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1.
Cell ; 143(3): 367-78, 2010 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-21029860

RESUMO

ATRX is an X-linked gene of the SWI/SNF family, mutations in which cause syndromal mental retardation and downregulation of α-globin expression. Here we show that ATRX binds to tandem repeat (TR) sequences in both telomeres and euchromatin. Genes associated with these TRs can be dysregulated when ATRX is mutated, and the change in expression is determined by the size of the TR, producing skewed allelic expression. This reveals the characteristics of the affected genes, explains the variable phenotypes seen with identical ATRX mutations, and illustrates a new mechanism underlying variable penetrance. Many of the TRs are G rich and predicted to form non-B DNA structures (including G-quadruplex) in vivo. We show that ATRX binds G-quadruplex structures in vitro, suggesting a mechanism by which ATRX may play a role in various nuclear processes and how this is perturbed when ATRX is mutated.


Assuntos
DNA Helicases/metabolismo , Proteínas Nucleares/metabolismo , Animais , Células Cultivadas , Imunoprecipitação da Cromatina , Cromossomos de Mamíferos/metabolismo , Ilhas de CpG , DNA Helicases/genética , DNA Ribossômico/metabolismo , Quadruplex G , Expressão Gênica , Estudo de Associação Genômica Ampla , Histonas/metabolismo , Humanos , Camundongos , Repetições Minissatélites , Mutação , Proteínas Nucleares/genética , Telômero/metabolismo , Proteína Nuclear Ligada ao X
2.
Nucleic Acids Res ; 50(8): 4500-4514, 2022 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-35451487

RESUMO

Histone H3.3 is an H3 variant which differs from the canonical H3.1/2 at four residues, including a serine residue at position 31 which is evolutionarily conserved. The H3.3 S31 residue is phosphorylated (H3.3 S31Ph) at heterochromatin regions including telomeres and pericentric repeats. However, the role of H3.3 S31Ph in these regions remains unknown. In this study, we find that H3.3 S31Ph regulates heterochromatin accessibility at telomeres during replication through regulation of H3K9/K36 histone demethylase KDM4B. In mouse embryonic stem (ES) cells, substitution of S31 with an alanine residue (H3.3 A31 -phosphorylation null mutant) results in increased KDM4B activity that removes H3K9me3 from telomeres. In contrast, substitution with a glutamic acid (H3.3 E31, mimics S31 phosphorylation) inhibits KDM4B, leading to increased H3K9me3 and DNA damage at telomeres. H3.3 E31 expression also increases damage at other heterochromatin regions including the pericentric heterochromatin and Y chromosome-specific satellite DNA repeats. We propose that H3.3 S31Ph regulation of KDM4B is required to control heterochromatin accessibility of repetitive DNA and preserve chromatin integrity.


Assuntos
Heterocromatina , Histonas , Animais , Camundongos , Histonas/genética , Histonas/metabolismo , Heterocromatina/genética , Histona Desmetilases/metabolismo , Fosforilação , Montagem e Desmontagem da Cromatina
3.
Mol Ther ; 29(9): 2841-2853, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-33940155

RESUMO

A primary challenge in lentiviral gene therapy of ß-hemoglobinopathies is to maintain low vector copy numbers to avoid genotoxicity while being reliably therapeutic for all genotypes. We designed a high-titer lentiviral vector, LVß-shα2, that allows coordinated expression of the therapeutic ßA-T87Q-globin gene and of an intron-embedded miR-30-based short hairpin RNA (shRNA) selectively targeting the α2-globin mRNA. Our approach was guided by the knowledge that moderate reduction of α-globin chain synthesis ameliorates disease severity in ß-thalassemia. We demonstrate that LVß-shα2 reduces α2-globin mRNA expression in erythroid cells while keeping α1-globin mRNA levels unchanged and ßA-T87Q-globin gene expression identical to the parent vector. Compared with the first ßA-T87Q-globin lentiviral vector that has received conditional marketing authorization, BB305, LVß-shα2 shows 1.7-fold greater potency to improve α/ß ratios. It may thus result in greater therapeutic efficacy and reliability for the most severe types of ß-thalassemia and provide an improved benefit/risk ratio regardless of the ß-thalassemia genotype.


