated in lung adenocarcinoma tissues and cells. The lower of BRD7 levels in patients with lung adenocarcinoma was associated with shortened disease-free survival. Furthermore, overexpression of BRD7 inhibited lung adenocarcinoma cell proliferation and migration. Inhibition of BRD7 expression promoted cell proliferation and migration by activating ERK phosphorylation. Overexpression of BRD7 inhibited cyclin D and myc expression. Our findings are consistent with a tumor suppressor role for BRD7 in lung adenocarcinoma tumorigenesis.


Assuntos
Adenocarcinoma/metabolismo , Movimento Celular , Proteínas Cromossômicas não Histona/biossíntese , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas Supressoras de Tumor/biossíntese , Células A549 , Adenocarcinoma/genética , Adenocarcinoma/patologia , Proteínas Cromossômicas não Histona/genética , Ciclina D/biossíntese , Ciclina D/genética , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Proteínas Supressoras de Tumor/genética
15.
Sci Rep ; 6: 31752, 2016 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-27538348

RESUMO

Heterochromatin formation in fission yeast depends on RNAi machinery and histone-modifying enzymes. One of the key histone-modifying complexes is Clr4-Rik1-Cul4 methyltransferase complex (CLRC), which mediates histone H3K9 methylation, a hallmark for heterochromatin. CLRC is composed of the Clr4 histone methyltransferase, Rik1, Raf1, Raf2 and Pcu4. However, transcriptional regulation of the CLRC subunits is not well understood. In this study, we identified Set3, a core subunit of the Set3/Hos2 histone deacetylase complex (Set3C), as a contributor to the integrity and silencing of heterochromatin at centromeres, telomeres and silent mating-type locus. This novel role of Set3 relies on its PHD finger, but is independent of deacetylase activity or structural integrity of Set3C. Set3 is not located to the centromeric region. Instead, Set3 is targeted to the promoters of clr4(+) and rik1(+), probably through its PHD finger. Set3 promotes transcription of clr4(+) and rik1(+). Consistently, the protein levels of Clr4 and Rik1 were reduced in the set3Δ mutant. The heterochromatin silencing defect in the set3Δ mutant could be rescued by overexpressing of clr4(+) or rik1(+). Our study suggests transcriptional activation of essential heterochromatin factors underlies the tight regulation of heterochromatin integrity.


Assuntos
Proteínas de Ciclo Celular/biossíntese , Proteínas Cromossômicas não Histona/biossíntese , Proteínas Culina/biossíntese , Regulação Fúngica da Expressão Gênica/fisiologia , Heterocromatina/metabolismo , Histona Desacetilases/metabolismo , Metiltransferases/biossíntese , Proteínas de Schizosaccharomyces pombe/biossíntese , Schizosaccharomyces/metabolismo , Transcrição Genética/fisiologia , Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Proteínas Culina/genética , Heterocromatina/genética , Histona Desacetilases/genética , Metiltransferases/genética , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/genética
16.
Oncol Rep ; 36(4): 2382-90, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27499261

RESUMO

DEK is overexpressed in multiple invasive tumors. However, the transcriptional regulatory mechanism of DEK remains unclear. In the present study, progressive-type truncation assay indicated that CpG2-2 (-167 bp/+35 bp) was the DEK core promoter, whose methylation inhibited DEK expression. Bisulfite genomic sequencing analysis indicated that the methylation levels of the DEK promoter in normal hepatic cells and tissues were higher than those in hepatocellular carcinoma (HCC) cells. TFSEARCH result revealed transcription factor binding sites in CpG2-2. Among the sites, the AP-2α binding site showed the most significant methylation difference; hence, AP-2α is a key transcription factor that regulates DEK expression. Point or deletion mutation of the AP-2α binding site significantly reduced the promoter activity. Chromatin immunoprecipitation assay demonstrated the binding of AP-2α to the core promoter. Furthermore, knock down of endogenous AP-2α downregulated DEK expression, whereas overexpression of AP-2α upregulated DEK expression. Thus, AP-2α is an important transcription factor of DEK expression, which is correlated with the methylation level of the DEK core promoter in HCC.


