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1.
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
2.
Nat Commun ; 11(1): 4522, 2020 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-32908144

RESUMO

A unique, protective cell envelope contributes to the broad drug resistance of the nosocomial pathogen Acinetobacter baumannii. Here we use transposon insertion sequencing to identify A. baumannii mutants displaying altered susceptibility to a panel of diverse antibiotics. By examining mutants with antibiotic susceptibility profiles that parallel mutations in characterized genes, we infer the function of multiple uncharacterized envelope proteins, some of which have roles in cell division or cell elongation. Remarkably, mutations affecting a predicted cell wall hydrolase lead to alterations in lipooligosaccharide synthesis. In addition, the analysis of altered susceptibility signatures and antibiotic-induced morphology patterns allows us to predict drug synergies; for example, certain beta-lactams appear to work cooperatively due to their preferential targeting of specific cell wall assembly machineries. Our results indicate that the pathogen may be effectively inhibited by the combined targeting of multiple pathways critical for envelope growth.


Assuntos
Infecções por Acinetobacter/tratamento farmacológico , Acinetobacter baumannii/genética , Antibacterianos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Infecção Hospitalar/tratamento farmacológico , Farmacorresistência Bacteriana Múltipla/genética , Infecções por Acinetobacter/microbiologia , Acinetobacter baumannii/efeitos dos fármacos , Antibacterianos/uso terapêutico , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Parede Celular/efeitos dos fármacos , Parede Celular/genética , Parede Celular/metabolismo , Infecção Hospitalar/microbiologia , Análise Mutacional de DNA , Elementos de DNA Transponíveis/genética , DNA Bacteriano/genética , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Sinergismo Farmacológico , Humanos , Testes de Sensibilidade Microbiana , Mutação
3.
BMC Infect Dis ; 20(1): 657, 2020 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-32894079

RESUMO

BACKGROUND: Tuberculosis is still a significant diagnostic and therapeutic challenge with high proportion of smear- and culture- negative incidences worldwide. The conventional diagnostic tests are time-consuming and have a low sensitivity. Digital PCR is a novel technology which can detect target sequences with relatively low abundance and obtain the absolute copy numbers of the targets. METHODS: We assessed the accuracy of dPCR in TB diagnosis using more than 250 specimens, and for the first time, we selected M.tuberculosis-specific IS1081 in addition to widely used IS6110 as the amplification targets for dPCR. The quantification of target DNA was calculated using QuantaSoft Version 1.7.4.0917 (BioRad), and SPSS version 13.0 software (SPSS Inc., Chicago, IL, USA) was used for statistical analyses. RESULTS: IS6110-dPCR was more sensitive than IS1081, with the sensitivity and specificity accounting for 40.6 and 93.4% respectively. When we classified the TB patients by personal factors for high copy number of M.tuberculosis derived DNA in plasma: bilateral TB, extrapulmonary TB and disseminated TB, the sensitivity of both IS6110- and IS1081- dPCR was the highest in patients with disseminated TB (IS6110, 100%; IS1081, 68.8%), while their sensitivity was a bit higher in patients with extrapulmonary TB (IS6110, 50.0%; IS1081, 39.3%) than that in bilateral TB (IS6110, 43.3%; IS1081, 33.3%). Compared with traditional TB diagnostic tests, joint detection IS6110 & IS1081-dPCR was not as sensitive as smear microscope or mycobacterial culture, but it was higher than IS6110 qPCR (p < 0.05) and was able to detect 47.4% of smear-negative TB patients. CONCLUSION: Our study suggested that plasma IS6110-dPCR is a rapid, moderate accurate and less-invasive method to detect M.tuberculosis DNA in plasma of TB patients and IS6110 & IS1081-dPCR has a potential to aid diagnosis of smear-negative TB.


