RESUMO
Proteins involved in transcriptional regulation harbor a demonstrated enrichment of mutations in neurodevelopmental disorders. The Sin3 (Swi-independent 3)/histone deacetylase (HDAC) complex plays a central role in histone deacetylation and transcriptional repression. Among the two vertebrate paralogs encoding the Sin3 complex, SIN3A variants cause syndromic intellectual disability, but the clinical consequences of SIN3B haploinsufficiency in humans are uncharacterized. Here, we describe a syndrome hallmarked by intellectual disability, developmental delay, and dysmorphic facial features with variably penetrant autism spectrum disorder, congenital malformations, corpus callosum defects, and impaired growth caused by disruptive SIN3B variants. Using chromosomal microarray or exome sequencing, and through international data sharing efforts, we identified nine individuals with heterozygous SIN3B deletion or single-nucleotide variants. Five individuals harbor heterozygous deletions encompassing SIN3B that reside within a â¼230 kb minimal region of overlap on 19p13.11, two individuals have a rare nonsynonymous substitution, and two individuals have a single-nucleotide deletion that results in a frameshift and predicted premature termination codon. To test the relevance of SIN3B impairment to measurable aspects of the human phenotype, we disrupted the orthologous zebrafish locus by genome editing and transient suppression. The mutant and morphant larvae display altered craniofacial patterning, commissural axon defects, and reduced body length supportive of an essential role for Sin3 function in growth and patterning of anterior structures. To investigate further the molecular consequences of SIN3B variants, we quantified genome-wide enhancer and promoter activity states by using H3K27ac ChIP-seq. We show that, similar to SIN3A mutations, SIN3B disruption causes hyperacetylation of a subset of enhancers and promoters in peripheral blood mononuclear cells. Together, these data demonstrate that SIN3B haploinsufficiency leads to a hitherto unknown intellectual disability/autism syndrome, uncover a crucial role of SIN3B in the central nervous system, and define the epigenetic landscape associated with Sin3 complex impairment.
Assuntos
Transtorno do Espectro Autista/genética , Haploinsuficiência/genética , Histona Desacetilases/metabolismo , Deficiência Intelectual/genética , Proteínas Repressoras/genética , Acetilação , Adolescente , Animais , Criança , Pré-Escolar , Variações do Número de Cópias de DNA/genética , Feminino , Histonas/química , Histonas/metabolismo , Humanos , Lactente , Larva/genética , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Modelos Moleculares , Mutação , Proteínas Repressoras/deficiência , Proteínas Repressoras/metabolismo , Síndrome , Adulto Jovem , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/deficiência , Proteínas de Peixe-Zebra/genéticaRESUMO
KAT5 encodes an essential lysine acetyltransferase, previously called TIP60, which is involved in regulating gene expression, DNA repair, chromatin remodeling, apoptosis, and cell proliferation; but it remains unclear whether variants in this gene cause a genetic disease. Here, we study three individuals with heterozygous de novo missense variants in KAT5 that affect normally invariant residues, with one at the chromodomain (p.Arg53His) and two at or near the acetyl-CoA binding site (p.Cys369Ser and p.Ser413Ala). All three individuals have cerebral malformations, seizures, global developmental delay or intellectual disability, and severe sleep disturbance. Progressive cerebellar atrophy was also noted. Histone acetylation assays with purified variant KAT5 demonstrated that the variants decrease or abolish the ability of the resulting NuA4/TIP60 multi-subunit complexes to acetylate the histone H4 tail in chromatin. Transcriptomic analysis in affected individual fibroblasts showed deregulation of multiple genes that control development. Moreover, there was also upregulated expression of PER1 (a key gene involved in circadian control) in agreement with sleep anomalies in all of the individuals. In conclusion, dominant missense KAT5 variants cause histone acetylation deficiency with transcriptional dysregulation of multiples genes, thereby leading to a neurodevelopmental syndrome with sleep disturbance, cerebellar atrophy, and facial dysmorphisms, and suggesting a recognizable syndrome.
