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1.
JAMA Neurol ; 78(10): 1236-1248, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34459874

RESUMO

Importance: Juvenile amyotrophic lateral sclerosis (ALS) is a rare form of ALS characterized by age of symptom onset less than 25 years and a variable presentation. Objective: To identify the genetic variants associated with juvenile ALS. Design, Setting, and Participants: In this multicenter family-based genetic study, trio whole-exome sequencing was performed to identify the disease-associated gene in a case series of unrelated patients diagnosed with juvenile ALS and severe growth retardation. The patients and their family members were enrolled at academic hospitals and a government research facility between March 1, 2016, and March 13, 2020, and were observed until October 1, 2020. Whole-exome sequencing was also performed in a series of patients with juvenile ALS. A total of 66 patients with juvenile ALS and 6258 adult patients with ALS participated in the study. Patients were selected for the study based on their diagnosis, and all eligible participants were enrolled in the study. None of the participants had a family history of neurological disorders, suggesting de novo variants as the underlying genetic mechanism. Main Outcomes and Measures: De novo variants present only in the index case and not in unaffected family members. Results: Trio whole-exome sequencing was performed in 3 patients diagnosed with juvenile ALS and their parents. An additional 63 patients with juvenile ALS and 6258 adult patients with ALS were subsequently screened for variants in the SPTLC1 gene. De novo variants in SPTLC1 (p.Ala20Ser in 2 patients and p.Ser331Tyr in 1 patient) were identified in 3 unrelated patients diagnosed with juvenile ALS and failure to thrive. A fourth variant (p.Leu39del) was identified in a patient with juvenile ALS where parental DNA was unavailable. Variants in this gene have been previously shown to be associated with autosomal-dominant hereditary sensory autonomic neuropathy, type 1A, by disrupting an essential enzyme complex in the sphingolipid synthesis pathway. Conclusions and Relevance: These data broaden the phenotype associated with SPTLC1 and suggest that patients presenting with juvenile ALS should be screened for variants in this gene.

2.
Mol Biol Evol ; 2021 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-34464971

RESUMO

Human centromeres are mainly composed of alpha satellite DNA hierarchically organized as higher-order repeats (HORs). Alpha satellite dynamics is shown by sequence homogenization in centromeric arrays and by its transfer to other centromeric locations, for example during the maturation of new centromeres. We identified during prenatal aneuploidy diagnosis by FISH a de novo insertion of alpha satellite DNA from the centromere of chromosome 18 (D18Z1) into cytoband 15q26. Although bound by CENP-B, this locus did not acquire centromeric functionality as demonstrated by lack of constriction and absence of CENP-A binding. The insertion was associated with a 2.8 kbp deletion and likely occurred in the paternal germline. The site was enriched in long terminal repeats (LTRs) and located ∼10 Mbp from the location where a centromere was ancestrally seeded and became inactive in the common ancestor of humans and apes 20-25 million years ago. Long-read mapping to the T2T-CHM13 human genome assembly revealed that the insertion derives from a specific region of chromosome 18 centromeric 12-mer HOR array in which the monomer size follows a regular pattern. The rearrangement did not directly disrupt any gene or predicted regulatory element and did not alter the methylation status of the surrounding region, consistent with the absence of phenotypic consequences in the carrier. This case demonstrates a likely rare but new class of structural variation that we name 'alpha satellite insertion'. It also expands our knowledge on alphoid DNA dynamics and conveys the possibility that alphoid arrays can relocate near vestigial centromeric sites.

3.
Elife ; 102021 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-34279216

RESUMO

Over 100 years of studies in Drosophila melanogaster and related species in the genus Drosophila have facilitated key discoveries in genetics, genomics, and evolution. While high-quality genome assemblies exist for several species in this group, they only encompass a small fraction of the genus. Recent advances in long-read sequencing allow high-quality genome assemblies for tens or even hundreds of species to be efficiently generated. Here, we utilize Oxford Nanopore sequencing to build an open community resource of genome assemblies for 101 lines of 93 drosophilid species encompassing 14 species groups and 35 sub-groups. The genomes are highly contiguous and complete, with an average contig N50 of 10.5 Mb and greater than 97% BUSCO completeness in 97/101 assemblies. We show that Nanopore-based assemblies are highly accurate in coding regions, particularly with respect to coding insertions and deletions. These assemblies, along with a detailed laboratory protocol and assembly pipelines, are released as a public resource and will serve as a starting point for addressing broad questions of genetics, ecology, and evolution at the scale of hundreds of species.

