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1.
Int J Mol Sci ; 25(13)2024 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-39000388

RESUMO

Biological invasions have been identified as the fifth cause of biodiversity loss, and their subsequent dispersal represents a major ecological challenge. The aquatic invasive species Ludwigia grandiflora subsp. hexapetala (Lgh) and Ludwigia peploides subsp. montevidensis (Lpm) are largely distributed in aquatic environments in North America and in Europe. However, they also present worrying terrestrial forms that are able to colonize wet meadows. To comprehend the mechanisms of the terrestrial adaptation of Lgh and Lpm, it is necessary to develop their genomic resources, which are currently poorly documented. We performed de novo assembly of the mitogenomes of Lgh and Lpm through hybrid assemblies, combining short reads (SR) and/or long reads (LR) before annotating both mitogenomes. We successfully assembled the mitogenomes of Lgh and Lpm into two circular molecules each, resulting in a combined total length of 711,578 bp and 722,518 bp, respectively. Notably, both the Lgh and Lpm molecules contained plastome-origin sequences, comprising 7.8% of the mitochondrial genome length. Additionally, we identified recombinations that were mediated by large repeats, suggesting the presence of multiple alternative conformations. In conclusion, our study presents the first high-quality mitogenomes of Lpm and Lgh, which are the only ones in the Myrtales order found as two circular molecules.


Assuntos
Genoma Mitocondrial , Edição de RNA , Recombinação Genética , Filogenia , Genômica/métodos , Genoma de Planta , Cromossomos de Plantas/genética
2.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 41(7): 853-857, 2024 Jul 10.
Artigo em Chinês | MEDLINE | ID: mdl-38946372

RESUMO

OBJECTIVE: To analyze a Chinese pedigree with a recombination occurring between the HLA-A/C loci in both parents. METHODS: A patient who was planning to undergo hematopoietic stem cell transplantation due to "aplastic anemia" in February 2022 was selected as the study subject. Peripheral blood samples were collected from the patient, his parents and brother. HLA-A/C/B/DRB1/DQB1 high-resolution typing was carried out by using sequence-based typing and sequence-specific oligonucleotides. The recombination was identified by pedigree analysis. The HLA haplotype of each individual was identified by genealogical analysis. The parentage possibility was determined by short tandem repeat analysis. HLA-A/C/B/DRB1/DRB345/DQA1/DQB1/DPA1/DPB1 were determined with next-generation high-throughput sequence-based typing. The recombination sites were analyzed by family study. RESULTS: The high parentage possibilities of the family was confirmed by short tandem repeat analysis. Recombination was found between the HLA-A*24:02 A*33:03/C*14:03 in the paternally transmitted haplotype, whilst HLA-A*01:01 A*03:01/C*08:02 was found in the maternally transmitted haplotype, which had resulted in two novel HLA haplotypes in the proband. CONCLUSION: A rare case with simultaneous recombination of the paternal and maternal HLA-A/C loci has been discovered, which may facilitate further study of the mechanisms of the HLA recombination.


Assuntos
Povo Asiático , Antígenos HLA-A , Haplótipos , Linhagem , Recombinação Genética , Adulto , Feminino , Humanos , Masculino , Povo Asiático/genética , População do Leste Asiático , Teste de Histocompatibilidade , Antígenos HLA-A/genética , Antígenos HLA-C/genética , Repetições de Microssatélites , Pais
3.
Arch Virol ; 169(7): 155, 2024 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-38951272

RESUMO

Given the high prevalence of avian leukosis virus subgroup K (ALV-K) in chickens in China, the positive rate of ALV-K in local chickens in Henan province was investigated, and the genetic region encoding the glycoprotein gp85 of isolates from positive chickens was analyzed. The positive rate of ALV-K in local chickens in Henan was found to be 87.2% (41/47). Phylogenetic analysis of gp85 sequences revealed six clusters that differed in their host range regions (hr1 and hr2) and variable regions (vr1, vr2, and vr3). Evidence of recombination of hr1, hr2, vr1, vr2, and vr3 was observed between the different clusters. The isolate HN23LS02 appears to have obtained its hr1 and hr2 regions from separate lineages via recombination but without having a significant affect on the replication capacity of the virus.


