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1.
Adv Exp Med Biol ; 1158: 247-255, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31452144

RESUMO

The maternally inherited mitochondrial DNA (mtDNA) is located inside every mitochondrion, in variable number of copies, and it contains 37 crucial genes for cellular bioenergetics. This chapter will discuss the unique features of this circular genome including heteroplasmy, haplogroups, among others, along with the corresponding clinical relevance for each. The discussion also covers the nuclear-encoded mitochondrial genes (N > 1000) and the epistatic interactions between mtDNA and the nuclear genome. Examples of mitochondrial diseases related to specific mtDNA mutation sites of relevance for humans are provided. This chapter aims to provide an overview of mitochondrial genetics as an emerging hot topic for the future of medicine.


Assuntos
Metabolismo Energético , Mitocôndrias , DNA Mitocondrial/genética , Metabolismo Energético/genética , Epistasia Genética , Genes Mitocondriais/genética , Genoma/genética , Humanos , Mitocôndrias/genética , Doenças Mitocondriais/genética , Mutação
2.
Nat Commun ; 10(1): 2756, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31227702

RESUMO

Flight loss in birds is as characteristic of the class Aves as flight itself. Although morphological and physiological differences are recognized in flight-degenerate bird species, their contributions to recurrent flight degeneration events across modern birds and underlying genetic mechanisms remain unclear. Here, in an analysis of 295 million nucleotides from 48 bird genomes, we identify two convergent sites causing amino acid changes in ATGLSer321Gly and ACOT7Ala197Val in flight-degenerate birds, which to our knowledge have not previously been implicated in loss of flight. Functional assays suggest that Ser321Gly reduces lipid hydrolytic ability of ATGL, and Ala197Val enhances acyl-CoA hydrolytic activity of ACOT7. Modeling simulations suggest a switch of main energy sources from lipids to carbohydrates in flight-degenerate birds. Our results thus suggest that physiological convergence plays an important role in flight degeneration, and anatomical convergence often invoked may not.


Assuntos
Evolução Biológica , Aves/fisiologia , Metabolismo Energético/genética , Voo Animal/fisiologia , Genoma/genética , Animais , Metabolismo dos Carboidratos/fisiologia , Genômica/métodos , Lipase/genética , Lipase/metabolismo , Lipólise/fisiologia , Palmitoil-CoA Hidrolase/genética , Palmitoil-CoA Hidrolase/metabolismo , Filogenia
3.
Cancer Sci ; 110(8): 2328-2336, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31228211

RESUMO

Changes of nuclear localization of lineage-specific genes from a transcriptionally inert to permissive environment are a crucial step in establishing the identity of a cell. Noncoding RNA transcription-mediated genome folding and activation of target gene expression have been found in a variety of cell types. Noncoding RNA ThymoD (thymocyte differentiation factor) transcription at superenhancers is essential for mouse T-cell lineage commitment. The cessation of ThymoD transcription abolishes transcription-mediated demethylation, recruiting looping factors such as the cohesin complex, CCCTC-binding factor (CTCF), ultimately leading to the phenotype of severe combined immunodeficiency and T-cell leukemia/lymphoma. In this review, we describe the functional role of RNA polymerase II-mediated transcription at enhancers and in genome folding. We also highlight the involvement of faulty activation or suppression of enhancer transcription and enhancer-promoter interaction in cancer development.


