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1.
Adv Exp Med Biol ; 1287: 31-46, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33034024

RESUMO

The endosomal pathway plays a pivotal role upon signal transduction in the Notch pathway. Recent work on lethal (2) giant discs (lgd) points to an additional critical role in avoiding uncontrolled ligand-independent signalling during trafficking of the Notch receptor through the endosomal pathway to the lysosome for degradation. In this chapter, we will outline the journey of Notch through the endosomal system and present an overview of the current knowledge about Lgd and its mammalian orthologs Lgd1/CC2D1b and Lgd2/CC2D1a. We will then discuss how Notch is activated in the absence of lgd function in Drosophila and ask whether there is evidence that a similar ligand-independent activation of the Notch pathway can also happen in mammals if the orthologs are inactivated.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/metabolismo , Neoplasias/metabolismo , Receptores Notch/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismo , Animais , Endossomos/metabolismo , Humanos
2.
Nat Commun ; 11(1): 4956, 2020 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-33009383

RESUMO

Tet-enzyme-mediated 5-hydroxymethylation of cytosines in DNA plays a crucial role in mouse embryonic stem cells (ESCs). In RNA also, 5-hydroxymethylcytosine (5hmC) has recently been evidenced, but its physiological roles are still largely unknown. Here we show the contribution and function of this mark in mouse ESCs and differentiating embryoid bodies. Transcriptome-wide mapping in ESCs reveals hundreds of messenger RNAs marked by 5hmC at sites characterized by a defined unique consensus sequence and particular features. During differentiation a large number of transcripts, including many encoding key pluripotency-related factors (such as Eed and Jarid2), show decreased cytosine hydroxymethylation. Using Tet-knockout ESCs, we find Tet enzymes to be partly responsible for deposition of 5hmC in mRNA. A transcriptome-wide search further reveals mRNA targets to which Tet1 and Tet2 bind, at sites showing a topology similar to that of 5hmC sites. Tet-mediated RNA hydroxymethylation is found to reduce the stability of crucial pluripotency-promoting transcripts. We propose that RNA cytosine 5-hydroxymethylation by Tets is a mark of transcriptome flexibility, inextricably linked to the balance between pluripotency and lineage commitment.


Assuntos
5-Metilcitosina/análogos & derivados , Diferenciação Celular , Proteínas de Ligação a DNA/metabolismo , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , RNA/metabolismo , 5-Metilcitosina/metabolismo , Animais , Especificidade de Anticorpos/imunologia , Sequência de Bases , Corpos Embrioides/metabolismo , Camundongos , Modelos Biológicos , Células-Tronco Pluripotentes/metabolismo , Ligação Proteica , Estabilidade de RNA/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcriptoma/genética
3.
Zhonghua Zhong Liu Za Zhi ; 42(8): 629-634, 2020 Aug 23.
Artigo em Chinês | MEDLINE | ID: mdl-32867453

RESUMO

Objective: To investigate the effect of esculin on the proliferation of triple negative breast cancer cells and its molecular mechanism. Methods: MDA-MB-231 cells were treated with 28, 56, 112, 225, 450 and 900 µmol/L of esculin for 24, 48 and 72 h, respectively, and the cell viability was detected by cell counting kit 8 (CCK-8) assay. In addition, MDA-MB-231 cells were treated with 0, 225, 450 and 900 µmol/L of esculin for 48 h. And then the changes in cell morphology were observed by inverted microscope. The clone-forming ability was detected by colony formation assay. The mRNA expression levels of FBI-1, p53 and p21 were detected using real-time fluorescence quantitative polymerase chain reaction. The protein expression levels of FBI-1, p53, p21 and Ki67 were detected by western blot. Results: Compared with the blank control group, the cell viability of MDA-MB-231 cells that treated with esculin significantly decreased in a dose-dependent and time-dependent manners. After treatment with esculin, MDA-MB-231 cells shrunk, flattened, adhered poorly to the culture dish and the cell spacing became larger. Meanwhile, shedding and incomplete cells appeared, of which 900 µmol/L of esculin treatment group showed the most dramatic changes. In addition, the colony formation ratios were decreased to (77.18±5.13)%, (65.94±4.98)% and (45.92±3.70)% in the 225, 450 and 900 µmol/L of esculin treatment groups compared with blank control, respectively (P<0.01). Furthermore, the mRNA and protein expressions of FBI-1 increased, while the levels of p53 and p21 mRNA and protein, as well as the protein expression of Ki67 decreased in a concentration-dependent manner (P<0.01). Conclusion: Esculin may regulate cell cycle-related p53-p21 pathway via FBI-1 mediated DNA replication, thus inhibit the proliferation of triple negative breast cancer cells.