4.
Am J Hum Genet ; 108(8): 1436-1449, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34216551

RESUMO

Despite widespread clinical genetic testing, many individuals with suspected genetic conditions lack a precise diagnosis, limiting their opportunity to take advantage of state-of-the-art treatments. In some cases, testing reveals difficult-to-evaluate structural differences, candidate variants that do not fully explain the phenotype, single pathogenic variants in recessive disorders, or no variants in genes of interest. Thus, there is a need for better tools to identify a precise genetic diagnosis in individuals when conventional testing approaches have been exhausted. We performed targeted long-read sequencing (T-LRS) using adaptive sampling on the Oxford Nanopore platform on 40 individuals, 10 of whom lacked a complete molecular diagnosis. We computationally targeted up to 151 Mbp of sequence per individual and searched for pathogenic substitutions, structural variants, and methylation differences using a single data source. We detected all genomic aberrations-including single-nucleotide variants, copy number changes, repeat expansions, and methylation differences-identified by prior clinical testing. In 8/8 individuals with complex structural rearrangements, T-LRS enabled more precise resolution of the mutation, leading to changes in clinical management in one case. In ten individuals with suspected Mendelian conditions lacking a precise genetic diagnosis, T-LRS identified pathogenic or likely pathogenic variants in six and variants of uncertain significance in two others. T-LRS accurately identifies pathogenic structural variants, resolves complex rearrangements, and identifies Mendelian variants not detected by other technologies. T-LRS represents an efficient and cost-effective strategy to evaluate high-priority genes and regions or complex clinical testing results.


Assuntos
Aberrações Cromossômicas , Análise Citogenética/métodos , Doenças Genéticas Inatas/diagnóstico , Doenças Genéticas Inatas/genética , Predisposição Genética para Doença , Genoma Humano , Mutação , Variações do Número de Cópias de DNA , Feminino , Testes Genéticos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Cariotipagem , Masculino , Análise de Sequência de DNA
5.
6.
Am J Med Genet A ; 185(7): 2136-2149, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33783941

RESUMO

Van den Ende-Gupta syndrome (VDEGS) is a rare autosomal recessive condition characterized by distinctive facial and skeletal features, and in most affected persons, by biallelic pathogenic variants in SCARF2. We review the type and frequency of the clinical features in 36 reported individuals with features of VDEGS, 15 (42%) of whom had known pathogenic variants in SCARF2, 6 (16%) with negative SCARF2 testing, and 15 (42%) not tested. We also report three new individuals with pathogenic variants in SCARF2 and clinical features of VDEGS. Of the six persons without known pathogenic variants in SCARF2, three remain unsolved despite extensive genetic testing. Three were found to have pathogenic ABL1 variants using whole exome sequencing (WES) or whole genome sequencing (WGS). Their phenotype was consistent with the congenital heart disease and skeletal malformations syndrome (CHDSKM), which has been associated with ABL1 variants. Of the three unsolved cases, two were brothers who underwent WGS and targeted long-range sequencing of both SCARF2 and ABL1, and the third person who underwent WES and RNA sequencing for SCARF2. Because these affected individuals with classical features of VDEGS lacked a detectable pathogenic SCARF2 variant, genetic heterogeneity is likely. Our study shows the importance of performing genetic testing on individuals with the VDEGS "phenotype," either as a targeted gene analysis (SCARF2, ABL1) or WES/WGS. Additionally, individuals with the combination of arachnodactyly and blepharophimosis should undergo echocardiography while awaiting results of molecular testing due to the overlapping physical features of VDEGS and CHDSKM.

