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1.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 36(11): 1077-1080, 2019 Nov 10.
Artigo em Chinês | MEDLINE | ID: mdl-31703129

RESUMO

OBJECTIVE: To assess the value of detecting multiple rearrangements of MLL gene in children with acute mononuclear leukemia (AML). METHODS: Eighty six children with AML were analyzed by fluorescence in situ hybridization (FISH), chromosomal karyotyping and multiplex reverse transcription-PCR (RT-PCR). RESULTS: Cross signals were detected by FISH in 26 cases, and 30.2% were detected with MLL gene rearrangements. R-band karyotyping analysis revealed 14 translocations with breakages involving 11q23 and 5 other aberrations, which yielded an overall detection rate of 22.1%. Multiple RT-PCR has detected 12 fusion genes produced by the MLL translocation, which yielded a detection rate of 14.0%. A significant difference was found in the detection rate of the three methods (P< 0.05). CONCLUSION: Combined use of FISH, chromosomal karyotyping and multiplex RT-PCR can improve the detection of MLL gene rearrangements and provide important clues for clinical diagnosis, treatment and prognosis of AML.


Assuntos
Histona-Lisina N-Metiltransferase/genética , Leucemia Mieloide Aguda/genética , Proteína de Leucina Linfoide-Mieloide/genética , Criança , Cromossomos Humanos Par 11 , Rearranjo Gênico , Humanos , Hibridização in Situ Fluorescente , Cariotipagem , Translocação Genética
2.
Rinsho Ketsueki ; 60(9): 1317-1323, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31597858

RESUMO

Acute lymphoblastic leukemia (ALL) in infants under 1 is a rare and dismal disease. It is associated with a unique and specific biology, and 80% of cases harbor a KMT2A (MLL) gene rearrangement (KMT2A-r). In contrast to ALL in older children, with a survival rate of 80% or more, the prognosis of infant ALL is very poor, at 40%. In addition, the unique pharmacodynamics exhibited by infants has historically led to independent therapeutic development either in the U.S., Europe, or Japan. To improve the prognosis of infant ALL, it is necessary to uncover a supplementary novel effective agent to be used in combination with the existing conventional multi-agent chemotherapy. Because of the rarity of the disease, this could be only established by an international study, for which the consensus has already been established through discussions between the U.S., Europe, and Japan. Additionally, severe late effects in survivors are also problematic. Establishing novel treatment strategies to reduce relapse rates, treatment-related toxicities, and critical late effects is strongly encouraged in near future.


Assuntos
Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Europa (Continente) , Rearranjo Gênico , Histona-Lisina N-Metiltransferase/genética , Humanos , Lactente , Japão , Proteína de Leucina Linfoide-Mieloide/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Prognóstico , Taxa de Sobrevida , Estados Unidos
3.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 36(8): 777-780, 2019 Aug 10.
Artigo em Chinês | MEDLINE | ID: mdl-31400126

RESUMO

OBJECTIVE: To assess the value of detecting the rearrangement of mixed lineage leukemia (MLL) gene in children with acute mononuclear leukemia (AML). METHODS: Dual-color fluorescence in situ hybridization (FISH) probe was used to detect MLL gene rearrangement in 68 children with AML by interphase FISH. The results were compared with that of conventional G banding chromosomal analysis. RESULTS: Among the 68 children, 28 were detected by FISH with positive hybridization signals, with a detection rate for MLL gene rearrangement being 41.2%. Twelve (17.6%) reciprocal translocations and interruption of 11q23 were detected by G banding analysis. The difference in the detection rates between the two methods was statistically significant (P< 0.05). CONCLUSION: The sensitivity of FISH assay for MLL gene rearrangement was significantly higher than that of G banding chromosomal karyotyping. Combined use of both methods for children with AML can improve the detection rate of MLL gene rearrangements and provide crucial clues for clinical diagnosis, treatment and prognosis.