Assuntos
Vetores Genéticos/administração & dosagem , RNA Interferente Pequeno/genética , alfa-Globinas/genética , Globinas beta/genética , Talassemia beta/genética , Linhagem Celular , Células Cultivadas , Regulação para Baixo , Células Eritroides/citologia , Células Eritroides/metabolismo , Genótipo , Humanos , Células K562 , Lentivirus/genética , Lentivirus/fisiologia , MicroRNAs/antagonistas & inibidores , Cultura Primária de Células , Carga Viral , Talassemia beta/terapia
4.
Proc Natl Acad Sci U S A ; 115(18): 4737-4742, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29669917

RESUMO

ATRX (alpha thalassemia/mental retardation X-linked) complexes with DAXX to deposit histone variant H3.3 into repetitive heterochromatin. Recent genome sequencing studies in cancers have revealed mutations in ATRX and their association with ALT (alternative lengthening of telomeres) activation. Here we report depletion of ATRX in mouse ES cells leads to selective loss in ribosomal RNA gene (rDNA) copy number. Supporting this, ATRX-mutated human ALT-positive tumors also show a substantially lower rDNA copy than ALT-negative tumors. Further investigation shows that the rDNA copy loss and repeat instability are caused by a disruption in H3.3 deposition and thus a failure in heterochromatin formation at rDNA repeats in the absence of ATRX. We also find that ATRX-depleted cells are reduced in ribosomal RNA transcription output and show increased sensitivity to RNA polymerase I (Pol I) transcription inhibitor CX5461. In addition, human ALT-positive cancer cell lines are also more sensitive to CX5461 treatment. Our study provides insights into the contribution of ATRX loss of function to tumorigenesis through the loss of rDNA stability and suggests the therapeutic potential of targeting Pol I transcription in ALT cancers.


Assuntos
DNA de Neoplasias/metabolismo , DNA Ribossômico/metabolismo , Dosagem de Genes , Mutação , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Proteína Nuclear Ligada ao X/metabolismo , Benzotiazóis/farmacologia , Linhagem Celular Tumoral , DNA de Neoplasias/genética , DNA Ribossômico/genética , Instabilidade Genômica , Humanos , Naftiridinas/farmacologia , Proteínas de Neoplasias/genética , Neoplasias/genética , Neoplasias/patologia , RNA Polimerase I/antagonistas & inibidores , RNA Polimerase I/genética , RNA Polimerase I/metabolismo , Transcrição Gênica/efeitos dos fármacos , Transcrição Gênica/genética , Proteína Nuclear Ligada ao X/genética
5.
Nucleic Acids Res ; 44(4): 1496-501, 2016 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-26773061

RESUMO

A number of studies have demonstrated that various components of the ATRX/DAXX/Histone H3.3 complex are important for heterochromatin silencing at multiple genomic regions. We provide an overview of the individual components (ATRX, DAXX and/or H3.3) tested in each study and propose a model where the ATRX/DAXX chaperone complex deposits H3.3 to maintain the H3K9me3 modification at heterochromatin throughout the genome.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , DNA Helicases/genética , Heterocromatina/genética , Histonas/genética , Proteínas Nucleares/genética , Montagem e Desmontagem da Cromatina/genética , Proteínas Correpressoras , Genoma Humano , Histona-Lisina N-Metiltransferase/genética , Humanos , Chaperonas Moleculares/genética , Complexos Multiproteicos/genética , Proteína Nuclear Ligada ao X
6.
Cell Mol Life Sci ; 73(9): 1871-9, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26883803

RESUMO

Imprinted genes are an exceptional cluster of genes which are expressed in a parent-of-origin dependent fashion. This allele-specific expression is dependent on differential DNA methylation which is established in the parental germlines in a sex-specific manner. The DNA methylation imprint is accompanied by heterochromatin modifications which must be continuously maintained through development. This review summarises the factors which are important for protecting the epigenetic modifications at imprinted differentially methylated regions (DMRs), including PGC7, ZFP57 and the ATRX/Daxx/H3.3 complex. We discuss how these factors maintain heterochromatin silencing, not only at imprinted DMRs, but also other heterochromatic regions in the genome.