Assuntos
Atrofia/genética , Doenças Cerebelares/genética , Deficiência Intelectual/genética , Lisina Acetiltransferase 5/genética , Anormalidades Múltiplas/diagnóstico por imagem , Anormalidades Múltiplas/genética , Anormalidades Múltiplas/fisiopatologia , Adolescente , Adulto , Atrofia/diagnóstico por imagem , Atrofia/fisiopatologia , Doenças Cerebelares/diagnóstico por imagem , Doenças Cerebelares/fisiopatologia , Pré-Escolar , Cromatina/genética , Montagem e Desmontagem da Cromatina/genética , Reparo do DNA/genética , Epilepsia/diagnóstico por imagem , Epilepsia/genética , Epilepsia/fisiopatologia , Feminino , Heterozigoto , Histonas/genética , Humanos , Deficiência Intelectual/diagnóstico por imagem , Deficiência Intelectual/fisiopatologia , Masculino , Mutação de Sentido Incorreto/genética , Processamento de Proteína Pós-Traducional/genéticaRESUMO
Objective: To investigate the changes in serum inflammatory cytokines and the predictive factors for the efficacy of sertraline following medication therapy in adolescents with first-episode major depressive disorder (MDD). Methods: A total of 61 adolescent patients with first-episode drug-naïve MDD were enrolled for the MDD group and 55 healthy adolescents were enrolled for the healthy control (HC) group. Sertraline tablets were administered to the MDD group for 8 weeks after enrollment, while no medication was given to the HC group. In the MDD group, blood samples were collected to measure the cytokine levels and clinical data, including scores for the 17-item Hamilton Depression Scale (HAMD-17) and the Connor-Davidson Resilience Scale (CD-RISC), were assessed at baseline and at the end of the 8-week medication, whereas in the HC group, blood samples and clinical data were collected only at baseline. The correlation between the levels of serum inflammatory cytokines and depression severity in the MDD group was analyzed and stepwise linear regression of HAMD-17 in the MDD group was performed to find serologic indicators that could be used to predict the efficacy of sertraline. Results: At baseline, the levels of interleukin (IL)-1ß and IL-6 in the MDD group were significantly higher than those in the HC group (all P<0.0001), while the tumor necrosis factor (TNF)-α level in the MDD group was significantly lower than that in the HC group ( P=0.006). After 8 weeks of medication treatment, the MDD group showed decreased levels of IL-1ß and IL-6 and increased level of TNF-α compared to the pre-treatment levels. In addition, the HAMD-17 score, CD-RISC total score, and scores for perceived competence, trust and tolerance, and control, three factors of CD-RISC, all improved after treatment. There was no significant difference in serum cytokine levels at baseline between the subgroup showing response to the treatment and the non-responding subgroup. There was a weak correlation between IL-6 levels before and after treatment and CD-RISC scores and the scores for the trust and tolerance factor of CD-RISC before and after treatment. The baseline IL-1ß and TNF-α levels did not show significant effect on posttreatment HAMD-17 scores. Conclusions: Serum cytokine levels of adolescents with first-episode MDD differ significantly from those of healthy adolescents. Although IL-6 was found to be correlated with depression severity, there was not enough support for it to be used as a predictor of the antidepression efficacy of sertraline.
Assuntos
Antígenos de Grupos Sanguíneos , Transtorno Depressivo Maior , Humanos , Adolescente , Sertralina/uso terapêutico , Transtorno Depressivo Maior/tratamento farmacológico , Citocinas , Fator de Necrose Tumoral alfa , Interleucina-6 , Inflamação/tratamento farmacológico , Antígenos de Grupos Sanguíneos/uso terapêuticoRESUMO
Objective: To explore the differential expression of microRNAs (miRNAs) in brain-derived exosomes (BDEs) of adolescent mice with depression-like behavior. Methods: The experimental group consisted of susceptible adolescent mice exposed to chronic social defeat stress (CSDS), and sucrose preference test (SPT) and open field test (OFT) were performed to evaluate their depression-like behaviors. BDEs were extracted by ultracentrifugation (UC). The morphology, particle size, and surface marker proteins of BDEs were examined by transmission electron microscopy, nano-flow cytometry and Western blot. The expression of miRNA in BDEs was evaluated by high-throughput RNA sequencing. GO enrichment analysis and KEGG pathway enrichment analysis were carried out based on bioinformatics. Results: The particle size of BDEs ranged between 50 to 100 nm and they displayed a typical disc-shaped vesicle structure. TSG101 and syntenin, the exosome-positive proteins, were detected. In the BDEs of mice with depression-like behaviors induced by CSDS, 13 miRNAs were significantly upregulated and 4 miRNAs were significantly downregulated. Go and KEGG analysis showed that differentially expressed miRNAs were significantly enriched in PI3K-Akt signaling pathway, axonal guidance, and hypoxic response. Conclusion: It was found in this study that exosomal miRNAs in brain tissue might be involved in such biological processes as insulin resistance, neuroplasticity, and hypoxic response, thereby regulating brain functions and causing depression-like behaviors.