Assuntos
Histona-Lisina N-Metiltransferase/genética , Leucemia Monocítica Aguda/genética , Proteína de Leucina Linfoide-Mieloide/genética , Criança , Cromossomos Humanos Par 11 , Rearranjo Gênico , Humanos , Hibridização in Situ Fluorescente , Translocação Genética
4.
Cytogenet Genome Res ; 158(4): 205-212, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31434093

RESUMO

EHMT2 (euchromatic histone lysine methyltransferase 2), a histone methyltransferase, has been shown to be involved in multiple human cancers. In this study, we determined mRNA and protein expression of EHMT2 in cervical cancer cells and normal cervical epithelial cells. EHMT2 was inhibited with short hairpin RNA (shEHMT2) in cervical cancer cells. Cell viability, colony proliferation, apoptosis, adhesion, and invasion assays and Western blot were performed to assess the function of EHMT2. As a result, EHMT2 was upregulated in human cervical cancer cells compared to normal cervical epithelial cells. Suppression of EHMT2 expression impairs cell proliferation and induces apoptosis. Furthermore, EHMT2 silencing inhibited cell adhesion and invasion. Finally, knockdown of EHMT2 resulted in a reduction of the expression of the tumorigenic proteins Bcl-2, Mcl-1, and Survivin and in an increase in the expression of the anti-malignant protein E-cadherin. In conclusion, our data suggest that EHMT2 plays a key role in cell proliferation and metastatic capacity in cervical cancer cells and could serve as a potential therapeutic target.


Assuntos
Inativação Gênica , Antígenos de Histocompatibilidade/genética , Histona-Lisina N-Metiltransferase/deficiência , Histona-Lisina N-Metiltransferase/genética , Invasividade Neoplásica/genética , Invasividade Neoplásica/prevenção & controle , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/patologia , Apoptose/genética , Caderinas/biossíntese , Adesão Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Antígenos de Histocompatibilidade/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Invasividade Neoplásica/patologia , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Metástase Neoplásica/prevenção & controle , Neoplasias do Colo do Útero/genética
6.
Nat Chem Biol ; 15(8): 822-829, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31285596

RESUMO

Here, we report the fragment-based discovery of BI-9321, a potent, selective and cellular active antagonist of the NSD3-PWWP1 domain. The human NSD3 protein is encoded by the WHSC1L1 gene located in the 8p11-p12 amplicon, frequently amplified in breast and squamous lung cancer. Recently, it was demonstrated that the PWWP1 domain of NSD3 is required for the viability of acute myeloid leukemia cells. To further elucidate the relevance of NSD3 in cancer biology, we developed a chemical probe, BI-9321, targeting the methyl-lysine binding site of the PWWP1 domain with sub-micromolar in vitro activity and cellular target engagement at 1 µM. As a single agent, BI-9321 downregulates Myc messenger RNA expression and reduces proliferation in MOLM-13 cells. This first-in-class chemical probe BI-9321, together with the negative control BI-9466, will greatly facilitate the elucidation of the underexplored biological function of PWWP domains.


Assuntos
Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Proteínas Nucleares/antagonistas & inibidores , Sistemas CRISPR-Cas , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Regulação da Expressão Gênica/efeitos dos fármacos , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Domínios Proteicos , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo
7.
Nat Commun ; 10(1): 2854, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253781

RESUMO

SETD1A, a Set1/COMPASS family member maintaining histone-H3-lysine-4 (H3K4) methylation on transcriptionally active promoters, is overexpressed in breast cancer. Here, we show that SETD1A supports mitotic processes and consequentially, its knockdown induces senescence. SETD1A, through promoter H3K4 methylation, regulates several genes orchestrating mitosis and DNA-damage responses, and its depletion causes chromosome misalignment and segregation defects. Cell cycle arrest in SETD1A knockdown senescent cells is independent of mutations in p53, RB and p16, known senescence mediators; instead, it is sustained through transcriptional suppression of SKP2, which degrades p27 and p21. Rare cells escaping senescence by restoring SKP2 expression display genomic instability. In > 200 cancer cell lines and in primary circulating tumor cells, SETD1A expression correlates with genes promoting mitosis and cell cycle suggesting a broad role in suppressing senescence induced by aberrant mitosis. Thus, SETD1A is essential to maintain mitosis and proliferation and its suppression unleashes the tumor suppressive effects of senescence.


