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1.
PLoS Comput Biol ; 16(9): e1008229, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32936825

RESUMO

Accurately predicting essential genes using computational methods can greatly reduce the effort in finding them via wet experiments at both time and resource scales, and further accelerate the process of drug discovery. Several computational methods have been proposed for predicting essential genes in model organisms by integrating multiple biological data sources either via centrality measures or machine learning based methods. However, the methods aiming to predict human essential genes are still limited and the performance still need improve. In addition, most of the machine learning based essential gene prediction methods are lack of skills to handle the imbalanced learning issue inherent in the essential gene prediction problem, which might be one factor affecting their performance. We propose a deep learning based method, DeepHE, to predict human essential genes by integrating features derived from sequence data and protein-protein interaction (PPI) network. A deep learning based network embedding method is utilized to automatically learn features from PPI network. In addition, 89 sequence features were derived from DNA sequence and protein sequence for each gene. These two types of features are integrated to train a multilayer neural network. A cost-sensitive technique is used to address the imbalanced learning problem when training the deep neural network. The experimental results for predicting human essential genes show that our proposed method, DeepHE, can accurately predict human gene essentiality with an average performance of AUC higher than 94%, the area under precision-recall curve (AP) higher than 90%, and the accuracy higher than 90%. We also compare DeepHE with several widely used traditional machine learning models (SVM, Naïve Bayes, Random Forest, and Adaboost) using the same features and utilizing the same cost-sensitive technique to against the imbalanced learning issue. The experimental results show that DeepHE significantly outperforms the compared machine learning models. We have demonstrated that human essential genes can be accurately predicted by designing effective machine learning algorithm and integrating representative features captured from available biological data. The proposed deep learning framework is effective for such task.


Assuntos
Biologia Computacional/métodos , Aprendizado Profundo , Genes Essenciais/genética , Análise de Sequência de DNA/métodos , DNA/genética , Humanos , Redes Neurais de Computação , Mapas de Interação de Proteínas/genética
2.
PLoS One ; 15(8): e0238322, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866178

RESUMO

Space-filling curves have been used for decades to study the folding principles of globular proteins, compact polymers, and chromatin. Formally, space-filling curves trace a single circuit through a set of points (x,y,z); informally, they correspond to a polymer melt. Although not quite a melt, the folding principles of Human chromatin are likened to the Hilbert curve: a type of space-filling curve. Hilbert-like curves in general make biologically compelling models of chromatin; in particular, they lack knots which facilitates chromatin folding, unfolding, and easy access to genes. Knot complexity has been intensely studied with the aid of Alexander polynomials; however, the approach does not generalize well to cases of more than one chromosome. Crossing complexity is an understudied alternative better suited for quantifying entanglement between chromosomes. Do Hilbert-like configurations limit crossing complexity between chromosomes? How does crossing complexity for Hilbert-like configurations compare to equilibrium configurations? To address these questions, we extend the Mansfield algorithm to enable sampling of Hilbert-like space filling curves on a simple cubic lattice. We use the extended algorithm to generate equilibrium, intermediate, and Hilbert-like configurational ensembles and compute crossing complexity between curves (chromosomes) in each configurational snapshot. Our main results are twofold: (a) Hilbert-like configurations limit entanglement between chromosomes and (b) Hilbert-like configurations do not limit entanglement in a model of S-phase DNA. Our second result is particularly surprising yet easily rationalized with a geometric argument. We explore ergodicity of the extended algorithm and discuss our results in the context of more sophisticated models of chromatin.


Assuntos
DNA/química , DNA/genética , Fase S/genética , Algoritmos , Cromatina/química , Cromatina/genética , Cromossomos/química , Cromossomos/genética , Humanos , Polímeros/química
3.
Mol Cell ; 79(6): 1037-1050.e5, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32882183

RESUMO

DNA double-stranded breaks (DSBs) are dangerous lesions threatening genomic stability. Fidelity of DSB repair is best achieved by recombination with a homologous template sequence. In yeast, transcript RNA was shown to template DSB repair of DNA. However, molecular pathways of RNA-driven repair processes remain obscure. Utilizing assays of RNA-DNA recombination with and without an induced DSB in yeast DNA, we characterize three forms of RNA-mediated genomic modifications: RNA- and cDNA-templated DSB repair (R-TDR and c-TDR) using an RNA transcript or a DNA copy of the RNA transcript for DSB repair, respectively, and a new mechanism of RNA-templated DNA modification (R-TDM) induced by spontaneous or mutagen-induced breaks. While c-TDR requires reverse transcriptase, translesion DNA polymerase ζ (Pol ζ) plays a major role in R-TDR, and it is essential for R-TDM. This study characterizes mechanisms of RNA-DNA recombination, uncovering a role of Pol ζ in transferring genetic information from transcript RNA to DNA.


