Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 7 de 7
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Nat Cell Biol ; 26(5): 770-783, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38600236

RESUMO

DNA-protein crosslinks (DPCs) arise from enzymatic intermediates, metabolism or chemicals like chemotherapeutics. DPCs are highly cytotoxic as they impede DNA-based processes such as replication, which is counteracted through proteolysis-mediated DPC removal by spartan (SPRTN) or the proteasome. However, whether DPCs affect transcription and how transcription-blocking DPCs are repaired remains largely unknown. Here we show that DPCs severely impede RNA polymerase II-mediated transcription and are preferentially repaired in active genes by transcription-coupled DPC (TC-DPC) repair. TC-DPC repair is initiated by recruiting the transcription-coupled nucleotide excision repair (TC-NER) factors CSB and CSA to DPC-stalled RNA polymerase II. CSA and CSB are indispensable for TC-DPC repair; however, the downstream TC-NER factors UVSSA and XPA are not, a result indicative of a non-canonical TC-NER mechanism. TC-DPC repair functions independently of SPRTN but is mediated by the ubiquitin ligase CRL4CSA and the proteasome. Thus, DPCs in genes are preferentially repaired in a transcription-coupled manner to facilitate unperturbed transcription.


Assuntos
DNA Helicases , Enzimas Reparadoras do DNA , Reparo do DNA , Proteínas de Ligação a Poli-ADP-Ribose , Proteólise , RNA Polimerase II , Transcrição Gênica , Enzimas Reparadoras do DNA/metabolismo , Enzimas Reparadoras do DNA/genética , Humanos , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/genética , DNA Helicases/metabolismo , DNA Helicases/genética , RNA Polimerase II/metabolismo , RNA Polimerase II/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , DNA/metabolismo , DNA/genética , Células HEK293 , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Dano ao DNA , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas de Transporte , Receptores de Interleucina-17
2.
Nat Commun ; 15(1): 3490, 2024 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-38664429

RESUMO

Congenital nucleotide excision repair (NER) deficiency gives rise to several cancer-prone and/or progeroid disorders. It is not understood how defects in the same DNA repair pathway cause different disease features and severity. Here, we show that the absence of functional ERCC1-XPF or XPG endonucleases leads to stable and prolonged binding of the transcription/DNA repair factor TFIIH to DNA damage, which correlates with disease severity and induces senescence features in human cells. In vivo, in C. elegans, this prolonged TFIIH binding to non-excised DNA damage causes developmental arrest and neuronal dysfunction, in a manner dependent on transcription-coupled NER. NER factors XPA and TTDA both promote stable TFIIH DNA binding and their depletion therefore suppresses these severe phenotypical consequences. These results identify stalled NER intermediates as pathogenic to cell functionality and organismal development, which can in part explain why mutations in XPF or XPG cause different disease features than mutations in XPA or TTDA.


Assuntos
Caenorhabditis elegans , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA , Endonucleases , Fator de Transcrição TFIIH , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/genética , Humanos , Animais , Fator de Transcrição TFIIH/metabolismo , Fator de Transcrição TFIIH/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Endonucleases/metabolismo , Endonucleases/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteína de Xeroderma Pigmentoso Grupo A/metabolismo , Proteína de Xeroderma Pigmentoso Grupo A/genética , Ligação Proteica , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Mutação , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética
3.
Nucleic Acids Res ; 51(17): 9055-9074, 2023 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-37470997

RESUMO

The SWI/SNF family of ATP-dependent chromatin remodeling complexes is implicated in multiple DNA damage response mechanisms and frequently mutated in cancer. The BAF, PBAF and ncBAF complexes are three major types of SWI/SNF complexes that are functionally distinguished by their exclusive subunits. Accumulating evidence suggests that double-strand breaks (DSBs) in transcriptionally active DNA are preferentially repaired by a dedicated homologous recombination pathway. We show that different BAF, PBAF and ncBAF subunits promote homologous recombination and are rapidly recruited to DSBs in a transcription-dependent manner. The PBAF and ncBAF complexes promote RNA polymerase II eviction near DNA damage to rapidly initiate transcriptional silencing, while the BAF complex helps to maintain this transcriptional silencing. Furthermore, ARID1A-containing BAF complexes promote RNaseH1 and RAD52 recruitment to facilitate R-loop resolution and DNA repair. Our results highlight how multiple SWI/SNF complexes perform different functions to enable DNA repair in the context of actively transcribed genes.


Assuntos
Proteínas Cromossômicas não Histona , Estruturas R-Loop , Montagem e Desmontagem da Cromatina/genética , Proteínas Cromossômicas não Histona/metabolismo , DNA , Reparo do DNA/genética , Recombinação Homóloga/genética , Humanos
4.
Nucleic Acids Res ; 51(18): e93, 2023 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-37522336