Assuntos
Vírus da Leucose Aviária , Leucose Aviária , Galinhas , Especificidade de Hospedeiro , Filogenia , Doenças das Aves Domésticas , Recombinação Genética , Proteínas do Envelope Viral , Animais , Vírus da Leucose Aviária/genética , Vírus da Leucose Aviária/classificação , Vírus da Leucose Aviária/isolamento & purificação , Galinhas/virologia , Leucose Aviária/virologia , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/metabolismo , Doenças das Aves Domésticas/virologia , China
4.
Commun Biol ; 7(1): 829, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38977904

RESUMO

Modern plant breeding, such as genomic selection and gene editing, is based on the knowledge of the genetic architecture of desired traits. Quantitative trait loci (QTL) analysis, which combines high throughput phenotyping and genotyping of segregating populations, is a powerful tool to identify these genetic determinants and to decipher the underlying mechanisms. However, meiotic recombination, which shuffles genetic information between generations, is limited: Typically only one to two exchange points, called crossovers, occur between a pair of homologous chromosomes. Here we test the effect on QTL analysis of boosting recombination, by mutating the anti-crossover factors RECQ4 and FIGL1 in Arabidopsis thaliana full hybrids and lines in which a single chromosome is hybrid. We show that increasing recombination ~6-fold empowers the detection and resolution of QTLs, reaching the gene scale with only a few hundred plants. Further, enhanced recombination unmasks some secondary QTLs undetected under normal recombination. These results show the benefits of enhanced recombination to decipher the genetic bases of traits.


Assuntos
Arabidopsis , Mapeamento Cromossômico , Locos de Características Quantitativas , Recombinação Genética , Arabidopsis/genética , Mapeamento Cromossômico/métodos , Proteínas de Arabidopsis/genética , Fenótipo , RecQ Helicases/genética , Melhoramento Vegetal/métodos , Cromossomos de Plantas/genética , Troca Genética
5.
BMC Biol ; 22(1): 151, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38977974

RESUMO

BACKGROUND: RNA-DNA hybrids or R-loops are associated with deleterious genomic instability and protective immunoglobulin class switch recombination (CSR). However, the underlying phenomenon regulating the two contrasting functions of R-loops is unknown. Notably, the underlying mechanism that protects R-loops from classic RNase H-mediated digestion thereby promoting persistence of CSR-associated R-loops during CSR remains elusive. RESULTS: Here, we report that during CSR, R-loops formed at the immunoglobulin heavy (IgH) chain are modified by ribose 2'-O-methylation (2'-OMe). Moreover, we find that 2'-O-methyltransferase fibrillarin (FBL) interacts with activation-induced cytidine deaminase (AID) associated snoRNA aSNORD1C to facilitate the 2'-OMe. Moreover, deleting AID C-terminal tail impairs its association with aSNORD1C and FBL. Disrupting FBL, AID or aSNORD1C expression severely impairs 2'-OMe, R-loop stability and CSR. Surprisingly, FBL, AID's interaction partner and aSNORD1C promoted AID targeting to the IgH locus. CONCLUSION: Taken together, our results suggest that 2'-OMe stabilizes IgH-associated R-loops to enable productive CSR. These results would shed light on AID-mediated CSR and explain the mechanism of R-loop-associated genomic instability.


Assuntos
Citidina Desaminase , Switching de Imunoglobulina , Estruturas R-Loop , Switching de Imunoglobulina/genética , Citidina Desaminase/metabolismo , Citidina Desaminase/genética , Citidina Desaminase/química , Animais , Camundongos , Metilação , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Pesadas de Imunoglobulinas/metabolismo , Recombinação Genética , RNA/metabolismo , RNA/genética
6.
Mol Biol Evol ; 41(7)2024 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-38950035