Assuntos
DNA/genética , RNA/genética , Saccharomyces cerevisiae/genética , Adolescente , Adulto , DNA/ultraestrutura , Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , Replicação do DNA/genética , DNA Complementar/genética , DNA Polimerase Dirigida por DNA/genética , DNA Polimerase Dirigida por DNA/ultraestrutura , Instabilidade Genômica/genética , Humanos , Pessoa de Meia-Idade , RNA/ultraestrutura , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Adulto Jovem
4.
Nat Commun ; 11(1): 4758, 2020 09 21.
Artigo em Inglês | MEDLINE | ID: mdl-32958811

RESUMO

Genetic programs operating in a history-dependent fashion are ubiquitous in nature and govern sophisticated processes such as development and differentiation. The ability to systematically and predictably encode such programs would advance the engineering of synthetic organisms and ecosystems with rich signal processing abilities. Here we implement robust, scalable history-dependent programs by distributing the computational labor across a cellular population. Our design is based on standardized recombinase-driven DNA scaffolds expressing different genes according to the order of occurrence of inputs. These multicellular computing systems are highly modular, do not require cell-cell communication channels, and any program can be built by differential composition of strains containing well-characterized logic scaffolds. We developed automated workflows that researchers can use to streamline program design and optimization. We anticipate that the history-dependent programs presented here will support many applications using cellular populations for material engineering, biomanufacturing and healthcare.


Assuntos
Modelos Genéticos , Biologia Sintética/métodos , Fenômenos Fisiológicos Celulares/genética , DNA/genética , DNA/metabolismo , Lógica , Recombinases/genética , Recombinases/metabolismo , Software , Fluxo de Trabalho
5.
Adv Exp Med Biol ; 1255: 1-6, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32949386

RESUMO

Clinical single-cell biomedicine has become a new emerging discipline, which integrates single-cell RNA and DNA sequencing, proteomics, and functions with clinical phenomes, therapeutic responses, and prognosis. It is of great value to discover disease-, phenome-, and therapy-specific diagnostic biomarkers and therapeutic targets on the basis of the principle of clinical single-cell biomedicine. This book reviews the roles of single-cell sequencing and methylation in diseases and explores disease-specific alterations of single-cell sequencing and methylation, especially focusing on potential applications of methodologies on human single-cell sequencing and methylation, on potential correlations between those changes with pulmonary diseases, and on potential roles of signaling pathways that cause heterogeneous cellular responses during treatment. This book also emphasizes the importance of methodologies in clinical practice and application, the potential of perspectives, challenges and solutions, and the significance of single-cell preparation standardization. Alterations of DNA and RNA methylation, demethylation in lung diseases, and a deep knowledge about the regulation and function of target gene methylation for diagnosing and treating diseases at the early stage are also provided. Importantly, this book aims to apply the measurement of single-cell sequencing and methylation for clinical diagnosis and treatment and to understand clinical values of those parameters and to headline and foresee the potential values of the application of single-cell sequencing in non-cancer diseases.


Assuntos
Metilação de DNA , Doença/genética , Análise de Sequência , Análise de Célula Única , DNA/genética , DNA/metabolismo , Humanos , Proteômica , RNA/genética , RNA/metabolismo
6.
Nucleic Acids Res ; 48(15): 8529-8544, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32738045

RESUMO

Myocyte enhancer factor-2B (MEF2B) has the unique capability of binding to its DNA target sites with a degenerate motif, while still functioning as a gene-specific transcriptional regulator. Identifying its DNA targets is crucial given regulatory roles exerted by members of the MEF2 family and MEF2B's involvement in B-cell lymphoma. Analyzing structural data and SELEX-seq experimental results, we deduced the DNA sequence and shape determinants of MEF2B target sites on a high-throughput basis in vitro for wild-type and mutant proteins. Quantitative modeling of MEF2B binding affinities and computational simulations exposed the DNA readout mechanisms of MEF2B. The resulting binding signature of MEF2B revealed distinct intricacies of DNA recognition compared to other transcription factors. MEF2B uses base readout at its half-sites combined with shape readout at the center of its degenerate motif, where A-tract polarity dictates nuances of binding. The predominant role of shape readout at the center of the core motif, with most contacts formed in the minor groove, differs from previously observed protein-DNA readout modes. MEF2B, therefore, represents a unique protein for studies of the role of DNA shape in achieving binding specificity. MEF2B-DNA recognition mechanisms are likely representative for other members of the MEF2 family.