Assuntos
Exossomos , MicroRNAs , Animais , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Exossomos/química , Fosfatidilinositol 3-Quinases/metabolismo , Depressão , Encéfalo/metabolismoRESUMO
Isochorismatase domain-containing 1 (ISOC1) plays a carcinogenic role in various tumors. However, its expression and role in hepatocellular carcinoma (HCC) have not been elucidated. This is the first study to investigate the involvement of ISOC1 in HCC growth and migration. ISOC1 expression was analyzed using public databases and clinical samples, and clinical specimens were analyzed by real-time quantitative polymerase chain reaction, western blotting, and immunohistochemistry. ISOC1 was also overexpressed in two HCC cell lines (Huh7 and HepG2) to explore how ISOC1 affects HCC cells. Finally, a nude mouse xenograft tumor model was used to investigate the role of ISOC1 in HCC cell tumorigenicity. ISOC1 was downregulated in HCC tissues compared to that in matched paracancerous tissues, and low ISOC1 expression was associated with a poor prognosis. The proliferation and single-cell colony-forming ability of the ISOC1-overexpressing cell lines Huh7 and HepG2 were significantly inhibited. Moreover, ISOC1 overexpression suppressed the migration and invasion abilities of HCC cells in vitro, and ISOC1 upregulation hindered tumor growth in the xenograft tumor model in vivo. Therefore, ISOC1 is a potential HCC suppressor protein.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Animais , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Hidrolases , Neoplasias Hepáticas/genética , CamundongosRESUMO
Intrahepatic cholangiocarcinoma (ICC), a type of bile duct cancer, has a high mortality rate. Gut microbiota, bile acid (BA) metabolism, and cytokines have not been characterized in patients with ICC, and better noninvasive diagnostic approaches for ICC are essential to be established. Therefore, in this study we aimed to improve our understanding of changes in gut microbiota, BA metabolism, and cytokines in patients with ICC. We found that the α-diversities and ß-diversities of ICC were highest and that the abundances of four genera (Lactobacillus, Actinomyces, Peptostreptococcaceae, and Alloscardovia) were increased in patients with ICC compared with those in patients with hepatocellular carcinoma or liver cirrhosis and in healthy individuals. The glycoursodeoxycholic acid and tauroursodeoxycholic acid (TUDCA) plasma-stool ratios were obviously increased in patients with ICC. Furthermore, the genera Lactobacillus and Alloscardovia that were positively correlated with TUDCA plasma-stool ratios were combined to discriminate ICC from the other three diseases. Vascular invasion (VI) frequently led to a poor prognosis in patients with ICC. Compared with patients with ICC without VI, patients with VI had a greater abundance of the family Ruminococcaceae, increased levels of plasma interleukin (IL)-4 and six conjugated BAs, and decreased levels of plasma IL-6 and chenodeoxycholic acid. A positive correlation between plasma taurocholic acid and IL-4 was observed in patients with ICC. Plasma TUDCA was negatively correlated with the abundance of the genus Pseudoramibacter and the survival time of patients with ICC, but had no effect on tumor size, as determined in two murine tumor models. Conclusion: In this study, we identified some biomarkers, including gut microbiota, BAs and inflammatory cytokines, for the diagnosis of ICC and prediction of VI in patients with ICC.
Assuntos
Ácidos e Sais Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Colangiocarcinoma/patologia , Citocinas/sangue , Microbioma Gastrointestinal/fisiologia , Actinobacteria/isolamento & purificação , Animais , Neoplasias dos Ductos Biliares/imunologia , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/microbiologia , Colangiocarcinoma/imunologia , Colangiocarcinoma/metabolismo , Colangiocarcinoma/microbiologia , Humanos , Lactobacillus/isolamento & purificação , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Invasividade NeoplásicaRESUMO
Histone acetyltransferases (HATs) assemble into multisubunit complexes in order to target distinct lysine residues on nucleosomal histones. Here, we characterize native HAT complexes assembled by the BRPF family of scaffold proteins. Their plant homeodomain (PHD)-Zn knuckle-PHD domain is essential for binding chromatin and is restricted to unmethylated H3K4, a specificity that is reversed by the associated ING subunit. Native BRPF1 complexes can contain either MOZ/MORF or HBO1 as catalytic acetyltransferase subunit. Interestingly, while the previously reported HBO1 complexes containing JADE scaffold proteins target histone H4, the HBO1-BRPF1 complex acetylates only H3 in chromatin. We mapped a small region to the N terminus of scaffold proteins responsible for histone tail selection on chromatin. Thus, alternate choice of subunits associated with HBO1 can switch its specificity between H4 and H3 tails. These results uncover a crucial new role for associated proteins within HAT complexes, previously thought to be intrinsic to the catalytic subunit.