Assuntos
Elementos Facilitadores Genéticos/genética , Genoma/genética , Neoplasias/genética , RNA não Traduzido/genética , Transcrição Genética/genética , Animais , Humanos , Regiões Promotoras Genéticas/genética
4.
Nat Chem ; 11(7): 629-637, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31209299

RESUMO

In DNA, the loss of a nucleobase by hydrolysis generates an abasic site. Formed as a result of DNA damage, as well as a key intermediate during the base excision repair pathway, abasic sites are frequent DNA lesions that can lead to mutations and strand breaks. Here we present snAP-seq, a chemical approach that selectively exploits the reactive aldehyde moiety at abasic sites to reveal their location within DNA at single-nucleotide resolution. Importantly, the approach resolves abasic sites from other aldehyde functionalities known to exist in genomic DNA. snAP-seq was validated on synthetic DNA and then applied to two separate genomes. We studied the distribution of thymine modifications in the Leishmania major genome by enzymatically converting these modifications into abasic sites followed by abasic site mapping. We also applied snAP-seq directly to HeLa DNA to provide a map of endogenous abasic sites in the human genome.


Assuntos
DNA/genética , Genoma/genética , Análise de Sequência de DNA/métodos , Aldeídos/química , Sequência de Bases , DNA/química , Dano ao DNA/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Leishmania major/genética , Sondas Moleculares/síntese química , Sondas Moleculares/química , Timina/química , Uracila-DNA Glicosidase/química
5.
J Anim Sci ; 97(7): 2793-2802, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31087081

RESUMO

The objectives of this study were to identify informative genomic regions that affect the exterior traits of purebred Korean Yorkshire pigs and to investigate and compare the accuracy of genomic prediction for response variables. Phenotypic data on body height (BH), body length (BL), and total teat number (TTN) from 2,432 Yorkshire pigs were used to obtain breeding values including as response variable the estimated breeding value (EBV) and 2 types of deregressed EBVs-one including the parent average (DEBVincPA) and the other excluding it (DEBVexcPA). A final genotype panel comprising 46,199 SNP markers was retained for analysis after quality control for common SNPs. The BayesB and BayesC methods-with various π and weighted response variables (EBV, DEBVincPA, or DEBVexcPA)-were used to estimate SNP effects, through the genome-wide association study. The significance of genomic windows (1 Mb) was obtained at 1.0% additive genetic variance and was subsequently used to identify informative genomic regions. Furthermore, SNPs with a high model frequency (≥0.90) were considered informative. The accuracy of genomic prediction was estimated using a 5-fold cross-validation with the K-means clustering method. Genomic accuracy was measured as the genomic correlation between the molecular breeding value and the individual weighted response variables (EBV, DEBVincPA, or DEBVexcPA). The number of identified informative windows (1 Mb) for BH, BL, and TTN was 4, 3, and 4, respectively. The number of significant SNPs for BH, BL, and TTN was 6, 4, and 5, respectively. Diversity π did not influence the accuracy of genomic prediction. The BayesB method showed slightly higher genomic accuracy for exterior traits than BayesC method in this study. In addition, the genomic accuracy using DEBVincPA as response variable was higher than that using other response variables. Therefore, the genomic accuracy using BayesB (π = 0.90) with DEBVinPA as a response variable was the most effective in this study. The genomic accuracy values for BH, BL, and TTN were calculated to be 0.52, 0.60, and 0.51, respectively.


Assuntos
Estudo de Associação Genômica Ampla/veterinária , Genoma/genética , Genômica , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , Suínos/genética , Animais , Cruzamento , Análise por Conglomerados , Confiabilidade dos Dados , Feminino , Marcadores Genéticos/genética , Testes Genéticos/veterinária , Genótipo , Masculino , Glândulas Mamárias Animais/anatomia & histologia , Fenótipo , Suínos/anatomia & histologia
6.
Genes Dev ; 33(15-16): 1008-1026, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31123061

RESUMO

Genome replication involves dealing with obstacles that can result from DNA damage but also from chromatin alterations, topological stress, tightly bound proteins or non-B DNA structures such as R loops. Experimental evidence reveals that an engaged transcription machinery at the DNA can either enhance such obstacles or be an obstacle itself. Thus, transcription can become a potentially hazardous process promoting localized replication fork hindrance and stress, which would ultimately cause genome instability, a hallmark of cancer cells. Understanding the causes behind transcription-replication conflicts as well as how the cell resolves them to sustain genome integrity is the aim of this review.