Assuntos
Proteínas de Ligação a DNA/ultraestrutura , DNA/ultraestrutura , Complexos Multiproteicos/ultraestrutura , Sequência de Aminoácidos/genética , Sítios de Ligação/genética , DNA/genética , Proteínas de Ligação a DNA/química , Humanos , Linfoma de Células B/genética , Linfoma de Células B/patologia , Proteínas de Domínio MADS/genética , Proteínas de Domínio MADS/ultraestrutura , Fatores de Transcrição MEF2/química , Fatores de Transcrição MEF2/ultraestrutura , Complexos Multiproteicos/genética , Conformação de Ácido Nucleico , Motivos de Nucleotídeos/genética , Ligação Proteica/genética
7.
Phys Rev Lett ; 125(4): 048104, 2020 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-32794805

RESUMO

The RNA world scenario posits replication by RNA polymerases. On early Earth, a geophysical setting is required to separate hybridized strands after their replication and to localize them against diffusion. We present a pointed heat source that drives exponential, RNA-catalyzed amplification of short RNA with high efficiency in a confined chamber. While shorter strands were periodically melted by laminar convection, the temperature gradient caused aggregated polymerase molecules to accumulate, protecting them from degradation in hot regions of the chamber. These findings demonstrate a size-selective pathway for autonomous RNA-based replication in natural nonequilibrium conditions.


Assuntos
Ecossistema , RNA/química , RNA/genética , Catálise , DNA/química , DNA/genética , DNA/metabolismo , Replicação do DNA , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/metabolismo , Planeta Terra , Evolução Molecular , Temperatura Alta , Biossíntese de Proteínas/genética , RNA/metabolismo
8.
Proc Natl Acad Sci U S A ; 117(33): 19661-19663, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32747537

RESUMO

The structural unit of eukaryotic chromatin is a nucleosome, comprising two histone H2A-H2B heterodimers and one histone (H3-H4)2 tetramer, wrapped around by ∼146 bp of DNA. The N-terminal flexible histone tails stick out from the histone core and have extensive posttranslational modifications, causing epigenetic changes of chromatin. Although crystal and cryogenic electron microscopy structures of nucleosomes are available, the flexible tail structures remain elusive. Using NMR, we have examined the dynamics of histone H3 tails in nucleosomes containing unmodified and tetra-acetylated H4 tails. In unmodified nucleosome, the H3 tail adopts a dynamic equilibrium structure between DNA-contact and reduced-contact states. In acetylated H4 nucleosome, however, the H3 tail equilibrium shifts to a mainly DNA-contact state with a minor reduced-contact state. The acetylated H4 tail is dynamically released from its own DNA-contact state to a reduced-contact state, while the H3 tail DNA-contact state becomes major. Notably, H3 K14 in the acetylated H4 nucleosome is much more accessible to acetyltransferase Gcn5 relative to unmodified nucleosome, possibly due to the formation of a favorable H3 tail conformation for Gcn5. In summary, each histone tail adopts a characteristic dynamic state but regulates one other, probably creating a histone tail network even on a nucleosome.


Assuntos
Histonas/química , Histonas/metabolismo , Nucleossomos/metabolismo , Acetilação , Motivos de Aminoácidos , DNA/genética , DNA/metabolismo , Histonas/genética , Humanos , Conformação de Ácido Nucleico , Nucleossomos/genética
9.
Mol Cell ; 79(6): 881-901, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32768408

RESUMO

Nucleosomes package genomic DNA into chromatin. By regulating DNA access for transcription, replication, DNA repair, and epigenetic modification, chromatin forms the nexus of most nuclear processes. In addition, dynamic organization of chromatin underlies both regulation of gene expression and evolution of chromosomes into individualized sister objects, which can segregate cleanly to different daughter cells at anaphase. This collaborative review shines a spotlight on technologies that will be crucial to interrogate key questions in chromatin and chromosome biology including state-of-the-art microscopy techniques, tools to physically manipulate chromatin, single-cell methods to measure chromatin accessibility, computational imaging with neural networks and analytical tools to interpret chromatin structure and dynamics. In addition, this review provides perspectives on how these tools can be applied to specific research fields such as genome stability and developmental biology and to test concepts such as phase separation of chromatin.