Assuntos
Senescência Celular/fisiologia , Regulação da Expressão Gênica/fisiologia , Histona-Lisina N-Metiltransferase/metabolismo , Mitose/fisiologia , Linhagem Celular Tumoral , Histona-Lisina N-Metiltransferase/genética , Histonas , Humanos , Metilação , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
8.
Anticancer Res ; 39(6): 2729-2737, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31177108

RESUMO

BACKGROUND/AIM: Salivary adenoid cystic carcinoma (SACC) is the most common malignancy of the salivary gland with a poor prognosis and survival. The present study aimed to investigate the role of histone methyltransferase WHSC1 in SACC. MATERIALS AND METHODS: Human SACC specimens were evaluated for WHSC1 expression by RT-PCR and immunohistochemistry. The effects of WHSC1 knockdown on SACC cells proliferation, cell cycle, clone and tumorsphere formation, and apoptosis as well as on the expression of related genes were examined. A xenograft mouse model of SACC was used to evaluate the in vivo effects of WHSC1 knockdown on SACC tumorigenesis. RESULTS: WHSC1 expression was up-regulated in human SACC tissues (p<0.01). WHSC1 knockdown in SACC cells significantly inhibited cell proliferation, clone and tumorsphere formation (p<0.05). Cell distribution at the S and G2/M phases was significantly reduced by WHSC1 knockdown (p<0.05). WHSC1 knockdown significantly increased apoptosis of SACC cells (p<0.05). c-Myc, survivin, Bcl-2 and cyclin B1 genes were significantly down-regulated by WHSC1 knockdown cells (p<0.05). WHSC1 knockdown significantly reduced H3K36me2 modification of the MYC gene promoter in SACC cells and tumorigenesis of SACC cells in vivo (p<0.05). CONCLUSION: Knockdown of WHSC1 inhibited cell proliferation, induced apoptosis and affected tumorigenesis in SACC.


Assuntos
Carcinoma Adenoide Cístico/patologia , Técnicas de Silenciamento de Genes/métodos , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Neoplasias das Glândulas Salivares/patologia , Regulação para Cima , Animais , Apoptose , Carcinoma Adenoide Cístico/genética , Carcinoma Adenoide Cístico/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Humanos , Masculino , Camundongos , Transplante de Neoplasias , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-myc/genética , Neoplasias das Glândulas Salivares/genética , Neoplasias das Glândulas Salivares/metabolismo , Transdução de Sinais
9.
Cancer Sci ; 110(8): 2600-2606, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31218784

RESUMO

The t(11;14)/CCND1-IGH, t(4;14)/NSD2(MMSET)-IGH, and t(14;16)/IGH-MAF gene rearrangements detected by fluorescence in situ hybridization (FISH) are used for risk stratification in patients with multiple myeloma (MM). Compared with conventional FISH techniques using fresh cells, immunohistochemistry (IHC) is much more cost- and time-efficient, and can be readily applied to routinely prepared formalin-fixed, paraffin-embedded (FFPE) materials. In this study, we performed tissue FISH and IHC employing FFPE specimens, and examined the usefulness of IHC as a tool for detecting CCND1, NSD2, and MAF gene rearrangements. CD138 signals were used to identify plasma cells in tissue FISH and IHC analyses. With cohort 1 (n = 70), we performed tissue FISH and subsequently IHC, and determined IHC cut-off points. In this cohort, the sensitivity and specificity for the 3 molecules were ≥.90 and ≥.96, respectively. With cohort 2, using MM cases with an unknown gene status (n = 120), we performed IHC, and the gene status was estimated using the cut-off points determined with cohort 1. The subsequent FISH analysis showed that the sensitivity and specificity for the 3 molecules were ≥.92 and ≥.98, respectively. CCND1, NSD2, and MAF gene rearrangements were estimated accurately by IHC, suggesting that conventional FISH assays can be replaced by IHC.


Assuntos
Ciclina D1/genética , Rearranjo Gênico/genética , Histona-Lisina N-Metiltransferase/genética , Mieloma Múltiplo/genética , Proteínas Proto-Oncogênicas c-maf/genética , Proteínas Repressoras/genética , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Humanos , Imuno-Histoquímica/métodos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade
10.
BMC Cancer ; 19(1): 455, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31092221

