Principles of genome evolution in the Drosophila melanogaster species group.

That closely related species often differ by chromosomal inversions was discovered by Sturtevant and Plunkett in 1926. Our knowledge of how these inversions originate is still very limited, although a prevailing view is that they are facilitated by ectopic recombination events between inverted repetitive sequences. The availability of genome sequences of related species now allows us to study in detail the mechanisms that generate interspecific inversions. We have analyzed the breakpoint regions of the 29 inversions that differentiate the chromosomes of Drosophila melanogaster and two closely related species, D. simulans and D. yakuba, and reconstructed the molecular events that underlie their origin. Experimental and computational analysis revealed that the breakpoint regions of 59% of the inversions (17/29) are associated with inverted duplications of genes or other nonrepetitive sequences. In only two cases do we find evidence for inverted repetitive sequences in inversion breakpoints. We propose that the presence of inverted duplications associated with inversion breakpoint regions is the result of staggered breaks, either isochromatid or chromatid, and that this, rather than ectopic exchange between inverted repetitive sequences, is the prevalent mechanism for the generation of inversions in the melanogaster species group. Outgroup analysis also revealed evidence for widespread breakpoint recycling. Lastly, we have found that expression domains in D. melanogaster may be disrupted in D. yakuba, bringing into question their potential adaptive significance

Higher order chromosome organization and recombination dynamics of meiotic prophase I in mouse spermatocytes

Meiotic recombination is required for parental chromosomes to find each other (pairing/synapsis) and to exchange genetic information thus allowing faithful segregation of chromosomes and the production of haploid gametes. At the start of meiotic prophase I, meiotic chromosomes organize into loop arrays that extrude out of the chromosome axis. Then, a large number of programmed double-strand breaks (DSBs) are formed at specific chromosomal locations or “hotspots” on parental chromosomes, which are repaired by homologous recombination (HR). HR produces either crossovers, which result in the exchange of flanking markers between homologs, or noncrossovers, which are short regions ofgene conversion to the donor genotype. Crossover formation is critical for proper chromosome segregation and crossovers arise from crossover precursors that form at a subset of DSBs that are designated to become future crossovers. Our current understanding of meiotic progression in mammals is largely derived from cytological observation.Many semi-redundant HR pathways can repair meiotic DSBs; however, the time at which different pathways are active, how the pathways interact, and the relative contribution of each pathway towards maintaining germline genomic integrity are poorly understood in vivoat endogenous sites, especially in a mammalian system. More importantly, how germline genome integrity is ensured at both the DNA level by recombination activity and by higher order chromosome structural changes has not been defined. Failure to maintain germline genome integrity can lead to aneuploidy, genetic disorders, birth defects and miscarriages. To define and dissect the temporal dynamics of different HR pathways and chromosome organization in vivo, I have established a novel and robust system to synchronize mouse spermatogenesis in F1 hybrid mice using the inhibitor WIN 18,446. My synchronizationprotocol allows the isolation of cells at specific stages of meiotic prophase I by flow cytometry, allowing me to analyze recombination outcomes at two meiotic hotspots and perform genome-wide Hi-C, a chromosome conformation capture method combined with high-throughput sequencing to investigate changes in higher order chromosome architecture during prophase I. Here, I provide the first direct molecular evidence that HR pathways that lead to to distinct meiotic outcomes aretemporally regulated. I have identified two novel classes of noncrossover pathways: 1) one that likely regulates the pairing/synapsis of parental chromosomes during early prophase I; and 2) one that derives from the crossover/noncrossover decision during mid-prophase I. My data show that crossover formation is suppressed until full synapsis is achieved at mid-prophase I, suggesting a previously unknown mechanism that prevents deleterious premature recombination. In addition, I show that alternative repair pathways are not activated until late prophase I, thus preventing designated crossover precursors from inappropriately forming noncrossovers. Furthermore, the Hi-C data I present provides evidence for dynamic genome reorganization during meiotic prophase I. There is evidence for loop array formation and loop extrusion as chromosomes condense. While topologically associating domains disappear at the onset of meiotic prophase I, chromosome compartments are well maintained. Most meiotic DSBs occur within a gene-dense open compartment A, suggesting that higher order chromosome structure plays an important role in meiotic recombination. Finally, interhomolog interactions and specialized chromosomal architecture in regions of pairing and synapsis could be inferred. Taken together, my data reveals that both chromosome recombination and chromosome structure are highly regulated to ensure chromosome pairing and segregation. These results provide important, novel insights to the field of meiosis and our understanding of germline genomic integrity and mammalian reproductive health

Exploring the Mechanism of Meiosis in \u3cem\u3eDrosophila melanogaster\u3c/em\u3e: Meiotic Functions of a Novel Cohesion Protein SOLO and a Translation Initiation Factor VASA