Assuntos
Elementos de DNA Transponíveis/genética , DNA Bacteriano/genética , Mycobacterium tuberculosis/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Tuberculose Pulmonar/diagnóstico , Adolescente , Adulto , Idoso , DNA Bacteriano/sangue , Confiabilidade dos Dados , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Adulto Jovem
4.
Nat Commun ; 11(1): 4058, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32792480

RESUMO

Tomatoes come in a multitude of shapes and flavors despite a narrow genetic pool. Here, we leverage whole-genome resequencing data available for 602 cultivated and wild accessions to determine the contribution of transposable elements (TEs) to tomato diversity. We identify 6,906 TE insertions polymorphisms (TIPs), which result from the mobilization of 337 distinct TE families. Most TIPs are low frequency variants and TIPs are disproportionately located within or adjacent to genes involved in environmental responses. In addition, genic TE insertions tend to have strong transcriptional effects and they can notably lead to the generation of multiple transcript isoforms. Using genome-wide association studies (GWAS), we identify at least 40 TIPs robustly associated with extreme variation in major agronomic traits or secondary metabolites and in most cases, no SNP tags the TE insertion allele. Collectively, these findings highlight the unique role of TE mobilization in tomato diversification, with important implications for breeding.


Assuntos
Elementos de DNA Transponíveis/genética , Lycopersicon esculentum/genética , Evolução Molecular , Genoma de Planta/genética , Estudo de Associação Genômica Ampla , Polimorfismo Genético/genética
5.
J Med Microbiol ; 69(8): 1089-1094, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32692646

RESUMO

Introduction. The bla CTX-M-3 gene has rarely been reported in Morganella morganii strains and its genetic environment has not yet been investigated.Aim. To identify the bla CTX-M-3 gene in M. morganii isolated from swine and characterize its genetic environment.Methodology. A M. morganii isolate (named MM1L5) from a deceased swine was identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and subjected to antimicrobial susceptibility testing. The bla genes were detected and then the genetic location and environment of bla CTX-M-3 were investigated by Southern blot and PCR mapping, respectively. The M. morganii bla CTX-M-3 gene was cloned and expressed in Escherichia coli.Results. Isolate MM1L5 harboured the bla CTX-M-3 and bla TEM-1 genes. The bla CTX-M-3 gene, located on the chromosome, was co-carried with an IS26 and bla TEM-1 gene by a novel 6361 bp IS26-flanked composite transposon, designated Tn6741. This transposon consisted of a novel bla CTX-M-3-containing module, IS26-ΔISEcp1-bla CTX-M-3-Δorf477-IS26 (named Tn6710), and a bla TEM-1-containing module, IS26-Δorf477-bla TEM-1-tnpR-IS26, differing from previous reports. Phylogenetic analysis showed a significant variation based on the sequence of Tn6741, as compared to those of other related transposons. Interestingly, although the cloned bla CTX-M-3 gene could confer resistance to ceftiofur, cefquinome, ceftriaxone and cefotaxime, one amino acid substitution (Ile-142-Thr) resulted in a significant reduction of resistance to these antimicrobials.Conclusion. This is the first time that bla CTX-M-3 has been identified on a chromosome from a M. morganii isolate. Furthermore, the bla CTX-M-3 gene was located with an IS26 element and bla TEM-1 gene on a novel IS26-flanked composite transposon, Tn6741, suggesting that Tn6741 might act as a reservoir for the bla CTX-M-3 and bla TEM-1 genes and may become an important vehicle for their dissemination among M. morganii.


Assuntos
Elementos de DNA Transponíveis/genética , Morganella morganii/genética , beta-Lactamases/genética , Animais , Anti-Infecciosos/farmacologia , Sequência de Bases , Clonagem Molecular , Morganella morganii/classificação , Morganella morganii/efeitos dos fármacos , Filogenia , Reação em Cadeia da Polimerase , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Suínos
6.
Nat Commun ; 11(1): 3739, 2020 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-32719317

RESUMO

The PIWI protein MIWI2 and its associated PIWI-interacting RNAs (piRNAs) instruct DNA methylation of young active transposable elements (TEs) in the male germline. piRNAs are proposed to recruit MIWI2 to the transcriptionally active TE loci by base pairing to nascent transcripts, however the downstream mechanisms and effector proteins utilized by MIWI2 in directing de novo TE methylation remain incompletely understood. Here, we show that MIWI2 associates with TEX15 in foetal gonocytes. TEX15 is predominantly a nuclear protein that is not required for piRNA biogenesis but is essential for piRNA-directed TE de novo methylation and silencing. In summary, TEX15 is an essential executor of mammalian piRNA-directed DNA methylation.


