Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Acta Neuropathol Commun ; 7(1): 77, 2019 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-31101070

RESUMO

The maintenance of genomic integrity is essential for normal cellular functions. However, it is difficult to maintain over a lifetime in postmitotic cells such as neurons, in which DNA damage increases with age and is exacerbated by multiple neurological disorders, including Alzheimer's disease (AD). Here we used immunohistochemical staining to detect DNA double strand breaks (DSBs), the most severe form of DNA damage, in postmortem brain tissues from patients with mild cognitive impairment (MCI) or AD and from cognitively unimpaired controls. Immunostaining for γH2AX-a post-translational histone modification that is widely used as a marker of DSBs-revealed increased proportions of γH2AX-labeled neurons and astrocytes in the hippocampus and frontal cortex of MCI and AD patients, as compared to age-matched controls. In contrast to the focal pattern associated with DSBs, some neurons and glia in humans and mice showed diffuse pan-nuclear patterns of γH2AX immunoreactivity. In mouse brains and primary neuronal cultures, such pan-nuclear γH2AX labeling could be elicited by increasing neuronal activity. To assess whether pan-nuclear γH2AX represents DSBs, we used a recently developed technology, DNA damage in situ ligation followed by proximity ligation assay, to detect close associations between γH2AX sites and free DSB ends. This assay revealed no evidence of DSBs in neurons or astrocytes with prominent pan-nuclear γH2AX labeling. These findings suggest that focal, but not pan-nuclear, increases in γH2AX immunoreactivity are associated with DSBs in brain tissue and that these distinct patterns of γH2AX formation may have different causes and consequences. We conclude that AD is associated with an accumulation of DSBs in vulnerable neuronal and glial cell populations from early stages onward. Because of the severe adverse effects this type of DNA damage can have on gene expression, chromatin stability and cellular functions, DSBs could be an important causal driver of neurodegeneration and cognitive decline in this disease.


Assuntos
Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Astrócitos/patologia , Quebras de DNA de Cadeia Dupla , Lobo Frontal/patologia , Hipocampo/patologia , Neurônios/patologia , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/metabolismo , Animais , Astrócitos/metabolismo , Disfunção Cognitiva/genética , Disfunção Cognitiva/patologia , Feminino , Lobo Frontal/metabolismo , Hipocampo/metabolismo , Histonas/metabolismo , Humanos , Masculino , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Neurônios/metabolismo
2.
Neurol Genet ; 4(3): e240, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29892709

RESUMO

OBJECTIVE: To describe the features of 2 unrelated adults with xeroderma pigmentosum complementation group F (XP-F) ascertained in a neurology care setting. METHODS: We report the clinical, imaging, molecular, and nucleotide excision repair (NER) capacity of 2 middle-aged women with progressive neurodegeneration ultimately diagnosed with XP-F. RESULTS: Both patients presented with adult-onset progressive neurologic deterioration involving chorea, ataxia, hearing loss, cognitive deficits, profound brain atrophy, and a history of skin photosensitivity, skin freckling, and/or skin neoplasms. We identified compound heterozygous pathogenic mutations in ERCC4 and confirmed deficient NER capacity in skin fibroblasts from both patients. CONCLUSIONS: These cases illustrate the role of NER dysfunction in neurodegeneration and how adult-onset neurodegeneration could be the major symptom bringing XP-F patients to clinical attention. XP-F should be considered by neurologists in the differential diagnosis of patients with adult-onset progressive neurodegeneration accompanied by global brain atrophy and a history of heightened sun sensitivity, excessive freckling, and skin malignancies.