Assuntos
Inativação Gênica , Impressão Genômica , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Cromossômicas não Histona , Proteínas Correpressoras , DNA Helicases/genética , DNA Helicases/metabolismo , Metilação de DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Heterocromatina/metabolismo , Histonas/genética , Histonas/metabolismo , Humanos , Chaperonas Moleculares , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas/genética , Proteínas/metabolismo , Proteínas Repressoras , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Nuclear Ligada ao X
7.
Hemasphere ; 8(5): e78, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38752170

RESUMO

α-Thalassemia represents one of the most important genetic modulators of ß-hemoglobinopathies. During this last decade, the ongoing interest in characterizing genotype-phenotype relationships has yielded incredible insights into α-globin gene regulation and its impact on ß-hemoglobinopathies. In this review, we provide a holistic update on α-globin gene expression stemming from DNA to RNA to protein, as well as epigenetic mechanisms that can impact gene expression and potentially influence phenotypic outcomes. Here, we highlight defined α-globin targeted strategies and rationalize the use of distinct molecular targets based on the restoration of balanced α/ß-like globin chain synthesis. Considering the therapies that either increase ß-globin synthesis or reactivate γ-globin gene expression, the modulation of α-globin chains as a disease modifier for ß-hemoglobinopathies still remains largely uncharted in clinical studies.

8.
Genome Biol ; 24(1): 284, 2023 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-38066546

RESUMO

BACKGROUND: Point mutations in histone variant H3.3 (H3.3K27M, H3.3G34R) and the H3.3-specific ATRX/DAXX chaperone complex are frequent events in pediatric gliomas. These H3.3 point mutations affect many chromatin modifications but the exact oncogenic mechanisms are currently unclear. Histone H3.3 is known to localize to nuclear compartments known as promyelocytic leukemia (PML) nuclear bodies, which are frequently mutated and confirmed as oncogenic drivers in acute promyelocytic leukemia. RESULTS: We find that the pediatric glioma-associated H3.3 point mutations disrupt the formation of PML nuclear bodies and this prevents differentiation down glial lineages. Similar to leukemias driven by PML mutations, H3.3-mutated glioma cells are sensitive to drugs that target PML bodies. We also find that point mutations in IDH1/2-which are common events in adult gliomas and myeloid leukemias-also disrupt the formation of PML bodies. CONCLUSIONS: We identify PML as a contributor to oncogenesis in a subset of gliomas and show that targeting PML bodies is effective in treating these H3.3-mutated pediatric gliomas.


Assuntos
Neoplasias Encefálicas , Glioma , Histonas , Adulto , Criança , Humanos , Neoplasias Encefálicas/genética , Glioma/genética , Histonas/genética , Mutação , Corpos Nucleares da Leucemia Promielocítica/genética , Corpos Nucleares da Leucemia Promielocítica/patologia
9.
Front Oncol ; 12: 1104129, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36686810

RESUMO

Pediatric high grade gliomas (HGG) are lethal tumors which are currently untreatable. A number of recent studies have provided much needed insights into the mutations and mechanisms which drive oncogenesis in pediatric HGGs. It is now clear that mutations in chromatin proteins, particularly H3.3 and its associated chaperone complex (ATRX), are a hallmark feature of pediatric HGGs. We review the current literature on the normal roles of the ATRX/H3.3 complex and how these functions are disrupted by oncogenic mutations. We discuss the current clinical trials and pre-clinical models that target chromatin and DNA, and how these agents fit into the ATRX/H3.3 mutation model. As chromatin mutations are a relatively new discovery in pediatric HGGs, developing clear mechanistic insights are a key step to improving therapies for these tumors.