Assuntos
Proteínas Argonauta/metabolismo , Proteínas de Ciclo Celular/metabolismo , Metilação de DNA/genética , Elementos de DNA Transponíveis/genética , Inativação Gênica , Animais , Proteínas Argonauta/genética , Feminino , Feto/citologia , Genoma , Células Germinativas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ligação Proteica , Testículo/metabolismo
7.
Nat Protoc ; 15(8): 2705-2727, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32681154

RESUMO

Invasive fungal infections caused by Candida species are life threatening with high mortality, posing a severe public health threat. New technologies for rapid, genome-wide identification of virulence genes and therapeutic targets are urgently needed. Our recent engineering of a piggyBac (PB) transposon-mediated mutagenesis system in haploid Candida albicans provides a powerful discovery tool, which we anticipate should be adaptable to other haploid Candida species. In this protocol, we use haploid C. albicans as an example to present an improved version of the mutagenesis system and provide a detailed description of the protocol for constructing high-quality mutant libraries. We also describe a method for quantitative PB insertion site sequencing, PBISeq. The PBISeq library preparation procedure exploits tagmentation to quickly and efficiently construct sequencing libraries. Finally, we present a pipeline to analyze PB insertion sites in a de novo assembled genome of our engineered haploid C. albicans strain. The entire protocol takes ~7 d from transposition induction to having a final library ready for sequencing. This protocol is highly efficient and less labor intensive than alternative approaches and significantly accelerates genetic studies of Candida.


Assuntos
Candida/genética , Elementos de DNA Transponíveis/genética , Genoma Fúngico/genética , Haploidia , Mutagênese Insercional/métodos
8.
Nat Commun ; 11(1): 3446, 2020 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-32651359

RESUMO

The piggyBac DNA transposon is used widely in genome engineering applications. Unlike other transposons, its excision site can be precisely repaired without leaving footprints and it integrates specifically at TTAA tetranucleotides. We present cryo-EM structures of piggyBac transpososomes: a synaptic complex with hairpin DNA intermediates and a strand transfer complex capturing the integration step. The results show that the excised TTAA hairpin intermediate and the TTAA target adopt essentially identical conformations, providing a mechanistic link connecting the two unique properties of piggyBac. The transposase forms an asymmetric dimer in which the two central domains synapse the ends while two C-terminal domains form a separate dimer that contacts only one transposon end. In the strand transfer structure, target DNA is severely bent and the TTAA target is unpaired. In-cell data suggest that asymmetry promotes synaptic complex formation, and modifying ends with additional transposase binding sites stimulates activity.


Assuntos
Elementos de DNA Transponíveis/genética , Transposases/metabolismo , Microscopia Crioeletrônica , Humanos , Espectroscopia de Ressonância Magnética , Ligação Proteica , Estrutura Secundária de Proteína , Transposases/genética
9.
Nat Commun ; 11(1): 3506, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32665538

RESUMO

Acute myeloid leukemia (AML) is characterised by a series of genetic and epigenetic alterations that result in deregulation of transcriptional networks. One understudied source of transcriptional regulators are transposable elements (TEs), whose aberrant usage could contribute to oncogenic transcriptional circuits. However, the regulatory influence of TEs and their links to AML pathogenesis remain unexplored. Here we identify six endogenous retrovirus (ERV) families with AML-associated enhancer chromatin signatures that are enriched in binding of key regulators of hematopoiesis and AML pathogenesis. Using both locus-specific genetic editing and simultaneous epigenetic silencing of multiple ERVs, we demonstrate that ERV deregulation directly alters the expression of adjacent genes in AML. Strikingly, deletion or epigenetic silencing of an ERV-derived enhancer suppresses cell growth by inducing apoptosis in leukemia cell lines. This work reveals that ERVs are a previously unappreciated source of AML enhancers that may be exploited by cancer cells to help drive tumour heterogeneity and evolution.