Assuntos
Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Acetilação , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Sequência de Aminoácidos , Cromatina/metabolismo , Proteínas de Ligação a DNA , Células HEK293 , Células HeLa , Histona Acetiltransferases/química , Histona Acetiltransferases/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Metilação , Dados de Sequência Molecular , Proteínas Nucleares/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Especificidade por Substrato , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/metabolismoRESUMO
During DNA replication, thousands of replication origins are activated across the genome. Chromatin architecture contributes to origin specification and usage, yet it remains unclear which chromatin features impact on DNA replication. Here, we perform a RNAi screen for chromatin regulators implicated in replication control by measuring RPA accumulation upon replication stress. We identify six factors required for normal rates of DNA replication and characterize a function of the bromodomain and PHD finger-containing protein 3 (BRPF3) in replication initiation. BRPF3 forms a complex with HBO1 that specifically acetylates histone H3K14, and genomewide analysis shows high enrichment of BRPF3, HBO1 and H3K14ac at ORC1-binding sites and replication origins found in the vicinity of TSSs. Consistent with this, BRPF3 is necessary for H3K14ac at selected origins and efficient origin activation. CDC45 recruitment, but not MCM2-7 loading, is impaired in BRPF3-depleted cells, identifying a BRPF3-dependent function of HBO1 in origin activation that is complementary to its role in licencing. We thus propose that BRPF3-HBO1 acetylation of histone H3K14 around TSS facilitates efficient activation of nearby replication origins.
Assuntos
Ciclo Celular/fisiologia , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Origem de Replicação/fisiologia , Acetilação , Ciclo Celular/genética , Linhagem Celular , Cromatina/metabolismo , Imunoprecipitação da Cromatina , Replicação do DNA/genética , Replicação do DNA/fisiologia , Histona Acetiltransferases/genética , Humanos , Imuno-Histoquímica , Origem de Replicação/genéticaRESUMO
Identification of over 500 epigenetic regulators in humans raises an interesting question regarding how chromatin dysregulation contributes to different diseases. Bromodomain and PHD finger-containing protein 1 (BRPF1) is a multivalent chromatin regulator possessing three histone-binding domains, one non-specific DNA-binding module, and several motifs for interacting with and activating three lysine acetyltransferases. Genetic analyses of fish brpf1 and mouse Brpf1 have uncovered an important role in skeletal, hematopoietic, and brain development, but it remains unclear how BRPF1 is linked to human development and disease. Here, we describe an intellectual disability disorder in ten individuals with inherited or de novo monoallelic BRPF1 mutations. Symptoms include infantile hypotonia, global developmental delay, intellectual disability, expressive language impairment, and facial dysmorphisms. Central nervous system and spinal abnormalities are also seen in some individuals. These clinical features overlap with but are not identical to those reported for persons with KAT6A or KAT6B mutations, suggesting that BRPF1 targets these two acetyltransferases and additional partners in humans. Functional assays showed that the resulting BRPF1 variants are pathogenic and impair acetylation of histone H3 at lysine 23, an abundant but poorly characterized epigenetic mark. We also found a similar deficiency in different lines of Brpf1-knockout mice. These data indicate that aberrations in the chromatin regulator gene BRPF1 cause histone H3 acetylation deficiency and a previously unrecognized intellectual disability syndrome.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Cromatina/metabolismo , Histonas/metabolismo , Deficiência Intelectual/genética , Mutação , Proteínas Nucleares/genética , Acetilação , Adolescente , Alelos , Animais , Proteínas de Transporte/genética , Criança , Cromatina/química , Proteínas de Ligação a DNA , Deficiências do Desenvolvimento/genética , Face/anormalidades , Feminino , Histona Acetiltransferases/genética , Humanos , Lisina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hipotonia Muscular/genética , SíndromeRESUMO
Protein lysine deacetylases have a pivotal role in numerous biological processes and can be divided into the Rpd3/Hda1 and sirtuin families, each having members in diverse organisms including prokaryotes. In vertebrates, the Rpd3/Hda1 family contains 11 members, traditionally referred to as histone deacetylases (HDAC) 1-11, which are further grouped into classes I, II and IV. Whereas most class I HDACs are subunits of multiprotein nuclear complexes that are crucial for transcriptional repression and epigenetic landscaping, class II members regulate cytoplasmic processes or function as signal transducers that shuttle between the cytoplasm and the nucleus. Little is known about class IV HDAC11, although its evolutionary conservation implies a fundamental role in various organisms.