3.
JAMA Neurol ; 75(8): 947-955, 2018 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-29710329

RESUMO

Importance: Identifying infectious causes of subacute or chronic meningitis can be challenging. Enhanced, unbiased diagnostic approaches are needed. Objective: To present a case series of patients with diagnostically challenging subacute or chronic meningitis using metagenomic next-generation sequencing (mNGS) of cerebrospinal fluid (CSF) supported by a statistical framework generated from mNGS of control samples from the environment and from patients who were noninfectious. Design, Setting, and Participants: In this case series, mNGS data obtained from the CSF of 94 patients with noninfectious neuroinflammatory disorders and from 24 water and reagent control samples were used to develop and implement a weighted scoring metric based on z scores at the species and genus levels for both nucleotide and protein alignments to prioritize and rank the mNGS results. Total RNA was extracted for mNGS from the CSF of 7 participants with subacute or chronic meningitis who were recruited between September 2013 and March 2017 as part of a multicenter study of mNGS pathogen discovery among patients with suspected neuroinflammatory conditions. The neurologic infections identified by mNGS in these 7 participants represented a diverse array of pathogens. The patients were referred from the University of California, San Francisco Medical Center (n = 2), Zuckerberg San Francisco General Hospital and Trauma Center (n = 2), Cleveland Clinic (n = 1), University of Washington (n = 1), and Kaiser Permanente (n = 1). A weighted z score was used to filter out environmental contaminants and facilitate efficient data triage and analysis. Main Outcomes and Measures: Pathogens identified by mNGS and the ability of a statistical model to prioritize, rank, and simplify mNGS results. Results: The 7 participants ranged in age from 10 to 55 years, and 3 (43%) were female. A parasitic worm (Taenia solium, in 2 participants), a virus (HIV-1), and 4 fungi (Cryptococcus neoformans, Aspergillus oryzae, Histoplasma capsulatum, and Candida dubliniensis) were identified among the 7 participants by using mNGS. Evaluating mNGS data with a weighted z score-based scoring algorithm reduced the reported microbial taxa by a mean of 87% (range, 41%-99%) when taxa with a combined score of 0 or less were removed, effectively separating bona fide pathogen sequences from spurious environmental sequences so that, in each case, the causative pathogen was found within the top 2 scoring microbes identified using the algorithm. Conclusions and Relevance: Diverse microbial pathogens were identified by mNGS in the CSF of patients with diagnostically challenging subacute or chronic meningitis, including a case of subarachnoid neurocysticercosis that defied diagnosis for 1 year, the first reported case of CNS vasculitis caused by Aspergillus oryzae, and the fourth reported case of C dubliniensis meningitis. Prioritizing metagenomic data with a scoring algorithm greatly clarified data interpretation and highlighted the problem of attributing biological significance to organisms present in control samples used for metagenomic sequencing studies.


Assuntos
Meningite/diagnóstico , Metagenoma/genética , Adolescente , Adulto , Animais , Aspergillus oryzae/genética , Candida/genética , Candidíase/líquido cefalorraquidiano , Candidíase/diagnóstico , Criança , Doença Crônica , Cryptococcus neoformans/genética , Feminino , Infecções por HIV/líquido cefalorraquidiano , Infecções por HIV/diagnóstico , HIV-1/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Histoplasma/genética , Histoplasmose/líquido cefalorraquidiano , Histoplasmose/diagnóstico , Humanos , Masculino , Meningite/líquido cefalorraquidiano , Meningite/microbiologia , Meningite Criptocócica/líquido cefalorraquidiano , Meningite Criptocócica/diagnóstico , Metagenômica , Pessoa de Meia-Idade , Neuroaspergilose/líquido cefalorraquidiano , Neuroaspergilose/diagnóstico , Neurocisticercose/líquido cefalorraquidiano , Neurocisticercose/diagnóstico , Análise de Sequência de RNA/métodos , Taenia solium/genética , Adulto Jovem
4.
Ann Neurol ; 78(5): 722-30, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26290222

RESUMO

OBJECTIVE: Identification of a particular cause of meningoencephalitis can be challenging owing to the myriad bacteria, viruses, fungi, and parasites that can produce overlapping clinical phenotypes, frequently delaying diagnosis and therapy. Metagenomic deep sequencing (MDS) approaches to infectious disease diagnostics are known for their ability to identify unusual or novel viruses and thus are well suited for investigating possible etiologies of meningoencephalitis. METHODS: We present the case of a 74-year-old woman with endophthalmitis followed by meningoencephalitis. MDS of her cerebrospinal fluid (CSF) was performed to identify an infectious agent. RESULTS: Sequences aligning to Balamuthia mandrillaris ribosomal RNA genes were identified in the CSF by MDS. Polymerase chain reaction subsequently confirmed the presence of B. mandrillaris in CSF, brain tissue, and vitreous fluid from the patient's infected eye. B. mandrillaris serology and immunohistochemistry for free-living amoebas on the brain biopsy tissue were positive. INTERPRETATION: The diagnosis was made using MDS after the patient had been hospitalized for several weeks and subjected to costly and invasive testing. MDS is a powerful diagnostic tool with the potential for rapid and unbiased pathogen identification leading to early therapeutic targeting.