Assuntos
Cromatina/genética , Cromossomos/genética , DNA/genética , Nucleossomos/genética , Reparo do DNA/genética , Replicação do DNA/genética , Epigênese Genética/genética , Humanos
10.
Arch Virol ; 165(10): 2397-2400, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32748177

RESUMO

Enterobacter aerogenes is a member of the ESKAPE group of bacteria, and multi-drug-resistant strains are increasingly being found. In this study, a novel bacteriophage, ATCEA85, which infects E. aerogenes, has been isolated and characterized. ATCEA85 is seen to have a circularly permuted linear double-stranded DNA genome of 47,484 base pairs in length. The closest related phage found in the databases is the Klebsiella phage Kp3, which exhibits 77% identity over a 34% query coverage. The G+C content of ATCEA85 is 56.2%, and 15 putative open reading frames are functionally annotated.


Assuntos
DNA Viral/genética , Enterobacter aerogenes/virologia , Genoma Viral , Fases de Leitura Aberta , Filogenia , Siphoviridae/genética , Composição de Bases , DNA/genética , Ontologia Genética , Anotação de Sequência Molecular , Siphoviridae/classificação , Siphoviridae/isolamento & purificação , Sequenciamento Completo do Genoma
12.
Mol Cell ; 79(6): 917-933.e9, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32755595

RESUMO

Despite key roles in sister chromatid cohesion and chromosome organization, the mechanism by which cohesin rings are loaded onto DNA is still unknown. Here we combine biochemical approaches and cryoelectron microscopy (cryo-EM) to visualize a cohesin loading intermediate in which DNA is locked between two gates that lead into the cohesin ring. Building on this structural framework, we design experiments to establish the order of events during cohesin loading. In an initial step, DNA traverses an N-terminal kleisin gate that is first opened upon ATP binding and then closed as the cohesin loader locks the DNA against the ATPase gate. ATP hydrolysis will lead to ATPase gate opening to complete DNA entry. Whether DNA loading is successful or results in loop extrusion might be dictated by a conserved kleisin N-terminal tail that guides the DNA through the kleisin gate. Our results establish the molecular basis for cohesin loading onto DNA.


Assuntos
Proteínas de Ciclo Celular/ultraestrutura , Cromátides/ultraestrutura , Proteínas Cromossômicas não Histona/ultraestrutura , DNA/ultraestrutura , Troca de Cromátide Irmã/genética , Adenosina Trifosfatases/genética , Proteínas de Ciclo Celular/genética , Cromátides/genética , Proteínas Cromossômicas não Histona/genética , Segregação de Cromossomos/genética , Microscopia Crioeletrônica , DNA/genética , Conformação de Ácido Nucleico , Conformação Proteica , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/ultraestrutura
13.
PLoS One ; 15(8): e0236709, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32790736

RESUMO

BACKGROUND: With the development of second-generation sequencing technology, more and more DNA sequence variations have been detected. Exon sequencing is the first choice for sequencing many cancer genes, and it can be better used to identify disease status by detecting gene variants. PCR sequence is an effective method to capture that sequence of an exon in the process of sequencing. Exon sequencing sequence contains PCR primer sequence, the correct position of the sequence can be determined by PCR primer sequence, which can be found in SNP, Indel mutation point by comparing the sequence of PCR primer sequence. RESULTS: In this paper, a matching algorithm based on the PCR primer sequence is proposed, which can effectively sequence the position of PCR primer sequence and find out the key position sequence. Then the sequencing sequence is sorted and the number of the same sequence is counted to reduce the matching times. Then, the sequenced sequence was matched with PCR primer sequence, so that the DNA position could be accurately matched and the variation in the sequenced sequence could be found more quickly. CONCLUSIONS: Compared with the traditional sequence matching method, PCR primer sequence matching method can match many sequences and find more variation. It also showed a high recall rate in the recall rate.