Assuntos
Bactérias/enzimologia , Histona Desacetilases/metabolismo , Leveduras/enzimologia , Animais , Citoplasma/enzimologia , Doença , Histona Desacetilases/química , Histona Desacetilases/classificação , Humanos , Camundongos , Transdução de SinaisRESUMO
α-Tubulin acetyltransferase 1 (ATAT1) catalyzes acetylation of α-tubulin at lysine 40 in various organisms ranging from Tetrahymena to humans. Despite the importance in mammals suggested by studies of cultured cells, the mouse Atat1 gene is non-essential for survival, raising an intriguing question about its real functions in vivo. To address this question, we systematically analyzed a mouse strain lacking the gene. The analyses revealed that starting at postnatal day 5, the mutant mice display enlarged lateral ventricles in the forebrain, resembling ventricular dilation in human patients with ventriculomegaly. In the mice, ventricular dilation is due to hypoplasia in the septum and striatum. Behavioral tests of the mice uncovered deficits in motor coordination. Birth-dating experiments revealed that neuronal migration to the mutant septum and striatum is impaired during brain development. In the mutant embryonic fibroblasts, we found mild defects in cell proliferation and primary cilium formation. Notably, in these cells, ATAT1 is indispensable for tubulin hyperacetylation in response to high salt, high glucose, and hydrogen peroxide-induced oxidative stress. We investigated the role of ATAT1 in the hematopoietic system using multicolor flow cytometry and found that this system remains normal in the mutant mice. Although tubulin acetylation was undetectable in a majority of mutant tissues, residual levels were detected in the heart, skeletal muscle, trachea, oviduct, thymus and spleen. This study thus not only establishes the importance of ATAT1 in regulating mouse forebrain development and governing tubulin hyperacetylation during stress responses, but also suggests the existence of an additional α-tubulin acetyltransferase.
Assuntos
Acetiltransferases/metabolismo , Proteínas dos Microtúbulos/metabolismo , Estresse Oxidativo , Prosencéfalo/metabolismo , Tubulina (Proteína)/metabolismo , Acetilação/efeitos dos fármacos , Acetiltransferases/genética , Animais , Comportamento Animal , Movimento Celular , Proliferação de Células , Células Cultivadas , Cílios/metabolismo , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Peróxido de Hidrogênio/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas dos Microtúbulos/genética , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Neurogênese , Estresse Oxidativo/efeitos dos fármacos , Prosencéfalo/crescimento & desenvolvimento , Prosencéfalo/patologiaRESUMO
Short-chain acylation of lysine residues has recently emerged as a group of reversible posttranslational modifications in mammalian cells. The diversity of acylation further broadens the landscape and complexity of the proteome. Identification of regulatory enzymes and effector proteins for lysine acylation is critical to understand functions of these novel modifications at the molecular level. Here, we report that the MYST family of lysine acetyltransferases (KATs) possesses strong propionyltransferase activity both in vitro and in cellulo Particularly, the propionyltransferase activity of MOF, MOZ, and HBO1 is as strong as their acetyltransferase activity. Overexpression of MOF in human embryonic kidney 293T cells induced significantly increased propionylation in multiple histone and non-histone proteins, which shows that the function of MOF goes far beyond its canonical histone H4 lysine 16 acetylation. We also resolved the X-ray co-crystal structure of MOF bound with propionyl-coenzyme A, which provides a direct structural basis for the propionyltransferase activity of the MYST KATs. Our data together define a novel function for the MYST KATs as lysine propionyltransferases and suggest much broader physiological impacts for this family of enzymes.