RESUMO

BACKGROUND: Wolf-Hirschhorn syndrome candidate gene-1 (WHSC1), a histone methyltransferase, has been found to be upregulated and its expression to be correlated with expression of enhancer of zeste homolog 2 (EZH2) in several cancers. In this study, we evaluated the role of WHSC1 and its therapeutic significance in ovarian clear cell carcinoma (OCCC). METHODS: First, we analyzed WHSC1 expression by quantitative PCR and immunohistochemistry using 23 clinical OCCC specimens. Second, the involvement of WHSC1 in OCCC cell proliferation was evaluated by MTT assays after siRNA-mediated WHSC1 knockdown. We also performed flow cytometry (FACS) to address the effect of WHSC1 on cell cycle. To examine the functional relationship between EZH2 and WHSC1, we knocked down EZH2 using siRNAs and checked the expression levels of WHSC1 and its histone mark H3K36m2 in OCCC cell lines. Finally, we checked WHSC1 expression after treatment with the selective inhibitor, GSK126. RESULTS: Both quantitative PCR and immunohistochemical analysis revealed that WHSC1 was significantly overexpressed in OCCC tissues compared with that in normal ovarian tissues. MTT assay revealed that knockdown of WHSC1 suppressed cell proliferation, and H3K36me2 levels were found to be decreased in immunoblotting. FACS revealed that WHSC1 knockdown affected the cell cycle. We also confirmed that WHSC1 expression was suppressed by EZH2 knockdown or inhibition, indicating that EZH2 is upstream of WHSC1 in OCCC cells. CONCLUSIONS: WHSC1 overexpression induced cell growth and its expression is, at least in part, regulated by EZH2. Further functional analysis will reveal whether WHSC1 is a promising therapeutic target for OCCC.


Assuntos
Adenocarcinoma de Células Claras/genética , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Histona-Lisina N-Metiltransferase/genética , Neoplasias Ovarianas/genética , Proteínas Repressoras/genética , Adenocarcinoma de Células Claras/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Humanos , Neoplasias Ovarianas/metabolismo , Proteínas Repressoras/metabolismo , Regulação para Cima
11.
Cell Prolif ; 52(4): e12611, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31054182

RESUMO

OBJECTIVES: Epigenetic modifiers were important players in the development of haematological malignancies and sensitivity to therapy. Mutations of SET domain-containing 2 (SETD2), a methyltransferase that catalyses the trimethylation of histone 3 on lysine 36 (H3K36me3), were found in various myeloid malignancies. However, the detailed mechanisms through which SETD2 confers chronic myeloid leukaemia progression and resistance to therapy targeting on BCR-ABL remain unclear. MATERIALS AND METHODS: The level of SETD2 in imatinib-sensitive and imatinib-resistant chronic myeloid leukaemia (CML) cells was examined by immunoblotting and quantitative real-time PCR. We analysed CD34+ CD38- leukaemic stem cells by flow cytometry and colony formation assays upon SETD2 knockdown or overexpression. The impact of SETD2 expression alterations or small-molecule inhibitor JIB-04 targeting H3K36me3 loss on imatinib sensitivity was assessed by IC50, cell apoptosis and proliferation assays. Finally, RNA sequencing and ChIP-quantitative PCR were performed to verify putative downstream targets. RESULTS: SETD2 was found to act as a tumour suppressor in CML. The novel oncogenic targets MYCN and ERG were shown to be the direct downstream targets of SETD2, where their overexpression induced by SETD2 knockdown caused imatinib insensitivity and leukaemic stem cell enrichment in CML cell lines. Treatment with JIB-04, an inhibitor that restores H3K36me3 levels through blockade of its demethylation, successfully improved the cell imatinib sensitivity and enhanced the chemotherapeutic effect. CONCLUSIONS: Our study not only emphasizes the regulatory mechanism of SETD2 in CML, but also provides promising therapeutic strategies for overcoming the imatinib resistance in patients with CML.


Assuntos
Regulação para Baixo/genética , Resistencia a Medicamentos Antineoplásicos/genética , Histona Metiltransferases/genética , Histona-Lisina N-Metiltransferase/genética , Mesilato de Imatinib/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Aminopiridinas/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Regulação para Baixo/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Fusão bcr-abl/genética , Humanos , Hidrazonas/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia
12.
Nat Genet ; 51(5): 844-856, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31040401