Assuntos
Cromatina/metabolismo , Leucemia Mieloide Aguda/genética , Animais , Linhagem Celular , Cromatina/genética , Elementos de DNA Transponíveis/genética , Retrovirus Endógenos/genética , Epigênese Genética/genética , Redes Reguladoras de Genes/genética , Redes Reguladoras de Genes/fisiologia , Genoma Humano/genética , Humanos , Sequências Repetitivas Dispersas/genética
10.
Proc Natl Acad Sci U S A ; 117(31): 18670-18679, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32675236

RESUMO

As the most abundant microbes on Earth, novel bacteriophages (phages; bacteria-specific viruses) are readily isolated from environmental samples. However, it remains challenging to characterize phage-bacteria interactions, such as the host receptor(s) phages bind to initiate infection. Here, we tested whether transposon insertion sequencing (INSeq) could be used to identify bacterial genes involved in phage binding. As proof of concept, results showed that INSeq screens successfully identified genes encoding known receptors for previously characterized viruses of Escherichia coli (phages T6, T2, T4, and T7). INSeq screens were then used to identify genes involved during infection of six newly isolated coliphages. Results showed that candidate receptors could be successfully identified for the majority (five of six) of the phages; furthermore, genes encoding the phage receptor(s) were the top hit(s) in the analyses of the successful screens. INSeq screens provide a generally useful method for high-throughput discovery of phage receptors. We discuss limitations of our approach when examining uncharacterized phages, as well as usefulness of the method for exploring the evolution of broad versus narrow use of cellular receptors among phages in the biosphere.


Assuntos
Proteínas de Bactérias/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Receptores Virais/genética , Proteínas de Bactérias/metabolismo , Bacteriófagos/metabolismo , Elementos de DNA Transponíveis/genética , DNA Bacteriano/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Biblioteca Gênica , Receptores Virais/metabolismo
11.
PLoS Genet ; 16(7): e1008931, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32644999

RESUMO

Shigella species are specialised lineages of Escherichia coli that have converged to become human-adapted and cause dysentery by invading human gut epithelial cells. Most studies of Shigella evolution have been restricted to comparisons of single representatives of each species; and population genomic studies of individual Shigella species have focused on genomic variation caused by single nucleotide variants and ignored the contribution of insertion sequences (IS) which are highly prevalent in Shigella genomes. Here, we investigate the distribution and evolutionary dynamics of IS within populations of Shigella dysenteriae Sd1, Shigella sonnei and Shigella flexneri. We find that five IS (IS1, IS2, IS4, IS600 and IS911) have undergone expansion in all Shigella species, creating substantial strain-to-strain variation within each population and contributing to convergent patterns of functional gene loss within and between species. We find that IS expansion and genome degradation are most advanced in S. dysenteriae and least advanced in S. sonnei; and using genome-scale models of metabolism we show that Shigella species display convergent loss of core E. coli metabolic capabilities, with S. sonnei and S. flexneri following a similar trajectory of metabolic streamlining to that of S. dysenteriae. This study highlights the importance of IS to the evolution of Shigella and provides a framework for the investigation of IS dynamics and metabolic reduction in other bacterial species.