7.
Nucleic Acids Res ; 49(2): 879-890, 2021 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-33406239

RESUMO

Programmed DNA double-strand breaks (DSBs) made during meiosis are repaired by recombination with the homologous chromosome to generate, at selected sites, reciprocal crossovers that are critical for the proper separation of homologs in the first meiotic division. Backup repair processes can compensate when the normal meiotic recombination processes are non-functional. We describe a novel backup repair mechanism that occurs when the homologous chromosome is not available in Drosophila melanogaster meiosis. In the presence of a previously described mutation (Mcm5A7) that disrupts chromosome pairing, DSB repair is initiated by homologous recombination but is completed by non-homologous end joining (NHEJ). Remarkably, this process yields precise repair products. Our results provide support for a recombination intermediate recently proposed in mouse meiosis, in which an oligonucleotide bound to the Spo11 protein that catalyzes DSB formation remains bound after resection. We propose that this oligonucleotide functions as a primer for fill-in synthesis to allow scarless repair by NHEJ. We argue that this is a conserved repair mechanism that is likely to be invoked to overcome occasional challenges in normal meiosis.


Assuntos
Proteínas de Ciclo Celular/fisiologia , Quebras de DNA de Cadeia Dupla , Reparo do DNA por Junção de Extremidades/genética , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/genética , Meiose/genética , Oligonucleotídeos/genética , Animais , Proteínas de Ciclo Celular/genética , Simulação por Computador , Troca Genética , DNA Ligase Dependente de ATP/fisiologia , Proteínas de Drosophila/genética , Endodesoxirribonucleases/fisiologia , Feminino , Masculino , Modelos Genéticos , Mutação de Sentido Incorreto , Mutação Puntual , Polimorfismo de Nucleotídeo Único , Rad51 Recombinase/fisiologia , Alinhamento de Sequência , Deleção de Sequência , Sequenciamento Completo do Genoma
8.
PLoS Genet ; 16(12): e1008911, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33332348

RESUMO

Ploidy is the number of whole sets of chromosomes in a species. Ploidy is typically a stable cellular feature that is critical for survival. Polyploidization is a route recognized to increase gene dosage, improve fitness under stressful conditions and promote evolutionary diversity. However, the mechanism of regulation and maintenance of ploidy is not well characterized. Here, we examine the spontaneous diploidization associated with mutations in components of the Saccharomyces cerevisiae centrosome, known as the spindle pole body (SPB). Although SPB mutants are associated with defects in spindle formation, we show that two copies of the mutant in a haploid yeast favors diploidization in some cases, leading us to speculate that the increased gene dosage in diploids 'rescues' SPB duplication defects, allowing cells to successfully propagate with a stable diploid karyotype. This copy number-based rescue is linked to SPB scaling: certain SPB subcomplexes do not scale or only minimally scale with ploidy. We hypothesize that lesions in structures with incompatible allometries such as the centrosome may drive changes such as whole genome duplication, which have shaped the evolutionary landscape of many eukaryotes.


Assuntos
Centrômero/genética , Cromossomos Fúngicos/genética , Diploide , Dosagem de Genes , Centrômero/metabolismo , Cromossomos Fúngicos/metabolismo , Saccharomyces cerevisiae , Corpos Polares do Fuso/genética , Corpos Polares do Fuso/metabolismo
9.
Genetics ; 216(3): 621-631, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-33158985

RESUMO

The term interchromosomal effect was originally used to describe a change in the distribution of exchange in the presence of an inversion. First characterized in the 1920s by early Drosophila researchers, it has been observed in multiple organisms. Nearly half a century later, the term began to appear in the human genetics literature to describe the hypothesis that parental chromosome differences, such as translocations or inversions, may increase the frequency of meiotic chromosome nondisjunction. Although it remains unclear if chromosome aberrations truly affect the segregation of structurally normal chromosomes in humans, the use of the term interchromosomal effect in this context persists. This article explores the history of the use of the term interchromosomal effect and discusses how chromosomes with structural aberrations are segregated during meiosis.