RESUMO

The oocyte epigenome plays critical roles in mammalian gametogenesis and embryogenesis. Yet, how it is established remains elusive. Here, we report that histone-lysine N-methyltransferase SETD2, an H3K36me3 methyltransferase, is a crucial regulator of the mouse oocyte epigenome. Deficiency in Setd2 leads to extensive alterations of the oocyte epigenome, including the loss of H3K36me3, failure in establishing the correct DNA methylome, invasion of H3K4me3 and H3K27me3 into former H3K36me3 territories and aberrant acquisition of H3K4me3 at imprinting control regions instead of DNA methylation. Importantly, maternal depletion of SETD2 results in oocyte maturation defects and subsequent one-cell arrest after fertilization. The preimplantation arrest is mainly due to a maternal cytosolic defect, since it can be largely rescued by normal oocyte cytosol. However, chromatin defects, including aberrant imprinting, persist in these embryos, leading to embryonic lethality after implantation. Thus, these data identify SETD2 as a crucial player in establishing the maternal epigenome that in turn controls embryonic development.


Assuntos
Desenvolvimento Embrionário/genética , Epigênese Genética , Impressão Genômica , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Animais , DNA (Citosina-5-)-Metiltransferases/deficiência , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA , Feminino , Código das Histonas/genética , Histona-Lisina N-Metiltransferase/deficiência , Histonas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Modelos Genéticos , Oócitos/metabolismo , Oogênese/genética , Gravidez
13.
BMC Genomics ; 20(1): 350, 2019 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-31068130

RESUMO

BACKGROUND: Histone H3 lysine 4 tri-methylation (H3K4me3) and histone H3 lysine 9 tri-methylation (H3K9me3) are widely perceived to be opposing and often mutually exclusive chromatin modifications. However, both are needed for certain light-activated genes in Neurospora crassa (Neurospora), including frequency (frq) and vivid (vvd). Except for these 2 loci, little is known about how H3K4me3 and H3K9me3 impact and contribute to light-regulated gene expression. RESULTS: In this report, we performed a multi-dimensional genomic analysis to understand the role of H3K4me3 and H3K9me3 using the Neurospora light response as the system. RNA-seq on strains lacking H3 lysine 4 methyltransferase (KMT2/SET-1) and histone H3 lysine 9 methyltransferase (KMT1/DIM-5) revealed some light-activated genes had altered expression, but the light response was largely intact. Comparing these 2 mutants to wild-type (WT), we found that roughly equal numbers of genes showed elevated and reduced expression in the dark and the light making the environmental stimulus somewhat ancillary to the genome-wide effects. ChIP-seq experiments revealed H3K4me3 and H3K9me3 had only minor changes in response to light in WT, but there were notable alterations in H3K4me3 in Δkmt1/Δdim-5 and H3K9me3 in Δkmt2/Δset-1 indicating crosstalk and redistribution between the modifications. Integrated analysis of the RNA-seq and ChIP-seq highlighted context-dependent roles for KMT2/SET1 and KMT1/DIM-5 as either co-activators or co-repressors with some overlap as co-regulators. At a small subset of loci, H3K4 methylation is required for H3K9me3-mediated facultative heterochromatin including, the central clock gene frequency (frq). Finally, we used sequential ChIP (re-ChIP) experiment to confirm Neurospora contains K4/K9 bivalent domains. CONCLUSIONS: Collectively, these data indicate there are obfuscated regulatory roles for H3K4 methylation and H3K9 methylation depending on genome location with some minor overlap and co-dependency.


Assuntos
Proteínas Fúngicas/metabolismo , Heterocromatina , Histona-Lisina N-Metiltransferase/metabolismo , Neurospora crassa/genética , Processamento de Proteína Pós-Traducional , Metilação de DNA , Proteínas Fúngicas/genética , Regulação Enzimológica da Expressão Gênica , Regulação Fúngica da Expressão Gênica , Histona-Lisina N-Metiltransferase/genética , Histonas/metabolismo , Luz , Neurospora crassa/enzimologia
14.
Cytogenet Genome Res ; 157(4): 231-238, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30933949