RESUMO

Natural selection is less efficient in the absence of recombination. As a result, nonrecombining sequences, such as sex chromosomes, tend to degenerate over time. Although the outcomes of recombination arrest are typically observed after many millions of generations, recent neo-sex chromosomes can give insight into the early stages of this process. Here, we investigate the evolution of neo-sex chromosomes in the Spanish marbled white butterfly, Melanargia ines, where a Z-autosome fusion has turned the homologous autosome into a nonrecombining neo-W chromosome. We show that these neo-sex chromosomes are likely limited to the Iberian population of M. ines, and that they arose around the time when this population split from North-African populations, around 1.5 million years ago. Recombination arrest of the neo-W chromosome has led to an excess of premature stop-codons and frame-shift mutations, and reduced gene expression compared to the neo-Z chromosome. Surprisingly, we identified two regions of ∼1 Mb at one end of the neo-W that are both less diverged from the neo-Z and less degraded than the rest of the chromosome, suggesting a history of rare but repeated genetic exchange between the two neo-sex chromosomes. These plateaus of neo-sex chromosome divergence suggest that neo-W degradation can be locally reversed by rare recombination between neo-W and neo-Z chromosomes.


Assuntos
Borboletas , Recombinação Genética , Cromossomos Sexuais , Animais , Cromossomos Sexuais/genética , Masculino , Borboletas/genética , Feminino , Evolução Molecular , Seleção Genética
7.
Mol Biol Evol ; 41(7)2024 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-38959451

RESUMO

Meiotic recombination is a fundamental feature of sexually reproducing species. It is often required for proper chromosome segregation and plays important role in adaptation and the maintenance of genetic diversity. The molecular mechanisms of recombination are remarkably conserved across eukaryotes, yet meiotic genes and proteins show substantial variation in their sequence and function, even between closely related species. Furthermore, the rate and distribution of recombination shows a huge diversity within and between chromosomes, individuals, sexes, populations, and species. This variation has implications for many molecular and evolutionary processes, yet how and why this diversity has evolved is not well understood. A key step in understanding trait evolution is to determine its genetic basis-that is, the number, effect sizes, and distribution of loci underpinning variation. In this perspective, I discuss past and current knowledge on the genetic basis of variation in recombination rate and distribution, explore its evolutionary implications, and present open questions for future research.


Assuntos
Variação Genética , Meiose , Recombinação Genética , Meiose/genética , Animais , Evolução Molecular , Evolução Biológica
8.
Appl Microbiol Biotechnol ; 108(1): 400, 2024 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-38951186

RESUMO

Over the past years, several methods have been developed for gene cloning. Choosing a cloning strategy depends on various factors, among which simplicity and affordability have always been considered. The aim of this study, on the one hand, is to simplify gene cloning by skipping in vitro assembly reactions and, on the other hand, to reduce costs by eliminating relatively expensive materials. We investigated a cloning system using Escherichia coli harboring two plasmids, pLP-AmpR and pScissors-CmR. The pLP-AmpR contains a landing pad (LP) consisting of two genes (λ int and λ gam) that allow the replacement of the transformed linear DNA using site-specific recombination. After the replacement process, the inducible expressing SpCas9 and specific sgRNA from the pScissors-CmR (CRISPR/Cas9) vector leads to the removal of non-recombinant pLP-AmpR plasmids. The function of LP was explored by directly transforming PCR products. The pScissors-CmR plasmid was evaluated for curing three vectors, including the origins of pBR322, p15A, and pSC101. Replacing LP with a PCR product and fast-eradicating pSC101 origin-containing vectors was successful. Recombinant colonies were confirmed following gene replacement and plasmid curing processes. The results made us optimistic that this strategy may potentially be a simple and inexpensive cloning method. KEY POINTS: •The in vivo cloning was performed by replacing the target gene with the landing pad. •Fast eradication of non-recombinant plasmids was possible by adapting key vectors. •This strategy is not dependent on in vitro assembly reactions and expensive materials.