10.
G3 (Bethesda) ; 10(11): 4271-4285, 2020 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-32972999

RESUMO

Balancers are rearranged chromosomes used in Drosophila melanogaster to maintain deleterious mutations in stable populations, preserve sets of linked genetic elements and construct complex experimental stocks. Here, we assess the phenotypes associated with breakpoint-induced mutations on commonly used third chromosome balancers and show remarkably few deleterious effects. We demonstrate that a breakpoint in p53 causes loss of radiation-induced apoptosis and a breakpoint in Fucosyltransferase A causes loss of fucosylation in nervous and intestinal tissue-the latter study providing new markers for intestinal cell identity and challenging previous conclusions about the regulation of fucosylation. We also describe thousands of potentially harmful mutations shared among X or third chromosome balancers, or unique to specific balancers, including an Ankyrin 2 mutation present on most TM3 balancers, and reiterate the risks of using balancers as experimental controls. We used long-read sequencing to confirm or refine the positions of two inversions with breakpoints lying in repetitive sequences and provide evidence that one of the inversions, In(2L)Cy, arose by ectopic recombination between foldback transposon insertions and the other, In(3R)C, cleanly separates subtelomeric and telomeric sequences and moves the subtelomeric sequences to an internal chromosome position. In addition, our characterization of In(3R)C shows that balancers may be polymorphic for terminal deletions. Finally, we present evidence that extremely distal mutations on balancers can add to the stability of stocks whose purpose is to maintain homologous chromosomes carrying mutations in distal genes. Overall, these studies add to our understanding of the structure, diversity and effectiveness of balancer chromosomes.


Assuntos
Cromossomos , Drosophila melanogaster , Animais , Inversão Cromossômica , Drosophila melanogaster/genética , Mutação , Fenótipo
11.
Am J Med Genet A ; 182(7): 1576-1591, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32500973

RESUMO

More than 50 individuals with activating variants in the receptor tyrosine kinase PDGFRB have been reported, separated based on clinical features into solitary myofibromas, infantile myofibromatosis, Penttinen syndrome with premature aging and osteopenia, Kosaki overgrowth syndrome, and fusiform aneurysms. Despite their descriptions as distinct clinical entities, review of previous reports demonstrates substantial phenotypic overlap. We present a case series of 12 patients with activating variants in PDGFRB and review of the literature. We describe five patients with PDGFRB activating variants whose clinical features overlap multiple diagnostic entities. Seven additional patients from a large family had variable expressivity and late-onset disease, including adult onset features and two individuals with sudden death. Three patients were treated with imatinib and had robust and rapid response, including the first two reported infants with multicentric myofibromas treated with imatinib monotherapy and one with a recurrent p.Val665Ala (Penttinen) variant. Along with previously reported individuals, our cohort suggests infants and young children had few abnormal features, while older individuals had multiple additional features, several of which appeared to worsen with advancing age. Our analysis supports a diagnostic entity of a spectrum disorders due to activating variants in PDGFRB. Differences in reported phenotypes can be dramatic and correlate with advancing age, genotype, and to mosaicism in some individuals.


Assuntos
Mesilato de Imatinib/uso terapêutico , Leucoencefalopatias/etiologia , Miofibromatose/congênito , Receptor beta de Fator de Crescimento Derivado de Plaquetas/genética , Adolescente , Adulto , Aneurisma/genética , Criança , Feminino , Estudos de Associação Genética , Humanos , Lactente , Leucoencefalopatias/tratamento farmacológico , Leucoencefalopatias/genética , Masculino , Miofibromatose/tratamento farmacológico , Miofibromatose/etiologia , Miofibromatose/genética , Linhagem , Inibidores de Proteínas Quinases/uso terapêutico
12.
Mol Genet Metab Rep ; 24: 100607, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32489882

RESUMO

Serine, a non-essential amino acid, has attracted clinical attention because of potential benefit in certain metabolic and neurological disorders. Despite the therapeutic potential, little is known about the pharmacokinetics of l-serine metabolism in humans. Here we present pharmacokinetic data at the time of treatment initiation as well as plasma serine levels during dose escalation from a single individual taking oral l-serine as part of a treatment regimen. Our results show that plasma serine levels rise and fall rapidly after oral l-serine intake, suggesting that the optimal dosing for oral l-serine supplementation is at least three times per day.