Assuntos
Carcinoma Hepatocelular/genética , Proteínas Cromossômicas não Histona/genética , Metilação de DNA/genética , Neoplasias Hepáticas/genética , Proteínas Oncogênicas/genética , Fator de Transcrição AP-2/genética , Sítios de Ligação , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/biossíntese , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Proteínas Oncogênicas/biossíntese , Proteínas de Ligação a Poli-ADP-Ribose , Regiões Promotoras Genéticas , Ativação Transcricional/genética
17.
G3 (Bethesda) ; 6(9): 2955-62, 2016 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-27449514

RESUMO

To identify genetic variants underlying changes in phenotypes within and between species, researchers often utilize transgenic animals to compare the function of alleles in different genetic backgrounds. In Drosophila, targeted integration mediated by the ΦC31 integrase allows activity of alternative alleles to be compared at the same genomic location. By using the same insertion site for each transgene, position effects are generally assumed to be controlled for because both alleles are surrounded by the same genomic context. Here, we test this assumption by comparing the activity of tan alleles from two Drosophila species, D. americana and D. novamexicana, at five different genomic locations in D. melanogaster We found that the relative effects of these alleles varied among insertion sites, with no difference in activity observed between them at two sites. One of these sites simply silenced both transgenes, but the other allowed expression of both alleles that was sufficient to rescue a mutant phenotype yet failed to reveal the functional differences between the two alleles. These results suggest that more than one insertion site should be used when comparing the activity of transgenes because failing to do so could cause functional differences between alleles to go undetected.


Assuntos
Alelos , Proteínas Cromossômicas não Histona/genética , Proteínas de Ligação a DNA/genética , Proteínas de Drosophila/genética , Drosophila/genética , Transgenes/genética , Animais , Animais Geneticamente Modificados , Proteínas Cromossômicas não Histona/biossíntese , Proteínas de Ligação a DNA/biossíntese , Proteínas de Drosophila/biossíntese , Genoma de Inseto , Genômica , Integrases/genética , Fenótipo , Especificidade da Espécie
18.
Oncotarget ; 7(29): 45803-45818, 2016 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-27329586

RESUMO

Development of colorectal cancer (CRC) involves sequential transformation of normal mucosal tissues into benign adenomas and then adenomas into malignant tumors. The identification of genes crucial for malignant transformation in colorectal adenomas (CRAs) has been based primarily on cross-sectional observations. In this study, we identified relevant genes using autologous samples. By performing genome-wide SNP genotyping and RNA sequencing analysis of adenocarcinomas, adenomatous polyps, and non-neoplastic colon tissues (referred as tri-part samples) from individual patients, we identified 68 genes with differential copy number alterations and progressively dysregulated expression. Aurora A, SKA3, and DSN1 protein levels were sequentially up-regulated in the samples, and this overexpression was associated with chromosome instability (CIN). Knockdown of SKA3 in CRC cells dramatically reduced cell growth rates and increased apoptosis. Depletion of SKA3 or DSN1 induced G2/M arrest and decreased migration, invasion, and anchorage-independent growth. AURKA and DSN1 are thus critical for chromosome 20q amplification-associated malignant transformation in CRA. Moreover, SKA3 at chromosome 13q was identified as a novel gene involved in promoting malignant transformation. Evaluating the expression of these genes may help identify patients with progressive adenomas, helping to improve treatment.