Assuntos
Algoritmos , Reação em Cadeia da Polimerase/métodos , DNA/química , DNA/genética , DNA/metabolismo , Primers do DNA/metabolismo , Éxons , Humanos , Mutação INDEL , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA
14.
Yakugaku Zasshi ; 140(8): 993-1000, 2020.
Artigo em Japonês | MEDLINE | ID: mdl-32741873

RESUMO

The human genome consists of more than 20000 genes and is essential for all biological phenomena. To understand these biological phenomena, including diseases, and to be able to modify them, approaches that enable optical control of the genome may be useful. Recently, we developed an optogenetic tool, named photoactivatable Cas9 (PA-Cas9). We divided Cas9 nuclease from the CRISPR-Cas9 system into two fragments and connected photo-inducible dimerization proteins, named Magnet system, to the fragments, leading to the development of PA-Cas9 of which nuclease activity is switchable with light. PA-Cas9 allows direct editing of DNA sequences by light stimulation. Additionally, we developed a light-inducible, RNA-guided programmable system for endogenous gene activation based on the CRISPR-Cas9 system. We demonstrated that this optogenetic tool allows rapid and reversible targeted gene activation by light. Using this tool, we exemplified optical control of neuronal differentiation of human induced pluripotent stem cells (iPSCs). The CRISPR-Cas9-based, photoactivatable transcription system offers a simple and versatile approach to precise gene activation. In addition to the CRISPR-Cas9-based optogenetic tools, we developed a photoactivatable Cre-loxP system. This tool allows optical control of DNA recombination reaction in an internal organ even by external, noninvasive illumination using LED light source. To date, genome engineering technology and optogenetics technology have emerged separately as different applications. Our studies described above merge these emerging research fields together.


Assuntos
Proteína 9 Associada à CRISPR , Sistemas CRISPR-Cas , Engenharia Genética , Luz , Optogenética , Ativação Transcricional , Animais , Diferenciação Celular , DNA/genética , Edição de Genes , Genoma Humano , Humanos , Células-Tronco Pluripotentes Induzidas/fisiologia , Camundongos , Recombinação Genética
15.
Nat Protoc ; 15(9): 3030-3063, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32807909

RESUMO

Materials that sense and respond to biological signals in their environment have a broad range of potential applications in drug delivery, medical devices and diagnostics. Nucleic acids are important biological cues that encode information about organismal identity and clinically relevant phenotypes such as drug resistance. We recently developed a strategy to design nucleic acid-responsive materials using the CRISPR-associated nuclease Cas12a as a user-programmable sensor and material actuator. This approach improves on the sensitivity of current DNA-responsive materials while enabling their rapid repurposing toward new sequence targets. Here, we provide a comprehensive resource for the design, synthesis and actuation of CRISPR-responsive hydrogels. First, we provide guidelines for the synthesis of Cas12a guide RNAs (gRNAs) for in vitro applications. We then outline methods for the synthesis of both polyethylene glycol-DNA (PEG-DNA) and polyacrylamide-DNA (PA-DNA) hydrogels, as well as their controlled degradation using Cas12a for the release of cargos, including small molecules, enzymes, nanoparticles and living cells within hours. Finally, we detail the design and assembly of microfluidic paper-based devices that use Cas12a-sensitive hydrogels to convert DNA inputs into a variety of visual and electronic readouts for use in diagnostics. Following the initial validation of the gRNA and Cas12a components (1 d), the synthesis and testing of either PEG-DNA or PA-DNA hydrogels require 3-4 d of laboratory time. Optional extensions, including the release of primary human cells or the design of the paper-based diagnostic, require an additional 2-3 d each.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Técnicas e Procedimentos Diagnósticos , Sistemas de Liberação de Medicamentos/métodos , Liberação Controlada de Fármacos , Materiais Inteligentes/química , Resinas Acrílicas/química , Proteínas de Bactérias/metabolismo , Sequência de Bases , Proteínas Associadas a CRISPR/metabolismo , DNA/química , DNA/genética , Endodesoxirribonucleases/metabolismo , Humanos , Células K562 , Polietilenoglicóis/química , RNA Guia/genética
16.
Nat Commun ; 11(1): 4072, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792663