Assuntos
Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Esculina/farmacologia , RNA Mensageiro/genética , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Neoplasias da Mama/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proteínas de Ligação a DNA , Regulação para Baixo/efeitos dos fármacos , Feminino , Humanos , Fatores de Transcrição , Neoplasias de Mama Triplo Negativas/patologia
4.
PLoS Genet ; 16(8): e1008976, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866141

RESUMO

Neural circuitry for mating and reproduction resides within the terminal segments of central nervous system (CNS) which express Hox paralogous group 9-13 (in vertebrates) or Abdominal-B (Abd-B) in Drosophila. Terminal neuroblasts (NBs) in A8-A10 segments of Drosophila larval CNS are subdivided into two groups based on expression of transcription factor Doublesex (Dsx). While the sex specific fate of Dsx-positive NBs is well investigated, the fate of Dsx-negative NBs is not known so far. Our studies with Dsx-negative NBs suggests that these cells, like their abdominal counterparts (in A3-A7 segments) use Hox, Grainyhead (Grh) and Notch to undergo cell death during larval development. This cell death also happens by transcriptionally activating RHG family of apoptotic genes through a common apoptotic enhancer in early to mid L3 stages. However, unlike abdominal NBs (in A3-A7 segments) which use increasing levels of resident Hox factor Abdominal-A (Abd-A) as an apoptosis trigger, Dsx-negative NBs (in A8-A10 segments) keep the levels of resident Hox factor Abd-B constant. These cells instead utilize increasing levels of the temporal transcription factor Grh and a rise in Notch activity to gain apoptotic competence. Biochemical and in vivo analysis suggest that Abdominal-A and Grh binding motifs in the common apoptotic enhancer also function as Abdominal-B and Grh binding motifs and maintains the enhancer activity in A8-A10 NBs. Finally, the deletion of this enhancer by the CRISPR-Cas9 method blocks the apoptosis of Dsx-negative NBs. These results highlight the fact that Hox dependent NB apoptosis in abdominal and terminal regions utilizes common molecular players (Hox, Grh and Notch), but seems to have evolved different molecular strategies to pattern CNS.


Assuntos
Apoptose/genética , Proteínas de Ligação a DNA/genética , Proteínas de Drosophila/genética , Receptores Notch/genética , Fatores de Transcrição/genética , Abdome/crescimento & desenvolvimento , Animais , Sistema Nervoso Central/crescimento & desenvolvimento , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Larva/genética , Larva/crescimento & desenvolvimento , Masculino , Células-Tronco Neurais/metabolismo , Sequências Reguladoras de Ácido Nucleico/genética
5.
Nat Commun ; 11(1): 4673, 2020 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-32938917

RESUMO

RAS-MAPK signaling mediates processes critical to normal development including cell proliferation, survival, and differentiation. Germline mutation of RAS-MAPK genes lead to the Noonan-spectrum of syndromes. Here, we present a patient affected by a 6p-interstitial microdeletion with unknown underlying molecular etiology. Examination of 6p-interstitial microdeletion cases reveals shared clinical features consistent with Noonan-spectrum disorders including short stature, facial dysmorphia and cardiovascular abnormalities. We find the RAS-responsive element binding protein-1 (RREB1) is the common deleted gene in multiple 6p-interstitial microdeletion cases. Rreb1 hemizygous mice display orbital hypertelorism and cardiac hypertrophy phenocopying the human syndrome. Rreb1 haploinsufficiency leads to sensitization of MAPK signaling. Rreb1 recruits Sin3a and Kdm1a to control H3K4 methylation at MAPK pathway gene promoters. Haploinsufficiency of SIN3A and mutations in KDM1A cause syndromes similar to RREB1 haploinsufficiency suggesting genetic perturbation of the RREB1-SIN3A-KDM1A complex represents a new category of RASopathy-like syndromes arising through epigenetic reprogramming of MAPK pathway genes.