Assuntos
Histona Acetiltransferases/metabolismo , Processamento de Proteína Pós-Traducional , Acetilação , Sequência de Aminoácidos , Células HEK293 , Histona Acetiltransferases/química , Humanos , Lisina/metabolismo , Modelos Moleculares , Conformação Proteica , ProteômicaRESUMO
Chromodomain helicase DNA-binding protein 4 (CHD4) is an ATP-dependent chromatin remodeler involved in epigenetic regulation of gene transcription, DNA repair, and cell cycle progression. Also known as Mi2ß, CHD4 is an integral subunit of a well-characterized histone deacetylase complex. Here we report five individuals with de novo missense substitutions in CHD4 identified through whole-exome sequencing and web-based gene matching. These individuals have overlapping phenotypes including developmental delay, intellectual disability, hearing loss, macrocephaly, distinct facial dysmorphisms, palatal abnormalities, ventriculomegaly, and hypogonadism as well as additional findings such as bone fusions. The variants, c.3380G>A (p.Arg1127Gln), c.3443G>T (p.Trp1148Leu), c.3518G>T (p.Arg1173Leu), and c.3008G>A, (p.Gly1003Asp) (GenBank: NM_001273.3), affect evolutionarily highly conserved residues and are predicted to be deleterious. Previous studies in yeast showed the equivalent Arg1127 and Trp1148 residues to be crucial for SNF2 function. Furthermore, mutations in the same positions were reported in malignant tumors, and a de novo missense substitution in an equivalent arginine residue in the C-terminal helicase domain of SMARCA4 is associated with Coffin Siris syndrome. Cell-based studies of the p.Arg1127Gln and p.Arg1173Leu mutants demonstrate normal localization to the nucleus and HDAC1 interaction. Based on these findings, the mutations potentially alter the complex activity but not its formation. This report provides evidence for the role of CHD4 in human development and expands an increasingly recognized group of Mendelian disorders involving chromatin remodeling and modification.
Assuntos
Trifosfato de Adenosina/metabolismo , Autoantígenos/genética , Montagem e Desmontagem da Cromatina/genética , Deficiência Intelectual/genética , Complexo Mi-2 de Remodelação de Nucleossomo e Desacetilase/genética , Mutação de Sentido Incorreto/genética , Anormalidades Múltiplas/genética , Adolescente , Animais , Núcleo Celular/metabolismo , Criança , Pré-Escolar , DNA Helicases/genética , Deficiências do Desenvolvimento/genética , Exoma/genética , Face/anormalidades , Feminino , Deformidades Congênitas da Mão/genética , Perda Auditiva/genética , Histona Desacetilase 1/metabolismo , Humanos , Masculino , Megalencefalia/genética , Camundongos , Micrognatismo/genética , Pescoço/anormalidades , Proteínas Nucleares/genética , Síndrome , Fatores de Transcrição/genéticaRESUMO
Translational control of gene expression plays a key role during the early phases of embryonic development. Here we describe a transcriptional regulator of mouse embryonic stem cells (mESCs), Yin-yang 2 (YY2), that is controlled by the translation inhibitors, Eukaryotic initiation factor 4E-binding proteins (4E-BPs). YY2 plays a critical role in regulating mESC functions through control of key pluripotency factors, including Octamer-binding protein 4 (Oct4) and Estrogen-related receptor-ß (Esrrb). Importantly, overexpression of YY2 directs the differentiation of mESCs into cardiovascular lineages. We show that the splicing regulator Polypyrimidine tract-binding protein 1 (PTBP1) promotes the retention of an intron in the 5'-UTR of Yy2 mRNA that confers sensitivity to 4E-BP-mediated translational suppression. Thus, we conclude that YY2 is a major regulator of mESC self-renewal and lineage commitment and document a multilayer regulatory mechanism that controls its expression.
Assuntos
Processamento Alternativo/fisiologia , Diferenciação Celular , Autorrenovação Celular/fisiologia , Células-Tronco Embrionárias/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Transcrição/metabolismo , Animais , Blastocisto/metabolismo , Proteínas de Transporte/metabolismo , Linhagem da Célula , Autorrenovação Celular/genética , Ribonucleoproteínas Nucleares Heterogêneas/genética , Íntrons , Camundongos , Camundongos Knockout , Modelos Biológicos , Fator 3 de Transcrição de Octâmero/metabolismo , Fosfoproteínas , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Biossíntese de Proteínas/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptores de Estrogênio/metabolismo , Fatores de Transcrição/genética , Transcrição Gênica/fisiologia , Fator de Transcrição YY1/metabolismoRESUMO
Lysine acetylation has recently emerged as an important post-translational modification in diverse organisms, but relatively little is known about its roles in mammalian development and stem cells. Bromodomain- and PHD finger-containing protein 1 (BRPF1) is a multidomain histone binder and a master activator of three lysine acetyltransferases, MOZ, MORF and HBO1, which are also known as KAT6A, KAT6B and KAT7, respectively. While the MOZ and MORF genes are rearranged in leukemia, the MORF gene is also mutated in prostate and other cancers and in four genetic disorders with intellectual disability. Here we show that forebrain-specific inactivation of the mouse Brpf1 gene causes hypoplasia in the dentate gyrus, including underdevelopment of the suprapyramidal blade and complete loss of the infrapyramidal blade. We trace the developmental origin to compromised Sox2+ neural stem cells and Tbr2+ intermediate neuronal progenitors. We further demonstrate that Brpf1 loss deregulates neuronal migration, cell cycle progression and transcriptional control, thereby causing abnormal morphogenesis of the hippocampus. These results link histone binding and acetylation control to hippocampus development and identify an important epigenetic regulator for patterning the dentate gyrus, a brain structure critical for learning, memory and adult neurogenesis.