Assuntos
Clonagem Molecular , Escherichia coli , Plasmídeos , Reação em Cadeia da Polimerase , Recombinação Genética , Escherichia coli/genética , Clonagem Molecular/métodos , Plasmídeos/genética , Reação em Cadeia da Polimerase/métodos , Vetores Genéticos/genética , Sistemas CRISPR-Cas
9.
PLoS Genet ; 20(7): e1011336, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38950081

RESUMO

Increasing natural resistance and resilience in plants is key for ensuring food security within a changing climate. Breeders improve these traits by crossing cultivars with their wild relatives and introgressing specific alleles through meiotic recombination. However, some genomic regions are devoid of recombination especially in crosses between divergent genomes, limiting the combinations of desirable alleles. Here, we used pooled-pollen sequencing to build a map of recombinant and non-recombinant regions between tomato and five wild relatives commonly used for introgressive tomato breeding. We detected hybrid-specific recombination coldspots that underscore the role of structural variations in modifying recombination patterns and maintaining genetic linkage in interspecific crosses. Crossover regions and coldspots show strong association with specific TE superfamilies exhibiting differentially accessible chromatin between somatic and meiotic cells. About two-thirds of the genome are conserved coldspots, located mostly in the pericentromeres and enriched with retrotransposons. The coldspots also harbor genes associated with agronomic traits and stress resistance, revealing undesired consequences of linkage drag and possible barriers to breeding. We presented examples of linkage drag that can potentially be resolved by pairing tomato with other wild species. Overall, this catalogue will help breeders better understand crossover localization and make informed decisions on generating new tomato varieties.


Assuntos
Genoma de Planta , Recombinação Genética , Solanum lycopersicum , Solanum lycopersicum/genética , Hibridização Genética , Ligação Genética , Melhoramento Vegetal , Retroelementos/genética , Troca Genética , Meiose/genética , Mapeamento Cromossômico , Cromossomos de Plantas/genética , Alelos
10.
BMC Plant Biol ; 24(1): 645, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38972991

RESUMO

Melia azedarach is a species of enormous value of pharmaceutical industries. Although the chloroplast genome of M. azedarach has been explored, the information of mitochondrial genome (Mt genome) remains surprisingly limited. In this study, we used a hybrid assembly strategy of BGI short-reads and Nanopore long-reads to assemble the Mt genome of M. azedarach. The Mt genome of M. azedarach is characterized by two circular chromosomes with 350,142 bp and 290,387 bp in length, respectively, which encodes 35 protein-coding genes (PCGs), 23 tRNA genes, and 3 rRNA genes. A pair of direct repeats (R1 and R2) were associated with genome recombination, resulting in two conformations based on the Sanger sequencing and Oxford Nanopore sequencing. Comparative analysis identified 19 homologous fragments between Mt and chloroplast genome, with the longest fragment of 12,142 bp. The phylogenetic analysis based on PCGs were consist with the latest classification of the Angiosperm Phylogeny Group. Notably, a total of 356 potential RNA editing sites were predicted based on 35 PCGs, and the editing events lead to the formation of the stop codon in the rps10 gene and the start codons in the nad4L and atp9 genes, which were verified by PCR amplification and Sanger sequencing. Taken together, the exploration of M. azedarach gap-free Mt genome provides a new insight into the evolution research and complex mitogenome architecture.


Assuntos
Genoma Mitocondrial , Filogenia , Recombinação Genética , Sequências Repetitivas de Ácido Nucleico/genética , Genoma de Cloroplastos , Genoma de Planta , Edição de RNA
11.
BMC Plant Biol ; 24(1): 644, 2024 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-38973002

RESUMO

BACKGROUND: Floating bamboo (Hygroryza aristata) is an endangered species with a narrow native distribution and is renowned for its unique aesthetic qualities, which holds significant ecological and ornamental value. However, the lack of genetic information research, with only one complete plastome available, significantly hampers conservation efforts and further research for this species. RESULTS: In this research, we sequenced and assembled the organelle genomes of floating bamboo, including the mitogenome (587,847 bp) and plastome (135,675 bp). The mitogenome can recombine into various configurations, which are mediated by 25 repeat pairs (13 SRs, 6 MRs, 1 LR, and 5 CRs). LR1 and SR5 are particularly notable as they have the ability to combine with other contigs, forming complex repeat units that facilitate further homologous recombination. The rate of homologous recombination varies significantly among species, yet there is still a pronounced positive correlation observed between the length of these repeat pairs and the rate of recombination they mediate. The mitogenome integrates seven intact protein-coding genes from the chloroplast. The codon usage patterns in both organelles are similar, with a noticeable bias towards C and T on the third codon. The gene map of Poales shows the entire loss of rpl6, succinate dehydrogenase subunits (sdh3 and sdh4). Additionally, the BOP clade retained more variable genes compared to the PACMAD clade. CONCLUSIONS: We provided a high-quality and well-annotated mitogenome for floating bamboo and demonstrated the presence of diverse configurations. Our study has revealed the correlation between repeat length and their corresponding recombination rate despite variations among species. Although the mitogenome can potentially exist in the form of a unicircular in vivo, this occurrence is rare and may not be stable.