Assuntos
Proteínas de Ligação a DNA/genética , Haploinsuficiência , Sistema de Sinalização das MAP Quinases/genética , Síndrome de Noonan/etiologia , Fatores de Transcrição/genética , Proteínas ras/metabolismo , Anormalidades Múltiplas/genética , Animais , Deleção Cromossômica , Cromossomos Humanos Par 6 , Proteínas de Ligação a DNA/metabolismo , Epigênese Genética , Feminino , Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Histonas/metabolismo , Humanos , Masculino , Metilação , Camundongos Endogâmicos C57BL , Camundongos Knockout , Complexo Correpressor Histona Desacetilase e Sin3/genética , Complexo Correpressor Histona Desacetilase e Sin3/metabolismo , Fatores de Transcrição/metabolismo , Proteínas ras/genética
6.
Nat Commun ; 11(1): 4666, 2020 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-32938943

RESUMO

Intercalated discs (ICD), specific cell-to-cell contacts that connect adjacent cardiomyocytes, ensure mechanical and electrochemical coupling during contraction of the heart. Mutations in genes encoding ICD components are linked to cardiovascular diseases. Here, we show that loss of Xinß, a newly-identified component of ICDs, results in cardiomyocyte proliferation defects and cardiomyopathy. We uncovered a role for Xinß in signaling via the Hippo-YAP pathway by recruiting NF2 to the ICD to modulate cardiac function. In Xinß mutant hearts levels of phosphorylated NF2 are substantially reduced, suggesting an impairment of Hippo-YAP signaling. Cardiac-specific overexpression of YAP rescues cardiac defects in Xinß knock-out mice-indicating a functional and genetic interaction between Xinß and YAP. Our study reveals a molecular mechanism by which cardiac-expressed intercalated disc protein Xinß modulates Hippo-YAP signaling to control heart development and cardiac function in a tissue specific manner. Consequently, this pathway may represent a therapeutic target for the treatment of cardiovascular diseases.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas do Citoesqueleto/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas com Domínio LIM/metabolismo , Miócitos Cardíacos/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Cardiomiopatia Dilatada/genética , Comunicação Celular , Proteínas de Ciclo Celular/genética , Proliferação de Células , Proteínas do Citoesqueleto/genética , Proteínas de Ligação a DNA/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Ventrículos do Coração/crescimento & desenvolvimento , Proteínas com Domínio LIM/genética , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação , Miócitos Cardíacos/citologia , Miócitos Cardíacos/patologia , Neurofibromina 2/genética , Neurofibromina 2/metabolismo , Proteínas Nucleares/genética , Transdução de Sinais
7.
Nat Commun ; 11(1): 4471, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32901010

RESUMO

A human cell contains hundreds to thousands of mitochondrial DNA (mtDNA) packaged into nucleoids. Currently, the segregation and allocation of nucleoids are thought to be passively determined by mitochondrial fusion and division. Here we provide evidence, using live-cell super-resolution imaging, that nucleoids can be actively transported via KIF5B-driven mitochondrial dynamic tubulation (MDT) activities that predominantly occur at the ER-mitochondria contact sites (EMCS). We further demonstrate that a mitochondrial inner membrane protein complex MICOS links nucleoids to Miro1, a KIF5B receptor on mitochondria, at the EMCS. We show that such active transportation is a mechanism essential for the proper distribution of nucleoids in the peripheral zone of the cell. Together, our work identifies an active transportation mechanism of nucleoids, with EMCS serving as a key platform for the interplay of nucleoids, MICOS, Miro1, and KIF5B to coordinate nucleoids segregation and transportation.


Assuntos
DNA Mitocondrial/metabolismo , Retículo Endoplasmático/metabolismo , Mitocôndrias/metabolismo , Dinâmica Mitocondrial/fisiologia , Animais , Transporte Biológico Ativo , Células COS , Células Cultivadas , Chlorocebus aethiops , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células HEK293 , Humanos , Cinesina/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Camundongos , Membranas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Modelos Biológicos , Ratos , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transfecção , Proteínas rho de Ligação ao GTP/metabolismo
8.
Nat Commun ; 11(1): 4515, 2020 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-32908127

RESUMO

The discovery of ancestral RAG transposons in early deuterostomia reveals the origin of vertebrate V(D)J recombination. Here, we analyze the functional regulation of a RAG transposon, ProtoRAG, in lancelet. We find that a specific interaction between the cis-acting element within the TIR sequences of ProtoRAG and a trans-acting factor, lancelet YY1-like (bbYY1), is important for the transcriptional regulation of lancelet RAG-like genes (bbRAG1L and bbRAG2L). Mechanistically, bbYY1 suppresses the transposition of ProtoRAG; meanwhile, bbYY1 promotes host DNA rejoins (HDJ) and TIR-TIR joints (TTJ) after TIR-dependent excision by facilitating the binding of bbRAG1L/2 L to TIR-containing DNA, and by interacting with the bbRAG1L/2 L complex. Our data thus suggest that bbYY1 has dual functions in fine-tuning the activity of ProtoRAG and maintaining the genome stability of the host.