Sister chromatid cohesion is essential for proper chromosome segregation during meiosis. However, the mechanism of meiotic cohesion in Drosophila is unclear. We describe a novel protein, SOLO (Sisters On the LOose) that is essential for meiotic cohesion in Drosophila melanogaster. solo mutations cause high nondisjunction of sister and homologous chromatids of sex chromosomes and autosomes in both sexes. In solo males, sister chromatids separate prematurely and segregate randomly during meiosis II. Although bivalents appear intact throughout meiosis I, sister centromeres lose cohesion prior to prometaphase I and orient nearly randomly on the meiosis I spindle. Centromeric foci of SMC1 are absent in solo males at all meiotic stages. SOLO and the cohesin protein SMC1 co-localize to meiotic centromeres from early prophase I until anaphase II in wild-type males but both proteins are removed prematurely from centromeres at anaphase I in mei-S332 mutants, coincident with premature loss of cohesion in those mutants. solo mutations in females cause reduced frequency of homologous recombination between X chromosomes and autosomes, partially due to the loss of inhibition of sister chromatid exchange. Synaptonemal complex assembly is severely disrupted in early meiotic stage in solo females. SOLO colocalizes with SMC1 and C(3)G in meiosis. Additionally, SOLO is required for stabilizing chiasmata generated from residual recombination events. The data about the phenotypes of solo males and females and colocalization patterns of SOLO strongly suggest SOLO is a component of potential cohesin in Drosophila meiosis. Drosophila males undergo meiosis without recombination. However, the underlying mechanism is not known. Mutations of vasa cause high frequency of X-Y exchange in meiosis. Chromatin bridges at anaphase I and II, due to dicentric recombination events, were observed in vasa males. vas and solo double mutant showed precocious segregation of homologs at metaphase I besides chromatin bridge at anaphase I and II. Our data thus for the first time demonstrate that inhibition of meiotic recombination during male meiosis requires vas function and interactions between vas and solo regulate chromosome dynamics in male meiosis

The effects of the promotor region of the 240bp repeats of the rRNA genes on x-y chromosome disjunction in Drosophila melanogaster males

Pairing between homologous chromosomes is essential for successful meiosis. In Drosophila melanogaster males, sex chromosome pairing during meiosis I is mediated by rDNA, located in heterochromatin. Several analyses of rDNA fragments showed that 240bp repeats in the intergenic spacer (IGS) have the ability to stimulate X-Y chromosome pairing and disjunction. In addition, point mutations within the promoter of the 240bp repeats failed to mediate X-Y chromosome pairing and disjunction. These previous studies imply that promoter activity of the 240bp repeats is involved in X-Y chromosome pairing in Drosophila males. In this study, I made a construct composed of 16 copies of the 72bp fragment within the 240bp repeat, which has promoter activity and obtained transformant lines with the construct. The construct was transferred to Df(1)X-1, an rDNA deficient X chromosome, by recombination. The effect of the transgene on the frequency of X-Y disjunction were analyzed both by cytological and genetic experiments. The transgene in Df(1)X-1 chromosome induced increased X-Y chromosome disjunction frequency. The result indicates that promoter activity of the 240bp repeats may be responsible for X-Y chromosome pairing in Drosophila males

Neighbourhood continuity is not required for correct testis gene expression in Drosophila.

It is now widely accepted that gene organisation in eukaryotic genomes is non-random and it is proposed that such organisation may be important for gene expression and genome evolution. In particular, the results of several large-scale gene expression analyses in a range of organisms from yeast to human indicate that sets of genes with similar tissue-specific or temporal expression profiles are clustered within the genome in gene expression neighbourhoods. While the existence of neighbourhoods is clearly established, the underlying reason for this facet of genome organisation is currently unclear and there is little experimental evidence that addresses the genomic requisites for neighbourhood organisation. We report the targeted disruption of three well-defined male-specific gene expression neighbourhoods in the Drosophila genome by the synthesis of precisely mapped chromosomal inversions. We compare gene expression in individuals carrying inverted chromosomes with their non-inverted but otherwise identical progenitors using whole-transcriptome microarray analysis, validating these data with specific quantitative real-time PCR assays. For each neighbourhood we generate and examine multiple inversions. We find no significant differences in the expression of genes that define each of the neighbourhoods. We further show that the inversions spatially separate both halves of a neighbourhood in the nucleus. Thus, models explaining neighbourhood organisation in terms of local sequence interactions, enhancer crosstalk, or short-range chromatin effects are unlikely to account for this facet of genome organisation. Our study challenges the notion that, at least in the case of the testis, expression neighbourhoods are a feature of eukaryotic genome organisation necessary for correct gene expression