Assuntos
Replicação do DNA/fisiologia , Instabilidade Genômica/genética , Transcrição Genética/fisiologia , Genoma/genética , Humanos , Neoplasias/fisiopatologia , Elongação da Transcrição Genética/fisiologia
7.
Molecules ; 24(9)2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31067825

RESUMO

G-quadruplex (G4) structures are highly stable four-stranded DNA and RNA secondary structures held together by non-canonical guanine base pairs. G4 sequence motifs are enriched at specific sites in eukaryotic genomes, suggesting regulatory functions of G4 structures during different biological processes. Considering the high thermodynamic stability of G4 structures, various proteins are necessary for G4 structure formation and unwinding. In a yeast one-hybrid screen, we identified Slx9 as a novel G4-binding protein. We confirmed that Slx9 binds to G4 DNA structures in vitro. Despite these findings, Slx9 binds only insignificantly to G-rich/G4 regions in Saccharomyces cerevisiae as demonstrated by genome-wide ChIP-seq analysis. However, Slx9 binding to G4s is significantly increased in the absence of Sgs1, a RecQ helicase that regulates G4 structures. Different genetic and molecular analyses allowed us to propose a model in which Slx9 recognizes and protects stabilized G4 structures in vivo.


Assuntos
Proteínas de Ligação a DNA/química , Quadruplex G , Proteínas Ribossômicas/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , DNA Helicases/química , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Genoma/genética , Conformação de Ácido Nucleico , Ligação Proteica , RecQ Helicases/química , RecQ Helicases/genética , Proteínas Ribossômicas/química , Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/química , Termodinâmica
8.
Nat Cell Biol ; 21(5): 542-551, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31048766

RESUMO

A diverse catalog of long noncoding RNAs (lncRNAs), which lack protein-coding potential, are transcribed from the mammalian genome. They are emerging as important regulators in gene expression networks by controlling nuclear architecture and transcription in the nucleus and by modulating mRNA stability, translation and post-translational modifications in the cytoplasm. In this Review, we highlight recent progress in cellular functions of lncRNAs at the molecular level in mammalian cells.


Assuntos
Fenômenos Fisiológicos Celulares/genética , Genoma/genética , Biossíntese de Proteínas/genética , RNA Longo não Codificante/genética , Animais , Redes Reguladoras de Genes/genética , Humanos , Processamento de Proteína Pós-Traducional/genética , Estabilidade de RNA/genética
9.
BMC Bioinformatics ; 20(1): 232, 2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31072311

RESUMO

BACKGROUND: Draft quality genomes for a multitude of organisms have become common due to the advancement of genome assemblers using long-read technologies with high error rates. Although current assemblies are substantially more contiguous than assemblies based on short reads, complete chromosomal assemblies are still challenging. Interspersed repeat families with multiple copy versions dominate the contig and scaffold ends of current long-read assemblies for complex genomes. These repeat families generally remain unresolved, as existing algorithmic solutions either do not scale to large copy numbers or can not handle the current high read error rates. RESULTS: We propose novel repeat resolution methods for large interspersed repeat families and assess their accuracy on simulated data sets with various distinct repeat structures and on drosophila melanogaster transposons. Additionally, we compare our methods to an existing long read repeat resolution tool and show the improved accuracy of our method. CONCLUSIONS: Our results demonstrate the applicability of our methods for the improvement of the contiguity of genome assemblies.