Assuntos
Genoma Mitocondrial , Poaceae , Poaceae/genética , Recombinação Genética , Sequências Repetitivas de Ácido Nucleico/genética , Genoma de Planta
12.
Mol Biol Evol ; 41(7)2024 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-38874402

RESUMO

Inferring the demographic history of populations provides fundamental insights into species dynamics and is essential for developing a null model to accurately study selective processes. However, background selection and selective sweeps can produce genomic signatures at linked sites that mimic or mask signals associated with historical population size change. While the theoretical biases introduced by the linked effects of selection have been well established, it is unclear whether ancestral recombination graph (ARG)-based approaches to demographic inference in typical empirical analyses are susceptible to misinference due to these effects. To address this, we developed highly realistic forward simulations of human and Drosophila melanogaster populations, including empirically estimated variability of gene density, mutation rates, recombination rates, purifying, and positive selection, across different historical demographic scenarios, to broadly assess the impact of selection on demographic inference using a genealogy-based approach. Our results indicate that the linked effects of selection minimally impact demographic inference for human populations, although it could cause misinference in populations with similar genome architecture and population parameters experiencing more frequent recurrent sweeps. We found that accurate demographic inference of D. melanogaster populations by ARG-based methods is compromised by the presence of pervasive background selection alone, leading to spurious inferences of recent population expansion, which may be further worsened by recurrent sweeps, depending on the proportion and strength of beneficial mutations. Caution and additional testing with species-specific simulations are needed when inferring population history with non-human populations using ARG-based approaches to avoid misinference due to the linked effects of selection.


Assuntos
Drosophila melanogaster , Modelos Genéticos , Densidade Demográfica , Seleção Genética , Animais , Drosophila melanogaster/genética , Humanos , Recombinação Genética , Genética Populacional/métodos , Simulação por Computador , Taxa de Mutação
13.
Mol Biol Evol ; 41(6)2024 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-38829800

RESUMO

It is commonly thought that the long-term advantage of meiotic recombination is to dissipate genetic linkage, allowing natural selection to act independently on different loci. It is thus theoretically expected that genes with higher recombination rates evolve under more effective selection. On the other hand, recombination is often associated with GC-biased gene conversion (gBGC), which theoretically interferes with selection by promoting the fixation of deleterious GC alleles. To test these predictions, several studies assessed whether selection was more effective in highly recombining genes (due to dissipation of genetic linkage) or less effective (due to gBGC), assuming a fixed distribution of fitness effects (DFE) for all genes. In this study, I directly derive the DFE from a gene's evolutionary history (shaped by mutation, selection, drift, and gBGC) under empirical fitness landscapes. I show that genes that have experienced high levels of gBGC are less fit and thus have more opportunities for beneficial mutations. Only a small decrease in the genome-wide intensity of gBGC leads to the fixation of these beneficial mutations, particularly in highly recombining genes. This results in increased positive selection in highly recombining genes that is not caused by more effective selection. Additionally, I show that the death of a recombination hotspot can lead to a higher dN/dS than its birth, but with substitution patterns biased towards AT, and only at selected positions. This shows that controlling for a substitution bias towards GC is therefore not sufficient to rule out the contribution of gBGC to signatures of accelerated evolution. Finally, although gBGC does not affect the fixation probability of GC-conservative mutations, I show that by altering the DFE, gBGC can also significantly affect nonsynonymous GC-conservative substitution patterns.