Assuntos
Proteínas de Transporte/genética , Giro Denteado/metabolismo , Epigênese Genética/genética , Histona Acetiltransferases/metabolismo , Morfogênese/genética , Acetilação , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas de Transporte/metabolismo , Diferenciação Celular/genética , Proteínas de Ligação a DNA , Giro Denteado/crescimento & desenvolvimento , Giro Denteado/patologia , Hipocampo/crescimento & desenvolvimento , Hipocampo/patologia , Histona Acetiltransferases/genética , Histonas/metabolismo , Humanos , Camundongos , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/patologia , Prosencéfalo/embriologia , Prosencéfalo/crescimento & desenvolvimento , Prosencéfalo/metabolismo , Processamento de Proteína Pós-Traducional/genética , Proteínas com Domínio T/genéticaRESUMO
To interpret epigenetic information, chromatin readers utilize various protein domains for recognition of DNA and histone modifications. Some readers possess multidomains for modification recognition and are thus multivalent. Bromodomain- and plant homeodomain-linked finger-containing protein 3 (BRPF3) is such a chromatin reader, containing two plant homeodomain-linked fingers, one bromodomain and a PWWP domain. However, its molecular and biological functions remain to be investigated. Here, we report that endogenous BRPF3 preferentially forms a tetrameric complex with HBO1 (also known as KAT7) and two other subunits but not with related acetyltransferases such as MOZ, MORF, TIP60, and MOF (also known as KAT6A, KAT6B, KAT5, and KAT8, respectively). We have also characterized a mutant mouse strain with a lacZ reporter inserted at the Brpf3 locus. Systematic analysis of ß-galactosidase activity revealed dynamic spatiotemporal expression of Brpf3 during mouse embryogenesis and high expression in the adult brain and testis. Brpf3 disruption, however, resulted in no obvious gross phenotypes. This is in stark contrast to Brpf1 and Brpf2, whose loss causes lethality at E9.5 and E15.5, respectively. In Brpf3-null mice and embryonic fibroblasts, RT-quantitative PCR uncovered no changes in levels of Brpf1 and Brpf2 transcripts, confirming no compensation from them. These results indicate that BRPF3 forms a functional tetrameric complex with HBO1 but is not required for mouse development and survival, thereby distinguishing BRPF3 from its paralogs, BRPF1 and BRPF2.
Assuntos
Embrião de Mamíferos/enzimologia , Desenvolvimento Embrionário , Histona Acetiltransferases/metabolismo , Complexos Multienzimáticos/metabolismo , Animais , Perda do Embrião/enzimologia , Perda do Embrião/genética , Células HEK293 , Histona Acetiltransferases/genética , Humanos , Camundongos , Camundongos Mutantes , Complexos Multienzimáticos/genéticaRESUMO
The adenovirus early region 1A (E1A) oncoprotein hijacks host cells via direct interactions with many key cellular proteins, such as KAT2B, also known as PCAF (p300/CBP associated factor). E1A binds the histone acetyltransferase (HAT) domain of KAT2B to repress its transcriptional activation. However, the molecular mechanism by which E1A inhibits the HAT activity is not known. Here we demonstrate that a short and relatively conserved N-terminal motif (cNM) in the intrinsically disordered E1A protein is crucial for KAT2B interaction, and inhibits its HAT activity through a direct competition with acetyl-CoA, but not its substrate histone H3. Molecular modeling together with a series of mutagenesis experiments suggests that the major helix of E1A cNM binds to a surface of the acetyl-CoA pocket of the KAT2B HAT domain. Moreover, transient expression of the cNM peptide is sufficient to inhibit KAT2B-specific H3 acetylation H3K14ac in vivo Together, our data define an essential motif cNM in N-terminal E1A as an acetyl-CoA entry blocker that directly associates with the entrance of acetyl-CoA binding pocket to block the HAT domain access to its cofactor.