RESUMO

Constitutional complex chromosomal rearrangements (CCRs) are rare events that typically involve 2 or more chromosomes with at least 3 breakpoints and can result in normal or abnormal phenotypes depending on whether they disturb the euchromatic neighborhood. Here, we report an unusual balanced CCR involving chromosomes 1, 9, and 10 that causes an unbalanced karyotype in a severely affected toddler. The CCR was initially reported as a maternal 2-way translocation but was reclassified as a 3-way translocation after a microarray analysis of the propositus revealed the involvement of another chromosome not identified by G-banding in his phenotypically normal mother. FISH assays on maternal metaphase cells confirmed that the 1qter region of der(1) was translocated to der(10), whereas the 10qter segment was translocated to der(9), which in turn donated a segment to der(1). Subsequently, this CCR was also identified in her phenotypically normal father (the patient's grandfather). Thus, the patient inherited the previously unreported pathogenic combination of der(1) with a loss of 1q43→qter (including AKT3, ZBTB18, HNRNPU, and SMYD3) and der(9) with a gain of 10q25.2→qter (including FGFR2), leading to a compound phenotype with key features of the 1q43→qter deletion and distal 10q trisomy syndromes. Our observations suggest that the loss of SMYD3 accounts for cardiac defects in a subset of patients. Moreover, due to recurrent miscarriages in this family, our findings allowed improved genetic counseling.


Assuntos
Anormalidades Múltiplas/genética , Cromossomos Humanos Par 10/genética , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 9/genética , Anormalidades Múltiplas/diagnóstico por imagem , Pré-Escolar , Hibridização Genômica Comparativa , Aconselhamento Genético , Histona-Lisina N-Metiltransferase/genética , Humanos , Hibridização in Situ Fluorescente/métodos , Masculino , Tomografia Computadorizada por Raios X , Translocação Genética
15.
PLoS Genet ; 15(4): e1008093, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-31009462

RESUMO

Chromosome and genome stability are important for normal cell function as instability often correlates with disease and dysfunction of DNA repair mechanisms. Many organisms maintain supernumerary or accessory chromosomes that deviate from standard chromosomes. The pathogenic fungus Zymoseptoria tritici has as many as eight accessory chromosomes, which are highly unstable during meiosis and mitosis, transcriptionally repressed, show enrichment of repetitive elements, and enrichment with heterochromatic histone methylation marks, e.g., trimethylation of H3 lysine 9 or lysine 27 (H3K9me3, H3K27me3). To elucidate the role of heterochromatin on genome stability in Z. tritici, we deleted the genes encoding the methyltransferases responsible for H3K9me3 and H3K27me3, kmt1 and kmt6, respectively, and generated a double mutant. We combined experimental evolution and genomic analyses to determine the impact of these deletions on chromosome and genome stability, both in vitro and in planta. We used whole genome sequencing, ChIP-seq, and RNA-seq to compare changes in genome and chromatin structure, and differences in gene expression between mutant and wildtype strains. Analyses of genome and ChIP-seq data in H3K9me3-deficient strains revealed dramatic chromatin reorganization, where H3K27me3 is mostly relocalized into regions that are enriched with H3K9me3 in wild type. Many genome rearrangements and formation of new chromosomes were found in the absence of H3K9me3, accompanied by activation of transposable elements. In stark contrast, loss of H3K27me3 actually increased the stability of accessory chromosomes under normal growth conditions in vitro, even without large scale changes in gene activity. We conclude that H3K9me3 is important for the maintenance of genome stability because it disallows H3K27me3 in regions considered constitutive heterochromatin. In this system, H3K27me3 reduces the overall stability of accessory chromosomes, generating a "metastable" state for these quasi-essential regions of the genome.


Assuntos
Instabilidade Genômica , Histonas/metabolismo , Lisina/metabolismo , Ascomicetos/genética , Ascomicetos/metabolismo , Cromossomos Fúngicos , Deleção de Genes , Heterocromatina/genética , Heterocromatina/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histonas/química , Metilação , Sequências Repetitivas de Ácido Nucleico , Ativação Transcricional
16.
Gene ; 705: 22-35, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31005612