Assuntos
Evolução Molecular , Conversão Gênica , Modelos Genéticos , Recombinação Genética , Seleção Genética , Aptidão Genética , Mutação , Composição de Bases , Ligação Genética
14.
Int J Mol Sci ; 25(12)2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38928003

RESUMO

Barley with high grain ß-glucan content is valuable for functional foods. The identification of loci for high ß-glucan content is, thus, of great importance for barley breeding. Segregation mapping for the content in ß-glucan and other barley grain components (starch, protein, lipid, ash, phosphorous, calcium, sodium) was performed using the progeny of the cross between Glacier AC38, a mutant with high amylose, and CDC Fibar, a high ß-glucan waxy cultivar. The offspring of this cross showed transgressive segregation for ß-glucan content. Linkage analysis based on single-nucleotide polymorphism (SNP) molecular markers was used for the genotyping of the parents and recombinant inbred lines (RILs). Two Quantitative Trait Loci (QTL) for ß-glucan content and several QTL for other grain components were found. The former ones, located on chromosomes 1H and 7H, explained 27.9% and 27.4% of the phenotypic variance, respectively. Glacier AC38 provided the allele for high ß-glucan content at the QTL on chromosome 1H, whereas CDC Fibar contributed the allele at the QTL on chromosome 7H. Their recombination resulted in a novel haplotype with higher ß-glucan content, up to 18.4%. Candidate genes are proposed for these two QTL: HvCslF9, involved in ß-glucan biosynthesis, for the QTL on chromosome 1H; Horvu_PLANET_7H01G069300, a gene encoding an ATP-Binding Cassette (ABC) transporter, for the QTL on chromosome 7H.


Assuntos
Mapeamento Cromossômico , Hordeum , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , beta-Glucanas , Hordeum/genética , Hordeum/metabolismo , beta-Glucanas/metabolismo , Fenótipo , Cromossomos de Plantas/genética , Grão Comestível/genética , Grão Comestível/metabolismo , Genótipo , Sementes/genética , Sementes/metabolismo , Sementes/química , Melhoramento Vegetal , Recombinação Genética/genética , Haplótipos
15.
Sci Rep ; 14(1): 13182, 2024 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-38849496

RESUMO

Recombinant HIV-1 genomes identified in three or more epidemiological unrelated individuals are defined as circulating recombinant forms (CRFs). CRFs can further recombine with other pure subtypes or recombinants to produce secondary recombinants. In this study, a new HIV-1 intersubtype CRF, designated CRF159_01103, isolated from three men who have sex with men with no epidemiological linkage, was identified in Baoding city, Hebei Province, China. CRF159_01103 was derived from CRF103_01B and CRF01_AE. Bayesian molecular clock analysis was performed on the CRF01-AE and CRF103_01B regions of CRF159_01103. The time of origin of CRF159_01103 was predicted to be 2018-2019, indicating that it is a recent recombinant virus. The emergence of CRF159_01103 has increased the complexity of the HIV-1 epidemic in Hebei Province.


Assuntos
Infecções por HIV , HIV-1 , Filogenia , Recombinação Genética , HIV-1/genética , HIV-1/classificação , HIV-1/isolamento & purificação , Humanos , China/epidemiologia , Infecções por HIV/virologia , Infecções por HIV/epidemiologia , Masculino , Genoma Viral , Homossexualidade Masculina , Teorema de Bayes
16.
Nature ; 630(8018): 994-1002, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38926616

RESUMO

Insertion sequence (IS) elements are the simplest autonomous transposable elements found in prokaryotic genomes1. We recently discovered that IS110 family elements encode a recombinase and a non-coding bridge RNA (bRNA) that confers modular specificity for target DNA and donor DNA through two programmable loops2. Here we report the cryo-electron microscopy structures of the IS110 recombinase in complex with its bRNA, target DNA and donor DNA in three different stages of the recombination reaction cycle. The IS110 synaptic complex comprises two recombinase dimers, one of which houses the target-binding loop of the bRNA and binds to target DNA, whereas the other coordinates the bRNA donor-binding loop and donor DNA. We uncovered the formation of a composite RuvC-Tnp active site that spans the two dimers, positioning the catalytic serine residues adjacent to the recombination sites in both target and donor DNA. A comparison of the three structures revealed that (1) the top strands of target and donor DNA are cleaved at the composite active sites to form covalent 5'-phosphoserine intermediates, (2) the cleaved DNA strands are exchanged and religated to create a Holliday junction intermediate, and (3) this intermediate is subsequently resolved by cleavage of the bottom strands. Overall, this study reveals the mechanism by which a bispecific RNA confers target and donor DNA specificity to IS110 recombinases for programmable DNA recombination.