Assuntos
Elementos de DNA Transponíveis/genética , Proteínas de Ligação a DNA/metabolismo , Anfioxos/genética , Recombinação V(D)J , Fator de Transcrição YY1/metabolismo , Animais , Técnicas de Silenciamento de Genes , Genes RAG-1 , Instabilidade Genômica , Células HEK293 , Células HeLa , Humanos , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Fator de Transcrição YY1/genética , Fator de Transcrição YY1/isolamento & purificação
9.
Nat Commun ; 11(1): 4642, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32934200

RESUMO

Epigenetic regulation plays an important role in governing stem cell fate and tumorigenesis. Lost expression of a key DNA demethylation enzyme TET2 is associated with human cancers and has been linked to stem cell traits in vitro; however, whether and how TET2 regulates mammary stem cell fate and mammary tumorigenesis in vivo remains to be determined. Here, using our recently established mammary specific Tet2 deletion mouse model, the data reveals that TET2 plays a pivotal role in mammary gland development and luminal lineage commitment. We show that TET2 and FOXP1 form a chromatin complex that mediates demethylation of ESR1, GATA3, and FOXA1, three key genes that are known to coordinately orchestrate mammary luminal lineage specification and endocrine response, and also are often silenced by DNA methylation in aggressive breast cancers. Furthermore, Tet2 deletion-PyMT breast cancer mouse model exhibits enhanced mammary tumor development with deficient ERα expression that confers tamoxifen resistance in vivo. As a result, this study elucidates a role for TET2 in governing luminal cell differentiation and endocrine response that underlies breast cancer resistance to anti-estrogen treatments.


Assuntos
Diferenciação Celular , Proteínas de Ligação a DNA/metabolismo , Estradiol/metabolismo , Estrogênios/metabolismo , Glândulas Mamárias Animais/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/fisiopatologia , Linhagem da Célula , Metilação de DNA , Proteínas de Ligação a DNA/genética , Sistema Endócrino/metabolismo , Epigênese Genética , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Glândulas Mamárias Animais/fisiopatologia , Camundongos , Camundongos Knockout , Proteínas Proto-Oncogênicas/genética
10.
Nat Commun ; 11(1): 4718, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32948777

RESUMO

Disturbances in glucose homeostasis and low-grade chronic inflammation culminate into metabolic syndrome that increase the risk for the development of type 2 diabetes mellitus (T2DM). The recently discovered group 2 innate lymphoid cells (ILC2s) are capable of secreting copious amounts of type 2 cytokines to modulate metabolic homeostasis in adipose tissue. In this study, we have established that expression of Death Receptor 3 (DR3), a member of the TNF superfamily, on visceral adipose tissue (VAT)-derived murine and peripheral blood human ILC2s is inducible by IL-33. We demonstrate that DR3 engages the canonical and/or non-canonical NF-κB pathways, and thus stimulates naïve and co-stimulates IL-33-activated ILC2s. Importantly, DR3 engagement on ILC2s significantly ameliorates glucose tolerance, protects against insulin-resistance onset and remarkably reverses already established insulin-resistance. Taken together, these results convey the potent role of DR3 as an ILC2 regulator and introduce DR3 agonistic treatment as a novel therapeutic avenue for treating T2DM.


Assuntos
Tecido Adiposo/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Linfócitos/metabolismo , Membro 25 de Receptores de Fatores de Necrose Tumoral/metabolismo , Adipócitos/metabolismo , Adolescente , Adulto , Idoso , Animais , Citocinas/metabolismo , Proteínas de Ligação a DNA/genética , Feminino , Glucose/metabolismo , Homeostase , Humanos , Imunidade Inata , Resistência à Insulina , Interleucina-33/metabolismo , Gordura Intra-Abdominal/metabolismo , Masculino , Síndrome Metabólica/complicações , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Membro 25 de Receptores de Fatores de Necrose Tumoral/uso terapêutico , Adulto Jovem
13.
Anticancer Res ; 40(10): 5883-5893, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32988919