RESUMO

Cpf1-linked base editors broaden the targeting scope of programmable cytidine deaminases by recognizing thymidine-rich protospacer-adjacent motifs (PAM) without inducing DNA double-strand breaks (DSBs). Here we present an unbiased in vitro method for identifying genome-wide off-target sites of Cpf1 base editors via whole genome sequencing. First, we treat human genomic DNA with dLbCpf1-BE ribonucleoprotein (RNP) complexes, which convert C-to-U at on-target and off-target sites and, then, with a mixture of E. coli uracil DNA glycosylase (UDG) and DNA glycosylase-lyase Endonuclease VIII, which removes uracil and produces single-strand breaks (SSBs) in vitro. Whole-genome sequencing of the resulting digested genome (Digenome-seq) reveals that, on average, dLbCpf1-BE induces 12 SSBs in vitro per crRNA in the human genome. Off-target sites with an editing frequency as low as 0.1% are successfully identified by this modified Digenome-seq method, demonstrating its high sensitivity. dLbCpf1-BEs and LbCpf1 nucleases often recognize different off-target sites, calling for independent analysis of each tool.


Assuntos
Citidina/metabolismo , Endonucleases/metabolismo , Sequenciamento Completo do Genoma/métodos , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Citidina/genética , DNA/genética , DNA/metabolismo , Endonucleases/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Edição de Genes , Genoma Humano/genética , Células HEK293 , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , RNA Guia/genética
17.
Nat Commun ; 11(1): 3839, 2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32737294

RESUMO

Chromatin regulates spatiotemporal gene expression during neurodevelopment, but it also mediates DNA damage repair essential to proliferating neural progenitor cells (NPCs). Here, we uncover molecularly dissociable roles for nucleosome remodeler Ino80 in chromatin-mediated transcriptional regulation and genome maintenance in corticogenesis. We find that conditional Ino80 deletion from cortical NPCs impairs DNA double-strand break (DSB) repair, triggering p53-dependent apoptosis and microcephaly. Using an in vivo DSB repair pathway assay, we find that Ino80 is selectively required for homologous recombination (HR) DNA repair, which is mechanistically distinct from Ino80 function in YY1-associated transcription. Unexpectedly, sensitivity to loss of Ino80-mediated HR is dependent on NPC division mode: Ino80 deletion leads to unrepaired DNA breaks and apoptosis in symmetric NPC-NPC divisions, but not in asymmetric neurogenic divisions. This division mode dependence is phenocopied following conditional deletion of HR gene Brca2. Thus, distinct modes of NPC division have divergent requirements for Ino80-dependent HR DNA repair.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/genética , Proteína BRCA2/genética , Cromatina/química , Proteínas de Ligação a DNA/genética , Células-Tronco Neurais/metabolismo , Neurogênese/genética , Reparo de DNA por Recombinação , ATPases Associadas a Diversas Atividades Celulares/deficiência , Animais , Apoptose/genética , Proteína BRCA2/deficiência , Divisão Celular , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , DNA/genética , DNA/metabolismo , Quebras de DNA de Cadeia Dupla , Proteínas de Ligação a DNA/deficiência , Embrião de Mamíferos , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Camundongos Transgênicos , Neocórtex/citologia , Neocórtex/crescimento & desenvolvimento , Neocórtex/metabolismo , Células-Tronco Neurais/citologia , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Fator de Transcrição YY1/genética , Fator de Transcrição YY1/metabolismo
18.
Nat Commun ; 11(1): 3831, 2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32737305

RESUMO

Long-term time series have provided evidence that anthropogenic pressures can threaten lakes. Yet it remains unclear how and the extent to which lake biodiversity has changed during the Anthropocene, in particular for microbes. Here, we used DNA preserved in sediments to compare modern micro-eukaryotic communities with those from the end of the 19th century, i.e., before acceleration of the human imprint on ecosystems. Our results obtained for 48 lakes indicate drastic changes in the composition of microbial communities, coupled with a homogenization of their diversity between lakes. Remote high elevation lakes were globally less impacted than lowland lakes affected by local human activity. All functional groups (micro-algae, parasites, saprotrophs and consumers) underwent significant changes in diversity. However, we show that the effects of anthropogenic changes have benefited in particular phototrophic and mixotrophic species, which is consistent with the hypothesis of a global increase of primary productivity in lakes.