Assuntos
DNA , RNA não Traduzido , Recombinação Genética , Domínio Catalítico , Microscopia Crioeletrônica , DNA/química , DNA/metabolismo , DNA/ultraestrutura , Elementos de DNA Transponíveis/genética , Modelos Moleculares , Conformação de Ácido Nucleico , Multimerização Proteica , Recombinases/química , Recombinases/genética , Recombinases/metabolismo , RNA não Traduzido/química , RNA não Traduzido/genética , RNA não Traduzido/metabolismo , RNA não Traduzido/ultraestrutura , Especificidade por Substrato
17.
Virulence ; 15(1): 2366874, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38869140

RESUMO

Recombinant Muscovy duck parvovirus (rMDPV) is a product of genetic recombination between classical Muscovy duck parvovirus (MDPV) and goose parvovirus (GPV). The recombination event took place within a 1.1-kb DNA segment located in the middle of the VP3 gene, and a 187-bp sequence extending from the P9 promoter to the 5' initiation region of the Rep1 ORF. This resulted in the alteration of five amino acids within VP3. Despite these genetic changes, the precise influence of recombination and amino acid mutations on the pathogenicity of rMDPV remains ambiguous. In this study, based on the rMDPV strain ZW and the classical MDPV strain YY, three chimeric viruses (rZW-mP9, rZW-mPR187, and rYY-rVP3) and the five amino acid mutations-introduced mutants (rZW-g5aa and rYY-5aa(ZW)) were generated using reverse genetic technology. When compared to the parental virus rZW, rZW-g5aa exhibited a prolonged mean death time (MDT) and a decreased median lethal dose (ELD50) in embryonated duck eggs. In contrast, rYY-5aa(ZW) did not display significant differences in MDT and ELD50 compared to rYY. In 2-day-old Muscovy ducklings, infection with rZW-g5aa and rYY-5aa(ZW) resulted in mortality rates of only 20% and 10%, respectively, while infections with the three chimeric viruses (rZW-mP9, rZW-mPR187, rYY-rVP3) and rZW still led to 100% mortality. Notably, rYY-rVP3, containing the VP3 region from strain ZW, exhibited 50% mortality in 6-day-old Muscovy ducklings and demonstrated significant horizontal transmission. Collectively, our findings indicate that recombination and consequent amino acid changes in VP3 have a synergistic impact on the heightened virulence of rMDPV in Muscovy ducklings.


Assuntos
Proteínas do Capsídeo , Patos , Infecções por Parvoviridae , Mutação Puntual , Doenças das Aves Domésticas , Recombinação Genética , Animais , Virulência , Infecções por Parvoviridae/virologia , Infecções por Parvoviridae/veterinária , Doenças das Aves Domésticas/virologia , Proteínas do Capsídeo/genética , Parvovirinae/genética , Parvovirinae/patogenicidade
18.
PLoS Genet ; 20(6): e1011162, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38885280

RESUMO

Very little is known about the process of meiosis in the apicomplexan parasite Cryptosporidium despite the essentiality of sex in its life cycle. Most cell lines only support asexual growth of Cryptosporidium parvum (C. parvum), but stem cell derived intestinal epithelial cells grown under air-liquid interface (ALI) conditions support the sexual cycle. To examine chromosomal dynamics during meiosis in C. parvum, we generated two transgenic lines of parasites that were fluorescently tagged with mCherry or GFP on chromosomes 1 or 5, respectively. Infection of ALI cultures or Ifngr1-/- mice with mCherry and GFP parasites resulted in cross-fertilization and the formation of "yellow" oocysts, which contain 4 haploid sporozoites that are the product of meiosis. Recombinant oocysts from the F1 generation were purified and used to infect HCT-8 cultures, and phenotypes of the progeny were observed by microscopy. All possible phenotypes predicted by independent segregation were represented equally (~25%) in the population, indicating that C. parvum chromosomes exhibit a Mendelian inheritance pattern. The most common pattern observed from the outgrowth of single oocysts included all possible parental and recombinant phenotypes derived from a single meiotic event, suggesting a high rate of crossover. To estimate the frequency of crossover, additional loci on chromosomes 1 and 5 were tagged and used to monitor intrachromosomal crosses in Ifngr1-/- mice. Both chromosomes showed a high frequency of crossover compared to other apicomplexans with map distances (i.e., 1% recombination) of 3-12 kb. Overall, a high recombination rate may explain many unique characteristics observed in Cryptosporidium spp. such as high rates of speciation, wide variation in host range, and rapid evolution of host-specific virulence factors.