13.
BMC Biol ; 18(1): 3, 2020 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-31915011

RESUMO

BACKGROUND: The lion (Panthera leo) is one of the most popular and iconic feline species on the planet, yet in spite of its popularity, the last century has seen massive declines for lion populations worldwide. Genomic resources for endangered species represent an important way forward for the field of conservation, enabling high-resolution studies of demography, disease, and population dynamics. Here, we present a chromosome-level assembly from a captive African lion from the Exotic Feline Rescue Center (Center Point, IN) as a resource for current and subsequent genetic work of the sole social species of the Panthera clade. RESULTS: Our assembly is composed of 10x Genomics Chromium data, Dovetail Hi-C, and Oxford Nanopore long-read data. Synteny is highly conserved between the lion, other Panthera genomes, and the domestic cat. We find variability in the length of runs of homozygosity across lion genomes, indicating contrasting histories of recent and possibly intense inbreeding and bottleneck events. Demographic analyses reveal similar ancient histories across all individuals during the Pleistocene except the Asiatic lion, which shows a more rapid decline in population size. We show a substantial influence on the reference genome choice in the inference of demographic history and heterozygosity. CONCLUSIONS: We demonstrate that the choice of reference genome is important when comparing heterozygosity estimates across species and those inferred from different references should not be compared to each other. In addition, estimates of heterozygosity or the amount or length of runs of homozygosity should not be taken as reflective of a species, as these can differ substantially among individuals. This high-quality genome will greatly aid in the continuing research and conservation efforts for the lion, which is rapidly moving towards becoming a species in danger of extinction.


Assuntos
Genoma , Leões/genética , Animais , Feminino , Leões/classificação , Sintenia
14.
Am J Med Genet A ; 182(3): 543-547, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31814264

RESUMO

We describe a 5-year-old male with developmental delay, behavioral problems, and dysmorphic features who was found by microarray to have a 93-kb duplication of uncertain significance that fully encompasses the third exon of CTNND2 (delta catenin). Mate-pair sequencing was used to determine that the duplication is tandem and is predicted to lead to CTNND2 haploinsufficiency. Haploinsufficiency for CTNND2 has been shown to result in developmental delay and intellectual disability, providing a unifying diagnosis for this patient. His features overlap those associated with the larger cri-du-chat deletion of this region, expanding the clinical phenotype of isolated CTNND2 variants. The use of mate-pair sequencing to determine the orientation of the small duplication was essential to the diagnosis and avoided the use of exome sequencing, which would not have defined the arrangement of the duplication. This is only the second reported patient, to our knowledge, with a single exon duplication of CTNND2.


Assuntos
Anormalidades Múltiplas/genética , Transtorno Autístico/genética , Cateninas/genética , Predisposição Genética para Doença , Atrofia Muscular/genética , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/patologia , Transtorno Autístico/diagnóstico , Transtorno Autístico/patologia , Criança , Pré-Escolar , Éxons/genética , Facies , Feminino , Genes Duplicados/genética , Haploinsuficiência/genética , Humanos , Masculino , Atrofia Muscular/diagnóstico , Atrofia Muscular/patologia , Linhagem , Comportamento Problema
15.
Am J Med Genet A ; 182(3): 437-440, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31833187

RESUMO

Catel-Manzke syndrome is characterized by hand anomalies, Robin sequence, cardiac defects, joint hyperextensibility, and characteristic facial features. Approximately 40 patients with Catel-Manzke have been reported, all with the pathognomonic bilateral or unilateral hyperphalangy caused by an accessory bone between the second metacarpal and proximal phalanx known as Manzke dysostosis. Here we present the first case of molecularly confirmed Catel-Manzke syndrome with Robin sequence but without Manzke dysostosis.


Assuntos
Deformidades Congênitas da Mão/genética , Hidroliases/genética , Disostose Mandibulofacial/genética , Síndrome de Pierre Robin/genética , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Anormalidades Múltiplas/patologia , Adolescente , Criança , Pré-Escolar , Feminino , Deformidades Congênitas da Mão/diagnóstico , Deformidades Congênitas da Mão/patologia , Humanos , Disostose Mandibulofacial/diagnóstico , Disostose Mandibulofacial/patologia , Mutação/genética , Síndrome de Pierre Robin/diagnóstico , Síndrome de Pierre Robin/patologia
16.
G3 (Bethesda) ; 10(2): 525-537, 2020 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-31882405