Assuntos
Bases de Dados Genéticas/normas , Genoma/genética , Análise de Sequência de DNA/métodos , Algoritmos , Humanos
10.
J Anim Sci ; 97(7): 3027-3033, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-30997484

RESUMO

An efficient strategy to improve QTL detection power is performing across-breed validation studies. Variants segregating across breeds are expected to be in high linkage disequilibrium (LD) with causal mutations affecting economically important traits. The aim of this study was to validate, in a Tropical Composite cattle (TC) population, QTL associations identified for sexual precocity traits in a Nellore and Brahman meta-analysis genome-wide association study. In total, 2,816 TC, 8,001 Nellore, and 2,210 Brahman animals were available for the analysis. For that, genomic regions significantly associated with puberty traits in the meta-analysis study were validated for the following sexual precocity traits in TC: age at first corpus luteum (AGECL), first postpartum anestrus interval (PPAI), and scrotal circumference at 18 months of age (SC). We considered validated QTL those underpinned by significant markers from the Nellore and Brahman meta-analysis (P ≤ 10-4) that were also significant for a TC trait, i.e., presenting a P-value of ≤10-3 for AGECL, PPAI, or SC. We also considered as validated QTL those regions where significant markers in the reference population were at ±250 kb from significant markers in the validation population. Using this criteria, 49 SNP were validated for AGECL, 4 for PPAI, and 14 for SC, from which 5 were in common with AGECL, totaling 62 validated SNP for these traits and 30 candidate genes surrounding them. Considering just candidate genes closest to the top SNP of each chromosome, for AGECL 8 candidate genes were identified: COL8A1, PENK, ENSBTAG00000047425, BPNT1, ADAMTS17, CCHCR1, SUFU, and ENSBTAG00000046374. For PPAI, 3 genes emerged as candidates (PCBP3, KCNK10, and MRPS5), and for SC 8 candidate genes were identified (SNORA70, TRAC, ASS1, BPNT1, LRRK1, PKHD1, PTPRM, and ENSBTAG00000045690). Several candidate regions presented here were previously associated with puberty traits in cattle. The majority of emerging candidate genes are related to biological processes involved in reproductive events, such as maintenance of gestation, and some are known to be expressed in reproductive tissues. Our results suggested that some QTL controlling early puberty seem to be segregating across cattle breeds adapted to tropical conditions.


Assuntos
Bovinos/genética , Cromossomos/genética , Estudo de Associação Genômica Ampla/veterinária , Genoma/genética , Polimorfismo de Nucleotídeo Único/genética , Reprodução/genética , Maturidade Sexual/genética , Animais , Cruzamento , Bovinos/crescimento & desenvolvimento , Bovinos/fisiologia , Feminino , Frequência do Gene , Genômica , Genótipo , Desequilíbrio de Ligação , Masculino , Fenótipo , Gravidez , Locos de Características Quantitativas/genética
11.
Methods Cell Biol ; 151: 197-218, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30948008

RESUMO

To characterize the complex regulatory control of gene expression using fluorescent protein reporters, it is often necessary to analyze large genomic regions. Bacteria artificial chromosome (BAC) vectors, which are able to support DNA fragments of up to 300kb, provide stable platforms for experimental manipulation. Using phage-based systems of homologous recombination, BACs can be efficiently engineered for a variety of aims. These include expressing fluorescent proteins to delineate gene expression boundaries using high-resolution, in vivo microscopy, tracing cell lineages using stable fluorescent proteins, perturbing endogenous protein function by expressing dominant negative forms, interfering with development by mis-expressing transcription factors, and identifying regulatory regions through deletion analysis. Here, we present a series of protocols for identifying BAC clones that contain genes of interest, modifying BACs for use as reporter constructs, and preparing BAC DNA for microinjection into fertilized eggs. Although the protocols here are tailored for use in echinoderm embryonic and larval stages, these methods are easily adaptable for use in other transgenic systems. As fluorescent protein technology continues to expand, so do the potential applications for recombinant BACs.