Assuntos
Criptosporidiose , Cryptosporidium parvum , Meiose , Oocistos , Recombinação Genética , Animais , Cryptosporidium parvum/genética , Camundongos , Criptosporidiose/parasitologia , Criptosporidiose/genética , Meiose/genética , Humanos , Receptores de Interferon/genética , Receptor de Interferon gama , Segregação de Cromossomos/genética , Esporozoítos/genética , Camundongos Knockout , Fenótipo
19.
Nature ; 630(8018): 984-993, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38926615

RESUMO

Genomic rearrangements, encompassing mutational changes in the genome such as insertions, deletions or inversions, are essential for genetic diversity. These rearrangements are typically orchestrated by enzymes that are involved in fundamental DNA repair processes, such as homologous recombination, or in the transposition of foreign genetic material by viruses and mobile genetic elements1,2. Here we report that IS110 insertion sequences, a family of minimal and autonomous mobile genetic elements, express a structured non-coding RNA that binds specifically to their encoded recombinase. This bridge RNA contains two internal loops encoding nucleotide stretches that base-pair with the target DNA and the donor DNA, which is the IS110 element itself. We demonstrate that the target-binding and donor-binding loops can be independently reprogrammed to direct sequence-specific recombination between two DNA molecules. This modularity enables the insertion of DNA into genomic target sites, as well as programmable DNA excision and inversion. The IS110 bridge recombination system expands the diversity of nucleic-acid-guided systems beyond CRISPR and RNA interference, offering a unified mechanism for the three fundamental DNA rearrangements-insertion, excision and inversion-that are required for genome design.


Assuntos
DNA , RNA não Traduzido , Recombinação Genética , Pareamento de Bases , Sequência de Bases , DNA/genética , DNA/metabolismo , Elementos de DNA Transponíveis/genética , Mutagênese Insercional/genética , Recombinases/metabolismo , Recombinases/genética , Recombinação Genética/genética , RNA não Traduzido/genética , RNA não Traduzido/metabolismo
20.
Nat Ecol Evol ; 8(7): 1337-1352, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38839849

RESUMO

Despite deep evolutionary conservation, recombination rates vary greatly across the genome and among individuals, sexes and populations. Yet the impact of this variation on adaptively diverging populations is not well understood. Here we characterized fine-scale recombination landscapes in an adaptively divergent pair of marine and freshwater populations of threespine stickleback from River Tyne, Scotland. Through whole-genome sequencing of large nuclear families, we identified the genomic locations of almost 50,000 crossovers and built recombination maps for marine, freshwater and hybrid individuals at a resolution of 3.8 kb. We used these maps to quantify the factors driving variation in recombination rates. We found strong heterochiasmy between sexes but also differences in recombination rates among ecotypes. Hybrids showed evidence of significant recombination suppression in overall map length and in individual loci. Recombination rates were lower not only within individual marine-freshwater-adaptive loci, but also between loci on the same chromosome, suggesting selection on linked gene 'cassettes'. Through temporal sampling along a natural hybrid zone, we found that recombinants showed traits associated with reduced fitness. Our results support predictions that divergence in cis-acting recombination modifiers, whose functions are disrupted in hybrids, may play an important role in maintaining differences among adaptively diverging populations.


Assuntos
Recombinação Genética , Smegmamorpha , Animais , Smegmamorpha/genética , Escócia , Masculino , Feminino , Aptidão Genética , Variação Genética
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