Assuntos
Elementos de DNA Transponíveis/genética , Disenteria/genética , Evolução Molecular , Shigella dysenteriae/genética , DNA Bacteriano/genética , Disenteria/microbiologia , Escherichia coli/genética , Escherichia coli/patogenicidade , Genoma Bacteriano/genética , Humanos , Shigella dysenteriae/patogenicidade
12.
PLoS Genet ; 16(7): e1008872, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32673310

RESUMO

Transposable elements (TEs) are genomic parasites that selfishly replicate at the expense of host fitness. Fifty years of evolutionary studies of TEs have concentrated on the deleterious genetic effects of TEs, such as their effects on disrupting genes and regulatory sequences. However, a flurry of recent work suggests that there is another important source of TEs' harmful effects-epigenetic silencing. Host genomes typically silence TEs by the deposition of repressive epigenetic marks. While this silencing reduces the selfish replication of TEs and should benefit hosts, a picture is emerging that the epigenetic silencing of TEs triggers inadvertent spreading of repressive marks to otherwise expressed neighboring genes, ultimately jeopardizing host fitness. In this Review, we provide a long-overdue overview of the recent genome-wide evidence for the presence and prevalence of TEs' epigenetic effects, highlighting both the similarities and differences across mammals, insects, and plants. We lay out the current understanding of the functional and fitness consequences of TEs' epigenetic effects, and propose possible influences of such effects on the evolution of both hosts and TEs themselves. These unique evolutionary consequences indicate that TEs' epigenetic effect is not only a crucial component of TE biology but could also be a significant contributor to genome function and evolution.


Assuntos
Elementos de DNA Transponíveis/genética , Epigênese Genética , Evolução Molecular , Inativação Gênica , Animais , Repressão Epigenética/genética , Regulação da Expressão Gênica/genética , Insetos/genética , Mamíferos/genética , Plantas/genética
13.
PLoS Genet ; 16(6): e1008861, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32525870

RESUMO

In metazoan germlines, the piRNA pathway acts as a genomic immune system, employing small RNA-mediated silencing to defend host DNA from the harmful effects of transposable elements (TEs). Expression of genomic TEs is proposed to initiate self regulation by increasing the production of repressive piRNAs, thereby "adapting" piRNA-mediated control to the most active TE families. Surprisingly, however, piRNA pathway proteins, which execute piRNA biogenesis and enforce silencing of targeted sequences, evolve rapidly and adaptively in animals. If TE silencing is ensured through piRNA biogenesis, what necessitates changes in piRNA pathway proteins? Here we used interspecific complementation to test for functional differences between Drosophila melanogaster and D. simulans alleles of three adaptively evolving piRNA pathway proteins: Armitage, Aubergine and Spindle-E. In contrast to piRNA-mediated transcriptional regulators examined in previous studies, these three proteins have cytoplasmic functions in piRNA maturation and post-transcriptional silencing. Across all three proteins we observed interspecific divergence in the regulation of only a handful of TE families, which were more robustly silenced by the heterospecific piRNA pathway protein. This unexpected result suggests that unlike transcriptional regulators, positive selection has not acted on cytoplasmic piRNA effector proteins to enhance their function in TE repression. Rather, TEs may evolve to "escape" silencing by host proteins. We further discovered that D. simulans alleles of aub and armi exhibit enhanced off-target effects on host transcripts in a D. melanogaster background, as well as modest reductions in the efficiency of piRNA biogenesis, suggesting that promiscuous binding of D. simulans Aub and Armi proteins to host transcripts reduces their participation in piRNA production. Avoidance of genomic auto-immunity may therefore be a critical target of selection. Our observations suggest that piRNA effector proteins are subject to an evolutionary trade-off between defending the host genome from the harmful effect of TEs while also minimizing collateral damage to host genes.