Una vista panorámica de los cromosomas en la profase I de la meiosis

The present review aims to summarize the research carried out in relation to meiosis in birds, especially by observing the protein axes of the chromosomes in prophase I of meiosis. This line of research, initially developed in Argentina, has provided key data in the study of the evolution of sex chromosomes and the mechanisms involved in the frequency and distribution of crossing over in birds, among other topics. Some of these contributions, in addition to those made by other authors, are described also providing the general theoretical framework or the hypotheses that support them.La presente revisión tiene por objetivo resumir las investigaciones realizadas en relación a la meiosis de las aves, especialmente mediante la observación de los ejes proteicos de los cromosomas en la profase I de la meiosis. Esta línea de investigación, desarrollada inicialmente en Argentina, ha aportado datos clave dentro del estudio de la evolución de los cromosomas sexuales y los mecanismos involucrados en la frecuencia y distribución del crossing over en las aves, entre otros temas. Algunas de estas contribuciones, además de las realizadas por otros autores, se describen proporcionando también el marco teórico general o las hipótesis que las sustentan.Fil: Pigozzi, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; Argentin

Holocentric plants of the genus Rhynchospora as a new model to study meiotic adaptations to chromosomal structural rearrangements

Climate change, world hunger and overpopulation are some of the biggest challenges the world is currently facing. Moreover, they are part of a multidimensional single scenario: as climate change continues to modify our planet, we might see a decrease of arable land and increase in extreme weather patterns, posing a threat to food security. This has a direct impact on regions with high population growth, where food security is already scarce. Considering additionally the unsustainability of intensive global food production and its contribution to greenhouse emissions and biodiversity loss, it´s clear that all these factors are interconnected (Cardinale et al., 2012; Prosekov & Ivanova, 2018; Wiebe et al., 2019). Plants are the main source of staple food in the world and are also the main actors in carbon fixation, they are therefore key protagonists in controlling climate change. Plants are also an essential habitat-defining element balancing our ecosystem. Thus, how we grow plants and crops will, aside from the obvious implications for food security, also have a profound impact on the climate and biodiversity. The natural variability of species is considered an immense pool of genes and traits, and their understanding is key to generate new useful knowledge. For instance, natural populations can be more tolerant to abiotic and biotic stresses, or carry traits that combined together in hybrids, might achieve a higher seed number, or a faster growth. Classical breeding has exploited unrelated varieties to achieve traits of interest like dwarfism and higher grain production. However, only a limited number of crop species have been the focus of recent scientific and technological approaches, and they do not represent the extremely vast natural diversity of species that could generate useful knowledge for future applications (Castle et al., 2006; Pingali, 2012). The key to this natural variability is a process called meiotic recombination, the exchange of genomic material between homologous parental chromosomes. Meiotic recombination takes place during meiosis, a specialized cell division in which sexually reproducing organisms reduce the genomic complement of their gametes by half in preparation for fertilization. Meiotic recombination takes place at the beginning of meiosis, in a stage called prophase I. To exchange DNA sequences, the strands of two homologous chromosomes must be fragmented. This specific process of physiologically induced DNA fragmentation is conserved in the vast majority of eukaryotes (Keeney et al., 1997). After the formation of double-strand breaks, the 3’ ends that are left are targeted by recombinases that help the strands search and invade templates for repair. After invasion, the 3’ end is extended by DNA synthesis, exposing sequences on the opposite strand that can anneal to the other 3’ end of the original double strand break. DNA synthesis at both ends generates a new structure called a double Holliday Junction (dHJ), forming a physical link between homologous chromosomes, named chiasma (Wyatt & West, 2014). The resolutions of these structures are called crossovers (COs), which is the molecular event representing the outcome of meiotic recombination. Other outcomes are possible, like noncrossovers (NCOs). In this case, the invading strand is ejected and anneals to the single-strand 3´end of the original double-strand break (Allers & Lichten, 2001). Crossovers can be divided into two main groups, called class I and class II. COs of the first group are considered to be sensitive to interference, which means that there are mechanisms that prevent two class I COs from happening in proximity of each other. Class II is insensitive to interference. Class I COs are the result of a pathway called ZMM, which involves a group of specialised proteins that are highly conserved among eukaryotes (Lambing et al., 2017; Mercier et al., 2015). Class I COs are the most common, studied and important type of COs. Centromeres are structures, located on regions of the chromosomes, that allow proper chromosome segregation during mitosis and meiosis. Centromeres have a profound effect on plant breeding and crop improvement, as it is known that meiotic recombination is suppressed at centromeres in most eukaryotes. This represents a great limitation for crop improvement, as many possibly useful traits might be in regions not subject to recombination and thus might not be available for breeding purposes. Additionally, the mechanisms behind how recombination is regulated and prevented from happening at centromeres are still unclear. In most model organisms centromeres are single entities localized on specific regions on the chromosomes. This configuration is called monocentric. However, another type of configuration can be found in nature, but is less studied. In fact, some organisms harbour multiple centromeric determinants distributed over their whole chromosomal length. This configuration is called holocentric. The Cyperaceae comprise a vast, diverse family of plants, with a cosmopolitan distribution in all habitats (Spalink et al., 2016). Despite the presence of this family worldwide, knowledge about it is limited. Few genomes are available and molecular insights are scarce. This family is also known to be mainly formed by holocentric species (Melters et al., 2012). Understanding if and how meiotic recombination is achieved in holocentric plants will generate new knowledge that in the future might unlock new traits in elite crops, previously unavailable to breeding, that could help humanity face global climatic, economic and social challenges. Recent studies have reported new knowledge about important meiotic, chromosome and genome adaptions found in species of the Cyperaceae family and in particular the genus Rhynchospora (Marques et al., 2015, 2016a). With the recent publication of the first reference genomes for several Rhynchospora species, we could already perform a comprehensive analysis of their unique genome features and trace the evolutionary history of their karyotypes and how these have been determined by chromosome fusions (Hofstatter et al., 2021, 2022). This new resource paves the way for future research utilising Rhynchospora as a model genus to study adaptations to holocentricity in plants. With this work, my intention is to shed light on the underexplored topic of holocentricity in plants. Using cutting edge techniques, I examine the conservation of meiotic recombination together with other species-specific adaptations like achiasmy and polyploidy in holocentrics. My results reveal new insights into how plant meiotic recombination is regulated when small centromere units are found distributed chromosome-wide, challenging the classic dogma of suppression of recombination at centromeres