10.
Nat Commun ; 9(1): 3142, 2018 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-30087349

RESUMO

An array of oncogenic histone point mutations have been identified across a number of different cancer studies. It has been suggested that some of these mutant histones can exert their effects by inhibiting epigenetic writers. Here, we report that the H3.3 G34R (glycine to arginine) substitution mutation, found in paediatric gliomas, causes widespread changes in H3K9me3 and H3K36me3 by interfering with the KDM4 family of K9/K36 demethylases. Expression of a targeted single-copy of H3.3 G34R at endogenous levels induced chromatin alterations that were comparable to a KDM4 A/B/C triple-knockout. We find that H3.3 G34R preferentially binds KDM4 while simultaneously inhibiting its enzymatic activity, demonstrating that histone mutations can act through inhibition of epigenetic erasers. These results suggest that histone point mutations can exert their effects through interactions with a range of epigenetic readers, writers and erasers.


Assuntos
Neoplasias Encefálicas/metabolismo , Cromatina/química , Glioblastoma/metabolismo , Histonas/metabolismo , Mutação , Mutação Puntual , Animais , Arginina/química , Biotinilação , Neoplasias Encefálicas/genética , Criança , Modelos Animais de Doenças , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Glioblastoma/genética , Glicina/química , Histonas/genética , Humanos , Camundongos , Ligação Proteica , Análise de Sequência de RNA , Transgenes
11.
Trends Cancer ; 2(3): 114-116, 2016 03.
Artigo em Inglês | MEDLINE | ID: mdl-28741530

RESUMO

Alternative lengthening of telomeres (ALT) is an enigmatic process that allows certain cancers to maintain telomeres in the absence of telomerase. ALT cancers are frequently defective for ATRX/DAXX, a chaperone complex that deposits histone variant H3.3 at telomeres. We propose that mutations in alpha thalassemia-mental retardation syndrome X-linked (ATRX)/death-domain associated protein (DAXX) prime ALT activation by disrupting telomeric heterochromatin.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Heterocromatina/metabolismo , Neoplasias/genética , Proteínas Nucleares/genética , Homeostase do Telômero , Proteína Nuclear Ligada ao X/genética , Proteínas Correpressoras , Humanos , Chaperonas Moleculares , Mutação , Telômero
12.
Cell Rep ; 11(3): 405-18, 2015 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-25865896

RESUMO

Histone H3.3 is a replication-independent histone variant, which replaces histones that are turned over throughout the entire cell cycle. H3.3 deposition at euchromatin is dependent on HIRA, whereas ATRX/Daxx deposits H3.3 at pericentric heterochromatin and telomeres. The role of H3.3 at heterochromatic regions is unknown, but mutations in the ATRX/Daxx/H3.3 pathway are linked to aberrant telomere lengthening in certain cancers. In this study, we show that ATRX-dependent deposition of H3.3 is not limited to pericentric heterochromatin and telomeres but also occurs at heterochromatic sites throughout the genome. Notably, ATRX/H3.3 specifically localizes to silenced imprinted alleles in mouse ESCs. ATRX KO cells failed to deposit H3.3 at these sites, leading to loss of the H3K9me3 heterochromatin modification, loss of repression, and aberrant allelic expression. We propose a model whereby ATRX-dependent deposition of H3.3 into heterochromatin is normally required to maintain the memory of silencing at imprinted loci.


Assuntos
Montagem e Desmontagem da Cromatina/fisiologia , DNA Helicases/genética , Inativação Gênica/fisiologia , Heterocromatina , Histonas/genética , Proteínas Nucleares/genética , Animais , Imunoprecipitação da Cromatina , DNA Helicases/metabolismo , Técnicas de Inativação de Genes , Loci Gênicos , Heterocromatina/metabolismo , Histonas/metabolismo , Camundongos , Proteínas Nucleares/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Proteína Nuclear Ligada ao X
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