Assuntos
Autoimunidade/genética , Elementos de DNA Transponíveis/imunologia , Drosophila simulans/genética , Evolução Molecular , Genoma de Inseto/imunologia , RNA Interferente Pequeno/biossíntese , Alelos , Animais , Animais Geneticamente Modificados , Citoplasma/genética , Citoplasma/metabolismo , Elementos de DNA Transponíveis/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/imunologia , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/imunologia , Drosophila melanogaster/metabolismo , Drosophila simulans/metabolismo , Feminino , Regulação da Expressão Gênica/imunologia , Genoma de Inseto/genética , Masculino , Mutação , Interferência de RNA/imunologia
14.
BMC Evol Biol ; 20(1): 66, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32503430

RESUMO

BACKGROUND: Alternative splicing (AS) generates various transcripts from a single gene and thus plays a significant role in transcriptomic diversity and proteomic complexity. Alu elements are primate-specific transposable elements (TEs) and can provide a donor or acceptor site for AS. In a study on TE-mediated AS, we recently identified a novel AluSz6-exonized ACTR8 transcript of the crab-eating monkey (Macaca fascicularis). In the present study, we sought to determine the molecular mechanism of AluSz6 exonization of the ACTR8 gene and investigate its evolutionary and functional consequences in the crab-eating monkey. RESULTS: We performed RT-PCR and genomic PCR to analyze AluSz6 exonization in the ACTR8 gene and the expression of the AluSz6-exonized transcript in nine primate samples, including prosimians, New world monkeys, Old world monkeys, and hominoids. AluSz6 integration was estimated to have occurred before the divergence of simians and prosimians. The Alu-exonized transcript obtained by AS was lineage-specific and expressed only in Old world monkeys and apes, and humans. This lineage-specific expression was caused by a single G duplication in AluSz6, which provides a new canonical 5' splicing site. We further identified other alternative transcripts that were unaffected by the AluSz6 insertion. Finally, we observed that the alternative transcripts were transcribed into new isoforms with C-terminus deletion, and in silico analysis showed that these isoforms do not have a destructive function. CONCLUSIONS: The single G duplication in the TE sequence is the source of TE exonization and AS, and this mutation may suffer a different fate of ACTR8 gene expression during primate evolution.


Assuntos
Regulação da Expressão Gênica , Proteínas dos Microfilamentos/genética , Mutação , Primatas/genética , Processamento Alternativo , Elementos Alu/genética , Animais , Elementos de DNA Transponíveis/genética , Evolução Molecular , Éxons/genética , Humanos
15.
BMC Evol Biol ; 20(1): 69, 2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32564776

RESUMO

BACKGROUND: East African lake cichlids are one of the most impressive examples of an adaptive radiation. Independently in Lake Victoria, Tanganyika, and Malawi, several hundreds of species arose within the last 10 million to 100,000 years. Whereas most analyses in cichlids focused on nucleotide substitutions across species to investigate the genetic bases of this explosive radiation, to date, no study has investigated the contribution of structural variants (SVs) in the evolution of adaptive traits across the three Great Lakes of East Africa. RESULTS: Here, we annotate and characterize the repertoires and evolutionary potential of different SV classes (deletion, duplication, inversion, insertions and translocations) in four cichlid species: Haplochromis burtoni, Metriaclima zebra, Neolamprologus brichardi and Pundamilia nyererei. We investigate the patterns of gain and loss evolution for each SV type, enabling the identification of lineage specific events. Both deletions and inversions show a significant overlap with SINE elements, while inversions additionally show a limited, but significant association with DNA transposons. Inverted regions are enriched for genes regulating behaviour, or involved in skeletal and visual system development. We also find that duplicated regions show enrichment for genes associated with "antigen processing and presentation" and other immune related categories. Our pipeline and results were further tested by PCR validation of selected deletions and inversions, which confirmed respectively 7 out of 10 and 6 out of 9 events. CONCLUSIONS: Altogether, we provide the first comprehensive overview of rearrangement evolution in East African cichlids, and some important insights into their likely contribution to adaptation.