The Transposon Galileo Generates Natural Chromosomal Inversions in Drosophila by Ectopic Recombination

Background: transposable elements (TEs) are responsible for the generation of chromosomal inversions in several groups of organisms. However, in Drosophila and other Dipterans, where inversions are abundant both as intraspecific polymorphisms and interspecific fixed differences, the evidence for a role of TEs is scarce. Previous work revealed that the transposon Galileo was involved in the generation of two polymorphic inversions of Drosophila buzzatii. Methodology/Principal Findings: to assess the impact of TEs in Drosophila chromosomal evolution and shed light on the mechanism involved, we isolated and sequenced the two breakpoints of another widespread polymorphic inversion from D. buzzatii, 2z3. In the non inverted chromosome, the 2z3 distal breakpoint was located between genes CG2046 and CG10326 whereas the proximal breakpoint lies between two novel genes that we have named Dlh and Mdp. In the inverted chromosome, the analysis of the breakpoint sequences revealed relatively large insertions (2,870-bp and 4,786-bp long) including two copies of the transposon Galileo (subfamily Newton), one at each breakpoint, plus several other TEs. The two Galileo copies: (i) are inserted in opposite orientation; (ii) present exchanged target site duplications; and (iii) are both chimeric. Conclusions/Significance: our observations provide the best evidence gathered so far for the role of TEs in the generation of Drosophila inversions. In addition, they show unequivocally that ectopic recombination is the causative mechanism. The fact that the three polymorphic D. buzzatii inversions investigated so far were generated by the same transposon family is remarkable and is conceivably due to Galileo's unusual structure and current (or recent) transpositional activity

Recurrent insertion and duplication generate networks of transposable element sequences in the Drosophila melanogaster genome.

BACKGROUND: The recent availability of genome sequences has provided unparalleled insights into the broad-scale patterns of transposable element (TE) sequences in eukaryotic genomes. Nevertheless, the difficulties that TEs pose for genome assembly and annotation have prevented detailed, quantitative inferences about the contribution of TEs to genomes sequences. RESULTS: Using a high-resolution annotation of TEs in Release 4 genome sequence, we revise estimates of TE abundance in Drosophila melanogaster. We show that TEs are non-randomly distributed within regions of high and low TE abundance, and that pericentromeric regions with high TE abundance are mosaics of distinct regions of extreme and normal TE density. Comparative analysis revealed that this punctate pattern evolves jointly by transposition and duplication, but not by inversion of TE-rich regions from unsequenced heterochromatin. Analysis of genome-wide patterns of TE nesting revealed a 'nesting network' that includes virtually all of the known TE families in the genome. Numerous directed cycles exist among TE families in the nesting network, implying concurrent or overlapping periods of transpositional activity. CONCLUSION: Rapid restructuring of the genomic landscape by transposition and duplication has recently added hundreds of kilobases of TE sequence to pericentromeric regions in D. melanogaster. These events create ragged transitions between unique and repetitive sequences in the zone between euchromatic and beta-heterochromatic regions. Complex relationships of TE nesting in beta-heterochromatic regions raise the possibility of a co-suppression network that may act as a global surveillance system against the majority of TE families in D. melanogaster.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are