RESUMO

BACKGROUND/AIM: Somatic mutations were investigated in 21 patients with postmenopausal estrogen receptor (ER)-positive and human epidermal growth factor receptor-2 (HER-2)-positive (ER+HER2+) breast cancer (BC) treated with neoadjuvant letrozole and lapatinib, to identify their distinct molecular landscape. PATIENTS AND METHODS: We used tissue samples of 21 patients from phase II Neo ALL-IN cohort, and somatic alterations were examined using targeted exome sequencing performed in Foundation Medicine, Inc. (FMI). RESULTS: TP53 (61.9%) and PIK3CA (57.1%) were the two most frequently mutated genes that were inter-correlated (p=0.026). They were associated with unfavorable clinical outcomes, particularly when accompanying PIK3CA mutations at exon 9 in helical domains. Meanwhile, MLL2 alteration was negatively associated with mutations of TP53 or PIK3CA, and it tended to be present in patients with low KI-67 levels and no initial nodal involvement. Moreover, patients with MLL2 mutations numerically showed more favorable overall response rates (ORR) (80% vs. 56.2%) and better 5-year event-free survival (EFS) rates (100% vs. 87.5%) compared to the wild-type. CONCLUSION: Mutations in TP53 and PIK3CA hotspot at exon 9 may be potential negative predictors of ER+HER2+ BC treated with neoadjuvant letrozole and lapatinib, while MLL2 inactivating mutation might confer therapeutic benefit in these patients.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Classe I de Fosfatidilinositol 3-Quinases/genética , Proteínas de Ligação a DNA/genética , Proteínas de Neoplasias/genética , Proteína Supressora de Tumor p53/genética , Idoso , Biomarcadores Tumorais/genética , Mama/efeitos dos fármacos , Mama/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Receptor alfa de Estrogênio/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lapatinib/administração & dosagem , Letrozol/administração & dosagem , Pessoa de Meia-Idade , Mutação/genética , Intervalo Livre de Progressão , Receptor ErbB-2/genética
14.
Nat Commun ; 11(1): 4782, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32963223

RESUMO

Polycomb and Trithorax group proteins maintain stable epigenetic memory of gene expression states for some genes, but many targets show highly dynamic regulation. Here we combine experiment and theory to examine the mechanistic basis of these different modes of regulation. We present a mathematical model comprising a Polycomb/Trithorax response element (PRE/TRE) coupled to a promoter and including Drosophila developmental timing. The model accurately recapitulates published studies of PRE/TRE mediated epigenetic memory of both silencing and activation. With minimal parameter changes, the same model can also recapitulate experimental data for a different PRE/TRE that allows dynamic regulation of its target gene. The model predicts that both cell cycle length and PRE/TRE identity are critical for determining whether the system gives stable memory or dynamic regulation. Our work provides a simple unifying framework for a rich repertoire of PRE/TRE functions, and thus provides insights into  genome-wide Polycomb/Trithorax regulation.


Assuntos
Proteínas Cromossômicas não Histona/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Epigenômica , Regulação da Expressão Gênica no Desenvolvimento/genética , Modelos Teóricos , Complexo Repressor Polycomb 1/genética , Animais , Divisão Celular , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Epigênese Genética , Feminino , Inativação Gênica , Complexo Repressor Polycomb 1/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Regiões Promotoras Genéticas , Elementos de Resposta
15.
Nat Commun ; 11(1): 4784, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32963245

RESUMO

Genomic integrity is threatened by cytotoxic DNA double-strand breaks (DSBs), which must be resolved efficiently to prevent sequence loss, chromosomal rearrangements/translocations, or cell death. Polymerase µ (Polµ) participates in DSB repair via the nonhomologous end-joining (NHEJ) pathway, by filling small sequence gaps in broken ends to create substrates ultimately ligatable by DNA Ligase IV. Here we present structures of human Polµ engaging a DSB substrate. Synapsis is mediated solely by Polµ, facilitated by single-nucleotide homology at the break site, wherein both ends of the discontinuous template strand are stabilized by a hydrogen bonding network. The active site in the quaternary Pol µ complex is poised for catalysis and nucleotide incoporation proceeds in crystallo. These structures demonstrate that Polµ may address complementary DSB substrates during NHEJ in a manner indistinguishable from single-strand breaks.