Assuntos
Ciclídeos/genética , Variação Genética , Crescimento e Desenvolvimento/genética , Imunidade/genética , Animais , Ciclídeos/crescimento & desenvolvimento , Ciclídeos/imunologia , Elementos de DNA Transponíveis/genética , Evolução Molecular , Fenótipo
16.
PLoS Genet ; 16(6): e1008864, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32584820

RESUMO

Cytosine methylation is an ancient epigenetic modification yet its function and extent within genomes is highly variable across eukaryotes. In mammals, methylation controls transposable elements and regulates the promoters of genes. In insects, DNA methylation is generally restricted to a small subset of transcribed genes, with both intergenic regions and transposable elements (TEs) depleted of methylation. The evolutionary origin and the function of these methylation patterns are poorly understood. Here we characterise the evolution of DNA methylation across the arthropod phylum. While the common ancestor of the arthropods had low levels of TE methylation and did not methylate promoters, both of these functions have evolved independently in centipedes and mealybugs. In contrast, methylation of the exons of a subset of transcribed genes is ancestral and widely conserved across the phylum, but has been lost in specific lineages. A similar set of genes is methylated in all species that retained exon-enriched methylation. We show that these genes have characteristic patterns of expression correlating to broad transcription initiation sites and well-positioned nucleosomes, providing new insights into potential mechanisms driving methylation patterns over hundreds of millions of years.


Assuntos
Artrópodes/genética , Metilação de DNA , Epigênese Genética , Evolução Molecular , Animais , Ilhas de CpG/genética , Elementos de DNA Transponíveis/genética , Éxons/genética , Filogenia , Regiões Promotoras Genéticas/genética
17.
Nat Commun ; 11(1): 3256, 2020 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-32591500

RESUMO

BRCA1 mutation carriers have a higher risk of developing triple-negative breast cancer (TNBC), which is a refractory disease due to its non-responsiveness to current clinical targeted therapies. Using the Sleeping Beauty transposon system in Brca1-deficient mice, we identified 169 putative cancer drivers, among which Notch1 is a top candidate for accelerating TNBC by promoting the epithelial-mesenchymal transition (EMT) and regulating the cell cycle. Activation of NOTCH1 suppresses mitotic catastrophe caused by BRCA1 deficiency by restoring S/G2 and G2/M cell cycle checkpoints, which may through activation of ATR-CHK1 signalling pathway. Consistently, analysis of human breast cancer tissue demonstrates NOTCH1 is highly expressed in TNBCs, and the activated form of NOTCH1 correlates positively with increased phosphorylation of ATR. Additionally, we demonstrate that inhibition of the NOTCH1-ATR-CHK1 cascade together with cisplatin synergistically kills TNBC by targeting the cell cycle checkpoint, DNA damage and EMT, providing a potent clinical option for this fatal disease.


Assuntos
Proteína BRCA1/deficiência , Carcinogênese/patologia , Receptor Notch1/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Alelos , Animais , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Proteína BRCA1/metabolismo , Morte Celular , Linhagem Celular Tumoral , Quinase 1 do Ponto de Checagem/metabolismo , Elementos de DNA Transponíveis/genética , Progressão da Doença , Transição Epitelial-Mesenquimal , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Knockout , Mitose , Mutação/genética , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/genética
18.
Nat Commun ; 11(1): 3224, 2020 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-32591528

RESUMO

In plants, epigenetic regulation is critical for silencing transposons and maintaining proper gene expression. However, its impact on the genome-wide transcription initiation landscape remains elusive. By conducting a genome-wide analysis of transcription start sites (TSSs) using cap analysis of gene expression (CAGE) sequencing, we show that thousands of TSSs are exclusively activated in various epigenetic mutants of Arabidopsis thaliana and referred to as cryptic TSSs. Many have not been identified in previous studies, of which up to 65% are contributed by transposons. They possess similar genetic features to regular TSSs and their activation is strongly associated with the ectopic recruitment of RNAPII machinery. The activation of cryptic TSSs significantly alters transcription of nearby TSSs, including those of genes important for development and stress responses. Our study, therefore, sheds light on the role of epigenetic regulation in maintaining proper gene functions in plants by suppressing transcription from cryptic TSSs.