Assuntos
Adenocarcinoma/patologia , Adenoma/patologia , Transformação Celular Neoplásica/genética , Neoplasias Colorretais/patologia , Adenocarcinoma/genética , Adenoma/genética , Adulto , Idoso , Área Sob a Curva , Aurora Quinase A/biossíntese , Proteínas Cromossômicas não Histona/biossíntese , Neoplasias Colorretais/genética , Progressão da Doença , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Proteínas Associadas aos Microtúbulos/biossíntese , Pessoa de Meia-Idade , Curva ROC , Transcriptoma , Regulação para Cima
19.
Dev Growth Differ ; 58(4): 367-82, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27125315

RESUMO

Some organisms, such as zebrafish, urodele amphibians, and newborn mice, have a capacity for heart regeneration following injury. However, adult mammals fail to regenerate their hearts. To know why newborn mice can regenerate their hearts, we focused on epigenetic factors, which are involved in cell differentiation in many tissues. Baf60c (BRG1/BRM-associated factor 60c), a component of ATP-dependent chromatin-remodeling complexes, has an essential role for cardiomyocyte differentiation at the early heart development. To address the function of Baf60c in postnatal heart homeostasis and regeneration, we examined the detailed expression/localization patterns of Baf60c in both mice and axolotls. In the mouse heart development, Baf60c was highly expressed in the entire heart at the early stages, but gradually downregulated at the postnatal stages. During heart regeneration in neonatal mice and axolotls, Baf60c expression was strongly upregulated after resection. Interestingly, the timing of Baf60c upregulation after resection was consistent with the temporal dynamics of cardiomyocyte proliferation. Moreover, knockdown of Baf60c downregulated proliferation of neonatal mouse cardiomyocytes. These data suggested that Baf60c plays an important role in cardiomyocyte proliferation in heart development and regeneration. This is the first study indicating that Baf60c contributes to the heart regeneration in vertebrates.


Assuntos
Proteínas de Anfíbios/biossíntese , Proteínas Cromossômicas não Histona/biossíntese , Regulação da Expressão Gênica , Coração/fisiologia , Proteínas Musculares/biossíntese , Regeneração/fisiologia , Ambystoma mexicanum , Animais , Proliferação de Células/fisiologia , Camundongos , Camundongos Endogâmicos ICR , Camundongos Transgênicos , Miócitos Cardíacos/metabolismo
20.
PLoS Genet ; 12(3): e1005898, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26938916

RESUMO

BAHD1 is a vertebrate protein that promotes heterochromatin formation and gene repression in association with several epigenetic regulators. However, its physiological roles remain unknown. Here, we demonstrate that ablation of the Bahd1 gene results in hypocholesterolemia, hypoglycemia and decreased body fat in mice. It also causes placental growth restriction with a drop of trophoblast glycogen cells, a reduction of fetal weight and a high neonatal mortality rate. By intersecting transcriptome data from murine Bahd1 knockout (KO) placentas at stages E16.5 and E18.5 of gestation, Bahd1-KO embryonic fibroblasts, and human cells stably expressing BAHD1, we also show that changes in BAHD1 levels alter expression of steroid/lipid metabolism genes. Biochemical analysis of the BAHD1-associated multiprotein complex identifies MIER proteins as novel partners of BAHD1 and suggests that BAHD1-MIER interaction forms a hub for histone deacetylases and methyltransferases, chromatin readers and transcription factors. We further show that overexpression of BAHD1 leads to an increase of MIER1 enrichment on the inactive X chromosome (Xi). In addition, BAHD1 and MIER1/3 repress expression of the steroid hormone receptor genes ESR1 and PGR, both playing important roles in placental development and energy metabolism. Moreover, modulation of BAHD1 expression in HEK293 cells triggers epigenetic changes at the ESR1 locus. Together, these results identify BAHD1 as a core component of a chromatin-repressive complex regulating placental morphogenesis and body fat storage and suggest that its dysfunction may contribute to several human diseases.


Assuntos
Proteínas Cromossômicas não Histona/genética , Proteínas Nucleares/genética , Placentação/genética , Esteroides/metabolismo , Fatores de Transcrição/genética , Animais , Cromatina/genética , Proteínas Cromossômicas não Histona/biossíntese , Proteínas de Ligação a DNA , Receptor alfa de Estrogênio/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Células HEK293 , Humanos , Camundongos , Camundongos Knockout , Proteínas Nucleares/biossíntese , Placenta/metabolismo , Gravidez , Fatores de Transcrição/biossíntese , Transcriptoma/genética
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