Assuntos
DNA/genética , Eucariotos/genética , Sedimentos Geológicos/análise , Lagos/análise , Alveolados/classificação , Alveolados/genética , Alveolados/isolamento & purificação , Biodiversidade , Evolução Biológica , Ecossistema , Eucariotos/classificação , Eucariotos/isolamento & purificação , História do Século XIX , História do Século XX , História do Século XXI , Atividades Humanas/história , Humanos , Microalgas/classificação , Microalgas/genética , Microalgas/isolamento & purificação , Microbiota/genética , Processos Fototróficos/fisiologia , Rhizaria/classificação , Rhizaria/genética , Rhizaria/isolamento & purificação , Estramenópilas/classificação , Estramenópilas/genética , Estramenópilas/isolamento & purificação
19.
Biosens Bioelectron ; 167: 112479, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32763826

RESUMO

COVID-19 pandemic outbreak is the most astounding scene ever experienced in the 21st century. It has been determined to be caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). With the global pandemic, the lack of efficient rapid and accurate molecular diagnostic testing tools has hindered the public opportunely response to the emerging viral threat. Herein, a DNA nanoscaffold hybrid chain reaction (DNHCR)-based nucleic acid assay strategy is reported for rapid detection of SARS-CoV-2 RNA. In this method, the DNA nanoscaffolds have been first constructed by the self-assembly of long DNA strands and self-quenching probes (H1). Then, the SARS-CoV-2 RNA will initiate the hybridization of H1 and free H2 DNA probes along the nanoscaffold, and an illuminated DNA nanostring is instantly obtained. By taking advantages of the localization design of the H1 probes and the temperature tolerance of the isothermal amplification, the proposed DNHCR method can detect target at short responding time (within 10 min) and mild condition (15 °C-35 °C). Moreover, the reliability of DNHCR method in serum and saliva samples have also been validated. Therefore, DNHCR-based method is expected to provide a simple and faster alternative to the traditional SARS-CoV-2 qRT-PCR assay.


Assuntos
Betacoronavirus , Técnicas Biossensoriais/métodos , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/virologia , Pneumonia Viral/diagnóstico , Pneumonia Viral/virologia , Betacoronavirus/genética , Betacoronavirus/isolamento & purificação , Técnicas de Laboratório Clínico/estatística & dados numéricos , Infecções por Coronavirus/epidemiologia , DNA/síntese química , DNA/química , DNA/genética , Estudos de Viabilidade , Humanos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/estatística & dados numéricos , Nanoestruturas/química , Nanotecnologia , Técnicas de Amplificação de Ácido Nucleico/métodos , Pandemias , Pneumonia Viral/epidemiologia , RNA Viral/análise , RNA Viral/genética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
20.
PLoS One ; 15(8): e0231683, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32764752

RESUMO

Aquatic macroinvertebrates play a crucial role in freshwater ecosystems, but their diversity remains poorly known, particularly in the tropics. This "taxonomic void" limits our understanding of biodiversity patterns and processes in freshwater ecosystems, and the scale at which they operate. We used DNA barcoding to estimate lineage diversity (and the diversity of unique haplotypes) in 224 specimens of freshwater macroinvertebrates at a small spatial scale within the Panama Canal Watershed (PCW). In addition, we compiled available barcoding data to assess macroinvertebrate diversity at a broader spatial scale spanning the Isthmus of Panama. Consistently across two species delimitation algorithms (i.e., ABGD and GMYC), we found high lineage diversity within the PCW, with ~ 100-106 molecular operational taxonomic units (MOTUs) across 168 unique haplotypes. We also found a high lineage diversity along the Isthmus of Panama, but this diversity peaked within the PCW. However, our rarefaction/extrapolation approach showed that this diversity remains under-sampled. As expected, these results indicate that the diversity of Neotropical freshwater macroinvertebrates is higher than previously thought, with the possibility of high endemicity even at narrow spatial scales. Consistent with previous work on aquatic insects and other freshwater taxa in this region, geographic isolation is likely a main factor shaping these patterns of diversity. However, other factors such as habitat variability and perhaps local adaptation might be reshaping these patterns of diversity at a local scale. Although further research is needed to better understand the processes driving diversification in freshwater macroinvertebrates, we suggest that Neotropical streams hold a high proportion of hidden biodiversity. Understanding this diversity is crucial in the face of increasing human disturbance.


Assuntos
Biologia de Ecossistemas de Água Doce/métodos , Insetos/classificação , Invertebrados/genética , Animais , Biodiversidade , DNA/genética , Código de Barras de DNA Taxonômico/métodos , Ecossistema , Água Doce , Insetos/genética , Panamá , Zona do Canal do Panamá , Filogenia , Rios
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