Assuntos
Cromossomos Artificiais Bacterianos/genética , Equinodermos/genética , Genômica/métodos , Microinjeções/métodos , Animais , Equinodermos/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento/genética , Vetores Genéticos , Genoma/genética , Genômica/tendências , Microinjeções/tendências , Recombinação Genética , Sequências Reguladoras de Ácido Nucleico
12.
Methods Cell Biol ; 151: 305-321, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30948015

RESUMO

The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 (CRISPR-associated nuclease 9) technology enables rapid, targeted, and efficient changes in the genomes of various model organisms. The short guide RNAs (gRNAs) of the CRISPR/Cas9 system can be designed to recognize target DNA within coding regions for functional gene knockouts. Several studies have demonstrated that the CRISPR/Cas9 system efficiently and specifically targets sea urchin genes and results in expected mutant phenotypes. In addition to disrupting gene functions, modifications and additions to the Cas9 protein enable alternative activities targeted to specific sites within the genome. This includes a fusion of cytidine deaminase to Cas9 (Cas9-DA) for single nucleotide conversion in targeted sites. In this chapter, we describe detailed methods for the CRISPR/Cas9 application in sea urchin embryos, including gRNA design, in vitro synthesis of single guide RNA (sgRNA), and the usages of the CRISPR/Cas9 technology for gene knockout and single nucleotide editing. Methods for genotyping the resultant embryos are also provided for assessing efficiencies of gene editing.


Assuntos
Sistemas CRISPR-Cas/genética , Edição de Genes/métodos , Técnicas de Inativação de Genes/métodos , Ouriços-do-Mar/genética , Animais , Citidina Desaminase/genética , Marcação de Genes/métodos , Vetores Genéticos , Genoma/genética , Ouriços-do-Mar/crescimento & desenvolvimento
13.
Methods Cell Biol ; 151: 55-61, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30948031

RESUMO

At the most fundamental level, the genome is the basis for questions about the mechanisms of development: how it works. This perspective provides a brief historical review of the sequencing of the echinoderm genome and the progress in answering this complex question, which depends on technological advances as well as intellectual ones.


Assuntos
Genoma/genética , Genômica/história , Ouriços-do-Mar/genética , Animais , Mapeamento Cromossômico , Equinodermos/genética , História do Século XX , História do Século XXI
14.
Methods Cell Biol ; 151: 65-88, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30948032

RESUMO

Echinoderms are important research models for a wide range of biological questions. In particular, echinoderm embryos are exemplary models for dissecting the molecular and cellular processes that drive development and testing how these processes can be modified through evolution to produce the extensive morphological diversity observed in the phylum. Modern attempts to characterize these processes depend on some level of genomic analysis; from querying annotated gene sets to functional genomics experiments to identify candidate cis-regulatory sequences. Given how essential these data have become, it is important that researchers using available datasets or performing their own genome-scale experiments understand the nature and limitations of echinoderm genomic analyses. In this chapter we highlight the current state of echinoderm genomic data and provide methodological considerations for common approaches, including analysis of transcriptome and functional genomics datasets.


Assuntos
Equinodermos/genética , Desenvolvimento Embrionário/genética , Perfilação da Expressão Gênica/métodos , Genômica/métodos , Animais , Equinodermos/crescimento & desenvolvimento , Genoma/genética , Genômica/tendências , Anotação de Sequência Molecular/métodos
15.
Methods Cell Biol ; 151: 89-113, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30948033

RESUMO

The discovery of gene regulatory networks (GRNs) has opened a gate to access the genomic mechanisms controlling development. GRNs are systems of transcriptional regulatory circuits that control the differential specification of cell fates during development by regulating gene expression. The experimental analysis of GRNs involves a collection of methods, each revealing aspects of the overall control process. This review provides an overview of experimental and computational methods that have been successfully applied for solving developmental GRNs in the sea urchin embryo. The key in this approach is to obtain experimental evidence for functional interactions between transcription factors and regulatory DNA. In the second part of this review, a more generally applicable strategy is discussed that shows a path from experimental evidence to annotation of regulatory linkages to the generation of GRN models.