RESUMO

Mixed-lineage leukaemia 1 (MLL1) enzyme plays major role in regulating genes associated with vertebrate development. Cell physiology and homeostasis is regulated by microRNAs in diverse microenvironment. In this investigation we have identified conserved miR-193a target sites within the 3'-UTR of MLL1 gene transcript. Utilizing wild type and mutated 3'-UTR constructs and luciferase reporter assays we have clearly demonstrated that miR-193a directly targets the 3'-UTR region of the MLL1 mRNA. Ectopic expression of miR-193a modulated global H3K4 mono-, di- and tri-methylation levels and affects the expression of CAV1, a gene which is specifically modulated by H3K4me3. To determine the implications of this in vitro finding in aberrant physiological conditions we analyzed prostate cancer tissue samples. In this context miR-193a RNA was undetectable and MLL1 was highly expressed with concomitantly high levels of H3K4me, H3K4me2, and H3K4me3 enrichment in the promoters of MLL1 responsive genes. Finally, we showed that prolonged ectopic expression of miR-193a inhibits growth and cell migration, and induces apoptosis. Thus, while our study unveils amplitude of the epigenome, including miRnome it establishes that; (i) miR-193a directly target MLL1 mRNA, (ii) miR-193a impair MLL1 protein production, (iii) miR-193a reduces the overall methylation marks of the genome.


Assuntos
Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , MicroRNAs/genética , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/metabolismo , Neoplasias da Próstata/genética , Regiões 3' não Traduzidas , Caveolina 1/genética , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Cromatina/metabolismo , Regulação para Baixo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Metilação , Neoplasias da Próstata/metabolismo
17.
Nat Commun ; 10(1): 1897, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015486

RESUMO

The cellular decision regarding whether to undergo proliferation or death is made at the restriction (R)-point, which is disrupted in nearly all tumors. The identity of the molecular mechanisms that govern the R-point decision is one of the fundamental issues in cell biology. We found that early after mitogenic stimulation, RUNX3 binds to its target loci, where it opens chromatin structure by sequential recruitment of Trithorax group proteins and cell-cycle regulators to drive cells to the R-point. Soon after, RUNX3 closes these loci by recruiting Polycomb repressor complexes, causing the cell to pass through the R-point toward S phase. If the RAS signal is constitutively activated, RUNX3 inhibits cell cycle progression by maintaining R-point-associated genes in an open structure. Our results identify RUNX3 as a pioneer factor for the R-point and reveal the molecular mechanisms by which appropriate chromatin modifiers are selectively recruited to target loci for appropriate R-point decisions.


Assuntos
Pontos de Checagem do Ciclo Celular/genética , Cromatina/química , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Animais , Butadienos/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cromatina/efeitos dos fármacos , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Subunidade alfa 3 de Fator de Ligação ao Core/antagonistas & inibidores , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 4 Dependente de Ciclina/genética , Quinase 4 Dependente de Ciclina/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Células HEK293 , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Imidazóis/farmacologia , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 1/genética , MAP Quinase Quinase 1/metabolismo , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/metabolismo , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/metabolismo , Nitrilos/farmacologia , Piperazinas/farmacologia , Proteínas do Grupo Polycomb/genética , Proteínas do Grupo Polycomb/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Proteínas ras/genética , Proteínas ras/metabolismo
18.
Cell Prolif ; 52(4): e12621, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31012192

RESUMO

OBJECTIVES: Long non-coding RNAs (LncRNAs) play an important role in hepatocellular carcinoma development, however, as a crucial driver of hepatocellular carcinoma (HCC) metastasis, their functions in tumour metastasis remain largely unknown. MATERIALS AND METHODS: The lncRNA TRERNA1 expression levels were detected in HCC by quantitative real-time PCR (qPCR). The function of TRERNA1 was examined by wound-healing assays, transwell assays and tail vein injection experiments. The potential regulatory mechanisms of TRERNA1 on its target genes were explored by ChIP, RIP, IP and WB assays. RESULTS: Elevated TRERNA1 levels promoted HCC cell migration and invasion in vitro and in vivo. TRERNA1 recruited EHMT2 to dimethylate H3K9 in the CDH1 promoter region. Furthermore, EHMT2 bound to SNAI1 to suppress CDH1 expression in HCC cells. After inhibiting TRERNA1, the expression level of CDH1 was restored and was involved in the regulation of the EHMT2/SNAI1 complex. The level of TRERNA1 was positively correlated with tumour metastasis and was negatively correlated with the expression of CDH1 in HCC tissues. CONCLUSIONS: For the first time, the current study reveals that TRERNA1 promotes cell metastasis and the invasion of HCC via the recruitment of EHMT2 and/or the EHMT2/SNAI1 complex to suppress CDH1. These data identify a novel mechanism that regulates TRERNA1 in metastatic HCC and provides a potential targeted therapy for HCC patients.