Assuntos
Proteínas E1A de Adenovirus/fisiologia , Lisina Acetiltransferases/antagonistas & inibidores , Acetilação , Proteínas E1A de Adenovirus/química , Motivos de Aminoácidos , Sequência de Aminoácidos , Humanos , Interações Hidrofóbicas e Hidrofílicas , Cinética , Modelos Moleculares , Homologia de Sequência de AminoácidosRESUMO
The HBO1 HAT protein is the major source of histone H4 acetylation in vivo and has been shown to play critical roles in gene regulation and DNA replication. A distinctive characteristic of HBO1 HAT complexes is the presence of three PHD finger domains in two different subunits: tumor suppressor proteins ING4/5 and JADE1/2/3. Biochemical and functional analyses indicate that these domains interact with histone H3 N-terminal tail region, but with a different specificity toward its methylation status. Their combinatorial action is essential in regulating chromatin binding and substrate specificity of HBO1 complexes, as well as cell growth. Importantly, localization analyses on the human genome indicate that HBO1 complexes are enriched throughout the coding regions of genes, supporting a role in transcription elongation. These results underline the importance and versatility of PHD finger domains in regulating chromatin association and histone modification crosstalk within a single protein complex.
Assuntos
Cromatina/metabolismo , Proteínas de Ligação a DNA/metabolismo , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Acetilação , Sítios de Ligação , Células Cultivadas , Proteínas de Ligação a DNA/genética , Células HeLa , Histona Acetiltransferases/genética , Histonas/genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Metilação , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMO
This paper was aimed to investigate the effect of Aralia echinocaulis containing serum on expression of ß-catenin, Wnt-1, Frizzed-2, TCF and Axin in Wnt/ß-catenin signaling pathway of primary osteoblasts. SD healthy female rats (n=80) were used to make A. echinocaulis containing serum by gastric perfusion for seven days with distilled water, A. echinocaulis decoction high dosage, middle dosage, and low dosage. In vitro, primary osteoblasts were cultured and identified. The third generation primary osteoblasts were taken and cultured for 48 h, then cells were treated with the different drug serums for 10 days and calcified nodules were counted by alizarin red staining. The cells were collected after treatment for 48 h and the expression levels of ß-catenin, Wnt-1, Frizzled-2, TCF and Axin were detected by Real-time PCR and Western blot. The results suggested that the in vitro cells were primary osteoblasts; and after treatment, various doses groups could promote the mineralization ability of primary osteoblasts, up-regulate the mRNA and protein expression levels of ß-catenin, Wnt-1, Frizzled-2, and TCF, and down-regulate the mRNA and protein expression levels of Axin. These findings indicated that A. echinocaulis containing serum can enhance the differentiation and proliferation of osteoblasts by regulating the expression levels of ß-catenin, Wnt-1, Frizzled-2, TCF and Axin in Wnt/ß-catenin signaling pathway of primary osteoblasts.
Assuntos
Aralia/química , Osteoblastos/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Feminino , Receptores Frizzled/metabolismo , Ratos , Ratos Sprague-Dawley , Proteína Wnt1/metabolismo , beta Catenina/metabolismoRESUMO
With hundreds of chromatin regulators identified in mammals, an emerging issue is how they modulate biological and pathological processes. BRPF1 (bromodomain- and PHD finger-containing protein 1) is a unique chromatin regulator possessing two PHD fingers, one bromodomain and a PWWP domain for recognizing multiple histone modifications. In addition, it binds to the acetyltransferases MOZ, MORF, and HBO1 (also known as KAT6A, KAT6B, and KAT7, respectively) to promote complex formation, restrict substrate specificity, and enhance enzymatic activity. We have recently showed that ablation of the mouse Brpf1 gene causes embryonic lethality at E9.5. Here we present systematic analyses of the mutant animals and demonstrate that the ablation leads to vascular defects in the placenta, yolk sac, and embryo proper, as well as abnormal neural tube closure. At the cellular level, Brpf1 loss inhibits proliferation of embryonic fibroblasts and hematopoietic progenitors. Molecularly, the loss reduces transcription of a ribosomal protein L10 (Rpl10)-like gene and the cell cycle inhibitor p27, and increases expression of the cell-cycle inhibitor p16 and a novel protein homologous to Scp3, a synaptonemal complex protein critical for chromosome association and embryo survival. These results uncover a crucial role of Brpf1 in controlling mouse embryo development and regulating cellular and gene expression programs.