Assuntos
Biologia Computacional/métodos , Redes Reguladoras de Genes/genética , Modelos Biológicos , Ouriços-do-Mar/genética , Animais , Embrião não Mamífero , Regulação da Expressão Gênica no Desenvolvimento , Genoma/genética , Ouriços-do-Mar/crescimento & desenvolvimento
17.
Nat Rev Gastroenterol Hepatol ; 16(7): 395-410, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30936469

RESUMO

Hepatocytes operate in highly structured repeating anatomical units termed liver lobules. Blood flow along the lobule radial axis creates gradients of oxygen, nutrients and hormones, which, together with morphogenetic fields, give rise to a highly variable microenvironment. In line with this spatial variability, key liver functions are expressed non-uniformly across the lobules, a phenomenon termed zonation. Technologies based on single-cell transcriptomics have constructed a global spatial map of hepatocyte gene expression in mice revealing that ~50% of hepatocyte genes are expressed in a zonated manner. This broad spatial heterogeneity suggests that hepatocytes in different lobule zones might have not only different gene expression profiles but also distinct epigenetic features, regenerative capacities, susceptibilities to damage and other functional aspects. Here, we present genomic approaches for studying liver zonation, describe the principles of liver zonation and discuss the intrinsic and extrinsic factors that dictate zonation patterns. We also explore the challenges and solutions for obtaining zonation maps of liver non-parenchymal cells. These approaches facilitate global characterization of liver function with high spatial resolution along physiological and pathological timescales.


Assuntos
Perfilação da Expressão Gênica/métodos , Genoma/genética , Hepatócitos/metabolismo , Fígado/citologia , Animais , Hepatócitos/citologia , Humanos , Fígado/metabolismo , Imagem Individual de Molécula , Análise de Célula Única , Transcriptoma/genética
18.
Essays Biochem ; 63(1): 177-186, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-30967478

RESUMO

Chromatin architecture has a significant impact on gene expression. Evidence in the last two decades support RNA as an important component of chromatin structure [Genes Dev. (2005) 19, 1635-1655; PLoS ONE (2007) 2, e1182; Nat. Genet. (2002) 30, 329-334]. Long non-coding RNAs (lncRNAs) are able to control chromatin structure through nucleosome positioning, interaction with chromatin re-modellers and chromosome looping. These functions are carried out in cis at the site of lncRNAs transcription or in trans at distant loci. While the evidence for a role in lncRNAs in regulating gene expression through chromatin interactions is increasing, there is still very little conclusive evidence for a potential role in looping organisation. Here, we review models for the involvement of lncRNAs in genome architecture and the experimental evidence to support them.


Assuntos
Cromatina/genética , Genoma/genética , RNA Longo não Codificante/genética , Cromatina/química , Montagem e Desmontagem da Cromatina/genética , DNA/química , DNA/genética , Humanos , Conformação de Ácido Nucleico , Conformação Proteica
19.
Nat Biotechnol ; 37(5): 547-554, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30936559

RESUMO

Trajectory inference approaches analyze genome-wide omics data from thousands of single cells and computationally infer the order of these cells along developmental trajectories. Although more than 70 trajectory inference tools have already been developed, it is challenging to compare their performance because the input they require and output models they produce vary substantially. Here, we benchmark 45 of these methods on 110 real and 229 synthetic datasets for cellular ordering, topology, scalability and usability. Our results highlight the complementarity of existing tools, and that the choice of method should depend mostly on the dataset dimensions and trajectory topology. Based on these results, we develop a set of guidelines to help users select the best method for their dataset. Our freely available data and evaluation pipeline ( https://benchmark.dynverse.org ) will aid in the development of improved tools designed to analyze increasingly large and complex single-cell datasets.


Assuntos
Biologia Computacional/métodos , Genoma/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Análise de Célula Única/métodos , Benchmarking , Sequenciamento de Nucleotídeos em Larga Escala/tendências , Análise de Célula Única/tendências
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