RESUMO

Genetic stability depends on the maintenance of a variety of chromosome structures and the precise repair of DNA breaks. During meiosis, programmed double-strand breaks (DSBs) made in prophase I are normally repaired as gene conversions or crossovers. DSBs can also be made by other mechanisms, such as the movement of transposable elements (TEs), which must also be resolved. Incorrect repair of these DNA lesions can lead to mutations, copy-number changes, translocations, and/or aneuploid gametes. In Drosophila melanogaster, as in most organisms, meiotic DSB repair occurs in the presence of a rapidly evolving multiprotein structure called the synaptonemal complex (SC). Here, whole-genome sequencing is used to investigate the fate of meiotic DSBs in D. melanogaster mutant females lacking functional SC, to assay for de novo CNV formation, and to examine the role of the SC in transposable element movement in flies. The data indicate that, in the absence of SC, copy-number variation still occurs and meiotic DSB repair by gene conversion occurs infrequently. Remarkably, an 856-kilobase de novo CNV was observed in two unrelated individuals of different genetic backgrounds and was identical to a CNV recovered in a previous wild-type study, suggesting that recurrent formation of large CNVs occurs in Drosophila. In addition, the rate of novel TE insertion was markedly higher than wild type in one of two SC mutants tested, suggesting that SC proteins may contribute to the regulation of TE movement and insertion in the genome. Overall, this study provides novel insight into the role that the SC plays in genome stability and provides clues as to why the sequence, but not structure, of SC proteins is rapidly evolving.


Assuntos
Elementos de DNA Transponíveis , Drosophila melanogaster/genética , Complexo Sinaptonêmico/genética , Animais , Quebras de DNA de Cadeia Dupla , Variações do Número de Cópias de DNA , Reparo do DNA , Feminino , Masculino
17.
PLoS Genet ; 15(11): e1008421, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31697682

RESUMO

Balancer chromosomes are multiply inverted and rearranged chromosomes that are widely used in Drosophila genetics. First described nearly 100 years ago, balancers are used extensively in stock maintenance and complex crosses. Recently, the complete molecular structures of several commonly used balancers were determined by whole-genome sequencing. This revealed a surprising amount of variation among balancers derived from a common progenitor, identified genes directly affected by inversion breakpoints, and cataloged mutations shared by balancers. These studies emphasized that it is important to choose the optimal balancer, because different inversions suppress meiotic recombination in different chromosomal regions. In this review, we provide a brief history of balancers in Drosophila, discuss how they are used today, and provide examples of unexpected recombination events involving balancers that can lead to stock breakdown.


Assuntos
Inversão Cromossômica/genética , Cromossomos de Insetos/genética , Heterocromatina/genética , Recombinação Homóloga/genética , Animais , Centrômero/genética , Drosophila melanogaster/genética , Genoma de Inseto/genética , Fenótipo , Deleção de Sequência/genética , Cromossomo X/genética
18.
Genetics ; 210(4): 1197-1212, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30249684

RESUMO

The number of chromosomes carried by an individual species is one of its defining characteristics. Some species, however, can also carry supernumerary chromosomes referred to as B chromosomes. B chromosomes were recently identified in a laboratory stock of Drosophila melanogaster-an established model organism with a wealth of genetic and genomic resources-enabling us to subject them to extensive molecular analysis. We isolated the B chromosomes by pulsed-field gel electrophoresis and determined their composition through next-generation sequencing. Although these B chromosomes carry no known euchromatic sequence, they are rich in transposable elements and long arrays of short nucleotide repeats, the most abundant being the uncharacterized AAGAT satellite repeat. Fluorescent in situ hybridization on metaphase chromosome spreads revealed this repeat is located on chromosome 4, strongly suggesting the origin of the B chromosomes is chromosome 4 Cytological and quantitative comparisons of signal intensity between chromosome 4 and the B chromosomes supports the hypothesis that the structure of the B chromosome is an isochromosome. We also report the identification of a new B chromosome variant in a related laboratory stock. This B chromosome has a similar repeat signature as the original but is smaller and much less prevalent. We examined additional stocks with similar genotypes and did not find B chromosomes, but did find these stocks lacked the AAGAT satellite repeat. Our molecular characterization of D. melanogaster B chromosomes is the first step toward understanding how supernumerary chromosomes arise from essential chromosomes and what may be necessary for their stable inheritance.