Assuntos
Arabidopsis/genética , Epigênese Genética , Regulação da Expressão Gênica de Plantas , Transcrição Genética , Sequência de Bases , Sequência Consenso/genética , Metilação de DNA/genética , DNA Polimerase beta/metabolismo , Elementos de DNA Transponíveis/genética , Genes de Plantas , Mutação/genética , RNA Polimerase II/metabolismo , Sítio de Iniciação de Transcrição , Transcriptoma/genética
19.
PLoS Genet ; 16(6): e1008894, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32598340

RESUMO

Meiotic crossovers (COs) are important for reshuffling genetic information between homologous chromosomes and they are essential for their correct segregation. COs are unevenly distributed along chromosomes and the underlying mechanisms controlling CO localization are not well understood. We previously showed that meiotic COs are mis-localized in the absence of AXR1, an enzyme involved in the neddylation/rubylation protein modification pathway in Arabidopsis thaliana. Here, we report that in axr1-/-, male meiocytes show a strong defect in chromosome pairing whereas the formation of the telomere bouquet is not affected. COs are also redistributed towards subtelomeric chromosomal ends where they frequently form clusters, in contrast to large central regions depleted in recombination. The CO suppressed regions correlate with DNA hypermethylation of transposable elements (TEs) in the CHH context in axr1-/- meiocytes. Through examining somatic methylomes, we found axr1-/- affects DNA methylation in a plant, causing hypermethylation in all sequence contexts (CG, CHG and CHH) in TEs. Impairment of the main pathways involved in DNA methylation is epistatic over axr1-/- for DNA methylation in somatic cells but does not restore regular chromosome segregation during meiosis. Collectively, our findings reveal that the neddylation pathway not only regulates hormonal perception and CO distribution but is also, directly or indirectly, a major limiting pathway of TE DNA methylation in somatic cells.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Cromossomos de Plantas/genética , Metilação de DNA , Meiose/genética , Proteínas de Arabidopsis/genética , Pareamento Cromossômico , Segregação de Cromossomos , Troca Genética , Quebras de DNA de Cadeia Dupla , Elementos de DNA Transponíveis/genética , Técnicas de Inativação de Genes , Plantas Geneticamente Modificadas
20.
PLoS Genet ; 16(6): e1008822, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32497040

RESUMO

Insecticide resistance in malaria vectors threatens to reverse recent gains in malaria control. Deciphering patterns of gene flow and resistance evolution in malaria vectors is crucial to improving control strategies and preventing malaria resurgence. A genome-wide survey of Anopheles funestus genetic diversity Africa-wide revealed evidences of a major division between southern Africa and elsewhere, associated with different population histories. Three genomic regions exhibited strong signatures of selective sweeps, each spanning major resistance loci (CYP6P9a/b, GSTe2 and CYP9K1). However, a sharp regional contrast was observed between populations correlating with gene flow barriers. Signatures of complex molecular evolution of resistance were detected with evidence of copy number variation, transposon insertion and a gene conversion between CYP6P9a/b paralog genes. Temporal analyses of samples before and after bed net scale up suggest that these genomic changes are driven by this control intervention. Multiple independent selective sweeps at the same locus in different parts of Africa suggests that local evolution of resistance in malaria vectors may be a greater threat than trans-regional spread of resistance haplotypes.


Assuntos
Anopheles/genética , Evolução Molecular , Genoma de Inseto/genética , Resistência a Inseticidas/genética , Malária/prevenção & controle , Mosquitos Vetores/genética , África , Alelos , Animais , Anopheles/parasitologia , Família 6 do Citocromo P450/genética , Variações do Número de Cópias de DNA , Elementos de DNA Transponíveis/genética , Fluxo Gênico , Loci Gênicos , Haplótipos , Humanos , Proteínas de Insetos/genética , Malária/parasitologia , Malária/transmissão , Metagenômica , Controle de Mosquitos/métodos , Polimorfismo Genético , Piretrinas , Sequenciamento Completo do Genoma
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