Assuntos
Amebíase/diagnóstico , Amebíase/genética , Balamuthia mandrillaris/genética , Meningoencefalite/diagnóstico , Meningoencefalite/genética , Análise de Sequência de RNA/métodos , Idoso , Amebíase/líquido cefalorraquidiano , Animais , Encéfalo/microbiologia , DNA de Protozoário/genética , Feminino , Genômica , Humanos , Meningoencefalite/líquido cefalorraquidiano , Reação em Cadeia da Polimerase , Corpo Vítreo/microbiologia
5.
Methods Mol Biol ; 1042: 227-35, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23980011

RESUMO

Recent advances have led to several systems to study transcription from defined loci in living cells. It has now become possible to address long-standing questions regarding the interplay between the processes of DNA damage repair and transcription-two disparate processes that can occur on the same stretch of chromatin and which both lead to extensive chromatin change. Here we describe the development of a system to create enzymatically induced DNA double-strand breaks (DSBs) at a site of inducible transcription and methods to study the interplay between these processes.


Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , Microscopia de Fluorescência/métodos , Transcrição Gênica/genética , Animais , Antibacterianos/farmacologia , Antineoplásicos/farmacologia , Linhagem Celular , Cromatina/genética , Montagem e Desmontagem da Cromatina/fisiologia , Reparo do DNA/fisiologia , Desoxirribonucleases de Sítio Específico do Tipo II/metabolismo , Doxiciclina/farmacologia , Humanos , Óperon Lac/genética , Lasers , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Transcrição Gênica/fisiologia
6.
Nat Struct Mol Biol ; 20(3): 317-25, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23377543

RESUMO

The pathogenic sequelae of BRCA1 mutation in human and mouse cells are mitigated by concomitant deletion of 53BP1, which binds histone H4 dimethylated at Lys20 (H4K20me2) to promote nonhomologous end joining, suggesting that a balance between BRCA1 and 53BP1 regulates DNA double strand-break (DSB) repair mechanism choice. Here we document that acetylation is a key determinant of this balance. TIP60 acetyltransferase deficiency reduced BRCA1 at DSB chromatin with commensurate increases in 53BP1, whereas HDAC inhibition yielded the opposite effect. TIP60-dependent H4 acetylation diminished 53BP1 binding to H4K20me2 in part through disruption of a salt bridge between H4K16 and Glu1551 in the 53BP1 Tudor domain. Moreover, TIP60 deficiency impaired homologous recombination and conferred sensitivity to PARP inhibition in a 53BP1-dependent manner. These findings demonstrate that acetylation in cis to H4K20me2 regulates relative BRCA1 and 53BP1 DSB chromatin occupancy to direct DNA repair mechanism.


Assuntos
Proteína BRCA1/metabolismo , Cromatina/metabolismo , Histonas/metabolismo , Recombinação Homóloga , Peptídeos e Proteínas de Sinalização Intracelular/química , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , ATPases Associadas a Diversas Atividades Celulares , Acetilação , Sequência de Aminoácidos , Proteína BRCA1/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Células Cultivadas , Cromatina/genética , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Quebras de DNA de Cadeia Dupla , DNA Helicases/genética , DNA Helicases/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Técnicas de Silenciamento de Genes , Histona Acetiltransferases/genética , Histona Acetiltransferases/metabolismo , Histona Desacetilases/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Lisina Acetiltransferase 5 , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Conformação Proteica , Proteína 1 de Ligação à Proteína Supressora de Tumor p53
8.
Cell ; 141(6): 970-81, 2010 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-20550933

RESUMO

DNA double-strand breaks (DSBs) initiate extensive local and global alterations in chromatin structure, many of which depend on the ATM kinase. Histone H2A ubiquitylation (uH2A) on chromatin surrounding DSBs is one example, thought to be important for recruitment of repair proteins. uH2A is also implicated in transcriptional repression; an intriguing yet untested hypothesis is that this function is conserved in the context of DSBs. Using a novel reporter that allows for visualization of repair protein recruitment and local transcription in single cells, we describe an ATM-dependent transcriptional silencing program in cis to DSBs. ATM prevents RNA polymerase II elongation-dependent chromatin decondensation at regions distal to DSBs. Silencing is partially dependent on E3 ubiquitin ligases RNF8 and RNF168, whereas reversal of silencing relies on the uH2A deubiquitylating enzyme USP16. These findings give insight into the role of posttranslational modifications in mediating crosstalk between diverse processes occurring on chromatin.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Cromatina/metabolismo , Quebras de DNA de Cadeia Dupla , Proteínas de Ligação a DNA/metabolismo , Inativação Gênica , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteínas Mutadas de Ataxia Telangiectasia , Linhagem Celular Tumoral , Dano ao DNA , Histonas/metabolismo , Humanos , Transcrição Gênica , Ubiquitina Tiolesterase/metabolismo , Ubiquitinação
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...