Assuntos
Quebras de DNA de Cadeia Dupla , DNA Polimerase Dirigida por DNA/química , DNA/química , Cristalografia por Raios X , Dano ao DNA , Reparo do DNA por Junção de Extremidades , DNA Ligase Dependente de ATP/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA/química , Humanos , Ligação de Hidrogênio , Modelos Moleculares , Conformação Proteica
16.
Nat Commun ; 11(1): 4534, 2020 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-32913330

RESUMO

Collisions between the DNA replication machinery and co-transcriptional R-loops can impede DNA synthesis and are a major source of genomic instability in cancer cells. How cancer cells deal with R-loops to proliferate is poorly understood. Here we show that the ATP-dependent chromatin remodelling INO80 complex promotes resolution of R-loops to prevent replication-associated DNA damage in cancer cells. Depletion of INO80 in prostate cancer PC3 cells leads to increased R-loops. Overexpression of the RNA:DNA endonuclease RNAse H1 rescues the DNA synthesis defects and suppresses DNA damage caused by INO80 depletion. R-loops co-localize with and promote recruitment of INO80 to chromatin. Artificial tethering of INO80 to a LacO locus enabled turnover of R-loops in cis. Finally, counteracting R-loops by INO80 promotes proliferation and averts DNA damage-induced death in cancer cells. Our work suggests that INO80-dependent resolution of R-loops promotes DNA replication in the presence of transcription, thus enabling unlimited proliferation in cancers.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/metabolismo , Proliferação de Células/genética , Replicação do DNA , Proteínas de Ligação a DNA/metabolismo , Neoplasias/genética , Estruturas R-Loop/genética , Apoptose/genética , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Montagem e Desmontagem da Cromatina , Dano ao DNA , Instabilidade Genômica , Humanos , Neoplasias/patologia , Transcrição Genética
17.
Elife ; 92020 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-32915136

RESUMO

In starving yeast exposed to thermal stress, a transient drop in intracellular pH helps to trigger the heat shock response.


Assuntos
Proteínas de Saccharomyces cerevisiae , Proteínas de Ligação a DNA , Proteínas de Choque Térmico , Resposta ao Choque Térmico , Concentração de Íons de Hidrogênio , Fatores de Transcrição
18.
Nan Fang Yi Ke Da Xue Xue Bao ; 40(7): 936-941, 2020 Jul 30.
Artigo em Chinês | MEDLINE | ID: mdl-32895148

RESUMO

OBJECTIVE: To observe the expression of HELQ and RAD51C in normal endometrial and endometrial stromal sarcoma (ESS) and analyze their correlation with the clinical features of the patients. METHODS: The expressions of HELQ and RAD51C proteins were detected by immunohistochemical staining in normal endometrial tissues (14 cases) and tumor tissues from patients with ESS (37 cases) treated in Hunan Provincial Cancer Hospital from January, 2013 to December, 2016. The correlations of the expressions of the two proteins with the patients'age, FIGO staging, tissue type, tumor size, and lymph node metastasis were analyzed. RESULTS: Immunohistochemical staining showed that the expressions of HELQ and RAD51C were both decreased in ESS patients compared with the normal group, and there was a positive correlation between HELQ and RAD51C expression (P < 0.05). HELQ expression in ESS was correlated with the tumor size and type. The expressions of HELQ and RAD51C were not correlated with the patients' age, FIGO stage and status of lymph node metastasis (P > 0.05). CONCLUSIONS: Homologous recombination- directed DNA repair involving HELQ and RAD51C may participate in the occurrence and progression of ESS.


Assuntos
Neoplasias do Endométrio , Sarcoma do Estroma Endometrial , DNA Helicases , Proteínas de Ligação a DNA , Endométrio , Feminino , Humanos , Metástase Linfática
19.
Mol Cell ; 79(3): 362-364, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32763223

RESUMO

In this issue of Molecular Cell, Wang et al. (2020) investigate stress-induced nuclear condensates of the RNA-binding protein TDP-43, uncovering a protective function for these granules as well as an RNA-dependent mechanism for scaffolding them.


Assuntos
Esclerose Amiotrófica Lateral , RNA Longo não Codificante , Proteínas de Ligação a DNA , Humanos , Proteínas de Ligação a RNA
20.
Mol Cell ; 79(3): 365-367, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32763224

RESUMO

In a recent issue of Molecular Cell, Wang et al. (2020) employ unbiased proteomics approaches and live-cell imaging to reveal a key role for the histone chaperone complex FACT (SPT16 and SSRP1) in governing Cas9 turnover at the DNA target site during genome and epigenome editing.


Assuntos
Edição de Genes , RNA Longo não Codificante , Sistemas CRISPR-Cas , Proteínas de Ligação a DNA , Histonas
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