Assuntos
Antígenos CD/genética , Caderinas/genética , Carcinoma Hepatocelular/genética , Antígenos de Histocompatibilidade/genética , Histona-Lisina N-Metiltransferase/genética , Neoplasias Hepáticas/genética , Regiões Promotoras Genéticas/genética , RNA Longo não Codificante/genética , Fatores de Transcrição da Família Snail/genética , Animais , Carcinoma Hepatocelular/fisiopatologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica/genética , Células Hep G2 , Humanos , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Interferência de RNA/fisiologia
19.
Mol Cell ; 74(5): 1010-1019.e6, 2019 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-30981630

RESUMO

The essential histone H3 lysine 79 methyltransferase Dot1L regulates transcription and genomic stability and is deregulated in leukemia. The activity of Dot1L is stimulated by mono-ubiquitination of histone H2B on lysine 120 (H2BK120Ub); however, the detailed mechanism is not understood. We report cryo-EM structures of human Dot1L bound to (1) H2BK120Ub and (2) unmodified nucleosome substrates at 3.5 Å and 4.9 Å, respectively. Comparison of both structures, complemented with biochemical experiments, provides critical insights into the mechanism of Dot1L stimulation by H2BK120Ub. Both structures show Dot1L binding to the same extended surface of the histone octamer. In yeast, this surface is used by silencing proteins involved in heterochromatin formation, explaining the mechanism of their competition with Dot1. These results provide a strong foundation for understanding conserved crosstalk between histone modifications found at actively transcribed genes and offer a general model of how ubiquitin might regulate the activity of chromatin enzymes.


Assuntos
Histona-Lisina N-Metiltransferase/química , Histonas/química , Lisina/química , Conformação Proteica , Sítios de Ligação , Microscopia Crioeletrônica , Genoma Humano/genética , Instabilidade Genômica/genética , Heterocromatina/química , Heterocromatina/genética , Histona-Lisina N-Metiltransferase/genética , Histonas/genética , Humanos , Leucemia/genética , Lisina/genética , Proteínas Nucleares/química , Proteínas Nucleares/genética , Nucleossomos/química , Nucleossomos/genética , Ligação Proteica , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Transcrição Genética , Ubiquitinação/genética
20.
Cytogenet Genome Res ; 157(4): 213-219, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30974445

RESUMO

Patients with childhood acute myeloid leukemia (AML) with complex karyotypes (CKs) have a dismal outcome. However, for patients with a KMT2A rearrangement (KMT2A-r), the prognosis appears to depend on the fusion partner gene rather than the karyotype structure. Thus, a precise characterization of KMT2A-r and the fusion partner genes, especially in CKs, is of interest for managing AML. We describe the clinical and molecular features of a child who presented with a large abdominal mass, AML, and a new CK, involving chromosomes 11, 16, and 19 leading to a KMT2A-MLLT1 fusion and 2 extra copies of the ELL gene, thus resulting in the concurrent overexpression of MLLT1 and ELL. Molecular cytogenetic studies defined the karyotype as 47,XY,der(11)t(11;16)(q23.3;p11.2),der(16)t(16;19)(p11.2;p13.3),der(19)t(11;19)(q23.3;p13.3),+der(19)t(16;19)(16pter→p11.2::19p13.3→19q11::19p11→19p13.3::16p11.2→16pter). Array CGH revealed a gain of 30.5 Mb in the 16p13.3p11.2 region and a gain of 18.1 Mb in the 19p13.3p12 region. LDI-PCR demonstrated the KMT2A-MLLT1 fusion. Reverse sequence analysis showed that the MLLT1 gene was fused to the 16p11.2 region. RT-qPCR quantification revealed that ELL and MLLT1 were overexpressed (4- and 10-fold, respectively). In summary, this is a pediatric case of AML presenting a novel complex t(11;16;19) variant with overexpression of ELL and MLLT1.


Assuntos
Histona-Lisina N-Metiltransferase/genética , Leucemia Mieloide Aguda/genética , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Fatores de Transcrição/genética , Fatores de Elongação da Transcrição/genética , Translocação Genética , Criança , Cromossomos Humanos Par 11/genética , Cromossomos Humanos Par 16/genética , Cromossomos Humanos Par 19/genética , Humanos , Cariótipo , Masculino , Proteínas de Fusão Oncogênica/genética , Regulação para Cima
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