Assuntos
Cromossomos de Insetos/genética , Drosophila melanogaster/genética , Evolução Molecular , Animais , Centrômero/genética , Mapeamento Cromossômico , Elementos de DNA Transponíveis/genética , Especiação Genética , Genômica , Heterocromatina/genética , Hibridização in Situ Fluorescente , Sequências Repetitivas de Ácido Nucleico/genética
19.
Curr Biol ; 28(18): 2984-2990.e3, 2018 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-30174188

RESUMO

Crossovers (COs) are formed during meiosis by the repair of programmed DNA double-strand breaks (DSBs) and are required for the proper segregation of chromosomes. More DSBs are made than COs, and the remaining DSBs are repaired as noncrossovers (NCOs). The distribution of recombination events along a chromosome occurs in a stereotyped pattern that is shaped by CO-promoting and CO-suppressing forces, collectively referred to as crossover patterning mechanisms. Chromosome inversions are structural aberrations that, when heterozygous, disrupt the recombination landscape by suppressing crossing over. In Drosophila species, the local suppression of COs by heterozygous inversions triggers an increase in crossing over on freely recombining chromosomes termed the interchromosomal (IC) effect [1, 2]. The molecular mechanism(s) by which heterozygous inversions suppress COs, whether noncrossover gene conversions (NCOGCs) are similarly affected, and what mediates the increase in COs in the rest of the genome remain open questions. By sequencing whole genomes of individual offspring from mothers containing heterozygous inversions, we show that, although COs are suppressed by inversions, NCOGCs occur throughout inversions at higher than wild-type frequencies. We confirm that CO frequency increases on the freely recombining chromosomes, yet CO interference remains intact. Intriguingly, NCOGCs do not increase in frequency on the freely recombining chromosomes and the total number of DSBs is approximately the same per genome. Together, our data show that heterozygous inversions change the recombination landscape by altering the relative proportions of COs and NCOGCs and suggest that DSB fate may be plastic until a CO assurance checkpoint has been satisfied.


Assuntos
Inversão Cromossômica , Drosophila melanogaster/genética , Genoma de Inseto , Recombinação Genética , Animais , Troca Genética , Quebras de DNA de Cadeia Dupla , Heterozigoto , Meiose/genética
20.
G3 (Bethesda) ; 8(10): 3131-3141, 2018 10 03.
Artigo em Inglês | MEDLINE | ID: mdl-30087105

RESUMO

The Drosophila genus is a unique group containing a wide range of species that occupy diverse ecosystems. In addition to the most widely studied species, Drosophila melanogaster, many other members in this genus also possess a well-developed set of genetic tools. Indeed, high-quality genomes exist for several species within the genus, facilitating studies of the function and evolution of cis-regulatory regions and proteins by allowing comparisons across at least 50 million years of evolution. Yet, the available genomes still fail to capture much of the substantial genetic diversity within the Drosophila genus. We have therefore tested protocols to rapidly and inexpensively sequence and assemble the genome from any Drosophila species using single-molecule sequencing technology from Oxford Nanopore. Here, we use this technology to present highly contiguous genome assemblies of 15 Drosophila species: 10 of the 12 originally sequenced Drosophila species (ananassae, erecta, mojavensis, persimilis, pseudoobscura, sechellia, simulans, virilis, willistoni, and yakuba), four additional species that had previously reported assemblies (biarmipes, bipectinata, eugracilis, and mauritiana), and one novel assembly (triauraria). Genomes were generated from an average of 29x depth-of-coverage data that after assembly resulted in an average contig N50 of 4.4 Mb. Subsequent alignment of contigs from the published reference genomes demonstrates that our assemblies could be used to close over 60% of the gaps present in the currently published reference genomes. Importantly, the materials and reagents cost for each genome was approximately $1,000 (USD). This study demonstrates the power and cost-effectiveness of long-read sequencing for genome assembly in Drosophila and provides a framework for the affordable sequencing and assembly of additional Drosophila genomes.


Assuntos
Drosophila/genética , Genoma , Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Animais , Biologia Computacional/métodos , Genômica/métodos , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala/economia , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Mutação INDEL , Nanoporos , Filogenia , Análise de Sequência de DNA
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