RESUMO

Transcription-coupled nucleotide excision repair (TC-NER) is an important DNA repair mechanism that protects against the negative effects of transcription-blocking DNA lesions. Hereditary TC-NER deficiencies cause pleiotropic and often severe neurodegenerative and progeroid symptoms. While multiple assays have been developed to determine TC-NER activity for clinical and research purposes, monitoring TC-NER is hampered by the low frequency of repair events occurring in transcribed DNA. 'Recovery of RNA Synthesis' is widely used as indirect TC-NER assay based on the notion that lesion-blocked transcription only resumes after successful TC-NER. Here, we show that measuring novel synthesis of a protein after its compound-induced degradation prior to DNA damage induction is an equally effective but more versatile manner to indirectly monitor DNA repair activity in transcribed genes. This 'Recovery of Protein Synthesis' (RPS) assay can be adapted to various degradable proteins and readouts, including imaging and immunoblotting. Moreover, RPS allows real-time monitoring of TC-NER activity in various living cells types and even in differentiated tissues of living organisms. To illustrate its utility, we show that DNA repair in transcribed genes declines in aging muscle tissue of C. elegans. Therefore, the RPS assay constitutes an important novel clinical and research tool to investigate transcription-coupled DNA repair.


Assuntos
Caenorhabditis elegans , Reparo do DNA , Biossíntese de Proteínas , Transcrição Gênica , Animais , Caenorhabditis elegans/fisiologia , DNA/metabolismo , Dano ao DNA , Envelhecimento/metabolismo , Músculos/metabolismo
5.
Commun Biol ; 4(1): 1336, 2021 11 25.
Artigo em Inglês | MEDLINE | ID: mdl-34824371

RESUMO

The 10-subunit TFIIH complex is vital to transcription and nucleotide excision repair. Hereditary mutations in its smallest subunit, TTDA/GTF2H5, cause a photosensitive form of the rare developmental disorder trichothiodystrophy. Some trichothiodystrophy features are thought to be caused by subtle transcription or gene expression defects. TTDA/GTF2H5 knockout mice are not viable, making it difficult to investigate TTDA/GTF2H5 in vivo function. Here we show that deficiency of C. elegans TTDA ortholog GTF-2H5 is, however, compatible with life, in contrast to depletion of other TFIIH subunits. GTF-2H5 promotes TFIIH stability in multiple tissues and is indispensable for nucleotide excision repair, in which it facilitates recruitment of TFIIH to DNA damage. Strikingly, when transcription is challenged, gtf-2H5 embryos die due to the intrinsic TFIIH fragility in absence of GTF-2H5. These results support the idea that TTDA/GTF2H5 mutations cause transcription impairment underlying trichothiodystrophy and establish C. elegans as model for studying pathogenesis of this disease.


Assuntos
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/fisiologia , Reparo do DNA/genética , DNA de Helmintos/fisiologia , Fatores de Transcrição/genética , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Fatores de Transcrição/metabolismo
6.
Cell Rep ; 34(2): 108608, 2021 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-33440146

RESUMO

Hereditary DNA repair defects affect tissues differently, suggesting that in vivo cells respond differently to DNA damage. Knowledge of the DNA damage response, however, is largely based on in vitro and cell culture studies, and it is currently unclear whether DNA repair changes depending on the cell type. Here, we use in vivo imaging of the nucleotide excision repair (NER) endonuclease ERCC-1/XPF-1 in C. elegans to demonstrate tissue-specific NER activity. In oocytes, XPF-1 functions as part of global genome NER (GG-NER) to ensure extremely rapid removal of DNA-helix-distorting lesions throughout the genome. In contrast, in post-mitotic neurons and muscles, XPF-1 participates in NER of transcribed genes only. Strikingly, muscle cells appear more resistant to the effects of DNA damage than neurons. These results suggest a tissue-specific organization of the DNA damage response and may help to better understand pleiotropic and tissue-specific consequences of accumulating DNA damage.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , DNA Helicases/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Endonucleases/metabolismo , Animais , Caenorhabditis elegans , Feminino , Músculos/metabolismo , Músculos/efeitos da radiação , Neurônios/metabolismo , Neurônios/efeitos da radiação , Oócitos/metabolismo , Oócitos/efeitos da radiação , Especificidade de Órgãos , Raios Ultravioleta
7.
Sci Rep ; 10(1): 5426, 2020 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-32214115

RESUMO

Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is a photoreceptor-specific chaperone that stabilizes the effector enzyme of phototransduction, cGMP phosphodiesterase 6 (PDE6). Mutations in the AIPL1 gene cause a severe inherited retinal dystrophy, Leber congenital amaurosis type 4 (LCA4), that manifests as the loss of vision during the first year of life. In this study, we generated three-dimensional (3D) retinal organoids (ROs) from human induced pluripotent stem cells (hiPSCs) derived from an LCA4 patient carrying a Cys89Arg mutation in AIPL1. This study aimed to (i) explore whether the patient hiPSC-derived ROs recapitulate LCA4 disease phenotype, and (ii) generate a clinically relevant resource to investigate the molecular mechanism of disease and safely test novel therapies for LCA4 in vitro. We demonstrate reduced levels of the mutant AIPL1 and PDE6 proteins in patient organoids, corroborating the findings in animal models; however, patient-derived organoids maintained retinal cell cytoarchitecture despite significantly reduced levels of AIPL1.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas do Olho/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Organoides/metabolismo , Retina/metabolismo , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Transporte/metabolismo , Linhagem Celular , Proteínas do Olho/genética , Humanos , Amaurose Congênita de Leber/genética , Amaurose Congênita de Leber/metabolismo , Mutação/genética
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...