Monocentric Chromosomes Research Articles

Repeat-based holocentromeres of the woodrush Luzula sylvatica reveal insights into the evolutionary transition to holocentricity

In most studied eukaryotes, chromosomes are monocentric, with centromere activity confined to a single region. However, the rush family (Juncaceae) includes species with both monocentric (Juncus) and holocentric (Luzula) chromosomes, where centromere activity is distributed along the entire chromosome length. Here, we combine chromosome-scale genome assembly, epigenetic analysis, immuno-FISH and super-resolution microscopy to study the transition to holocentricity in Luzula sylvatica. We report repeat-based holocentromeres with an irregular distribution of features along the chromosomes. Luzula sylvatica holocentromeres are predominantly associated with two satellite DNA repeats (Lusy1 and Lusy2), while CENH3 also binds satellite-free gene-poor regions. Comparative repeat analysis suggests that Lusy1 plays a crucial role in centromere function across most Luzula species. Furthermore, synteny analysis between L. sylvatica (n = 6) and Juncus effusus (n = 21) suggests that holocentric chromosomes in Luzula could have arisen from chromosome fusions of ancestral monocentric chromosomes, accompanied by the expansion of CENH3-associated satellite repeats.

How diverse a monocentric chromosome can be? Repeatome and centromeric organization of Juncus effusus (Juncaceae).

Juncus is the largest genus of Juncaceae and was considered holocentric for a long time. Recent findings, however, indicated that 11 species from different clades of the genus have monocentric chromosomes. Thus, the Juncus centromere organization and evolution need to be reassessed. We aimed to investigate the major repetitive DNA sequences of two accessions of Juncus effusus and its centromeric structure by employing whole-genome analyses, fluorescent in situ hybridization, CENH3 immunodetection, and chromatin immunoprecipitation sequencing. We showed that the repetitive fraction of the small J. effusus genome (~270 Mbp/1C) is mainly composed of Class I and Class II transposable elements (TEs) and satellite DNAs. Three identified satellite DNA families were mainly (peri)centromeric, with two being associated with the centromeric protein CENH3, but not strictly centromeric. Two types of centromere organization were discerned in J. effusus: type 1 was characterized by a single CENH3 domain enriched with JefSAT1-155 or JefSAT2-180, whereas type 2 showed multiple CENH3 domains interrupted by other satellites, TEs or genes. Furthermore, while type 1 centromeres showed a higher degree of satellite identity along the array, type 2 centromeres had less homogenized arrays along the multiple CENH3 domains per chromosome. Although the analyses confirmed the monocentric organization of J. effusus chromosomes, our data indicate a more dynamic arrangement of J. effusus centromeres than observed for other plant species, suggesting it may constitute a transient state between mono- and holocentricity.

Regional centromere configuration in the fungal pathogens of the Pneumocystis genus.

Centromeres are constricted chromosomal regions that are essential for cell division. In eukaryotes, centromeres display a remarkable architectural and genetic diversity. The basis of centromere-accelerated evolution remains elusive. Here, we focused on Pneumocystis species, a group of mammalian-specific fungal pathogens that form a sister taxon with that of the Schizosaccharomyces pombe, an important genetic model for centromere biology research. Methods allowing reliable continuous culture of Pneumocystis species do not currently exist, precluding genetic manipulation. CENP-A, a variant of histone H3, is the epigenetic marker that defines centromeres in most eukaryotes. Using heterologous complementation, we show that the Pneumocystis CENP-A ortholog is functionally equivalent to CENP-ACnp1 of S. pombe. Using organisms from a short-term in vitro culture or infected animal models and chromatin immunoprecipitation (ChIP)-Seq, we identified CENP-A bound regions in two Pneumocystis species that diverged ~35 million years ago. Each species has a unique short regional centromere (<10 kb) flanked by heterochromatin in 16-17 monocentric chromosomes. They span active genes and lack conserved DNA sequence motifs and repeats. These features suggest an epigenetic specification of centromere function. Analysis of centromeric DNA across multiple Pneumocystis species suggests a vertical transmission at least 100 million years ago. The common ancestry of Pneumocystis and S. pombe centromeres is untraceable at the DNA level, but the overall architectural similarity could be the result of functional constraint for successful chromosomal segregation.IMPORTANCEPneumocystis species offer a suitable genetic system to study centromere evolution in pathogens because of their phylogenetic proximity with the non-pathogenic yeast S. pombe, a popular model for cell biology. We used this system to explore how centromeres have evolved after the divergence of the two clades ~ 460 million years ago. To address this question, we established a protocol combining short-term culture and ChIP-Seq to characterize centromeres in multiple Pneumocystis species. We show that Pneumocystis have short epigenetic centromeres that function differently from those in S. pombe.

Repositioning of centromere-associated repeats during karyotype evolution in Oryzias fishes.

The karyotype, which is the number and shape of chromosomes, is a fundamental characteristic of all eukaryotes. Karyotypic changes play an important role in many aspects of evolutionary processes, including speciation. In organisms with monocentric chromosomes, it was previously thought that chromosome number changes were mainly caused by centric fusions and fissions, whereas chromosome shape changes, that is, changes in arm numbers, were mainly due to pericentric inversions. However, recent genomic and cytogenetic studies have revealed examples of alternative cases, such as tandem fusions and centromere repositioning, found in the karyotypic changes within and between species. Here, we employed comparative genomic approaches to investigate whether centromere repositioning occurred during karyotype evolution in medaka fishes. In the medaka family (Adrianichthyidae), the three phylogenetic groups differed substantially in their karyotypes. The Oryzias latipes species group has larger numbers of chromosome arms than the other groups, with most chromosomes being metacentric. The O. javanicus species group has similar numbers of chromosomes to the O. latipes species group, but smaller arm numbers, with most chromosomes being acrocentric. The O. celebensis species group has fewer chromosomes than the other two groups and several large metacentric chromosomes that were likely formed by chromosomal fusions. By comparing the genome assemblies of O. latipes, O. javanicus, and O. celebensis, we found that repositioning of centromere-associated repeats might be more common than simple pericentric inversion. Our results demonstrated that centromere repositioning may play a more important role in karyotype evolution than previously appreciated.

Revision of the South American Ninetinae genus Guaranita (Araneae, Pholcidae)

The southern South American genus Guaranita includes tiny spiders (body length ~1 mm) that lead reclusive lives under ground-objects and run rapidly when disturbed. As a result, they have been poorly collected and studied. Here we report on a recent collection of Guaranita spiders from Argentina, describing one new species (G. auadae Huber sp. nov.) and the previously unknown female of G. dobby Torres et al., 2016. In addition, we provide CO1 barcodes for all (now five) known species, first SEM data, and first chromosome data for the genus. The diploid number of Guaranita goloboffi Huber, 2000 (2n♂ = 11) is among the lowest in araneomorph spiders with monocentric chromosome structure.

Holocentric chromosomes in animals and plants: what do we know about these points outside the curve?

Usually, members of the Eukarya domain have kinetochores grouped at a single point, characterizing monocentric chromosomes. However, in some taxa the protein apparatus that composes the kinetochore is distributed continuously or discretely along the length of the chromosome, condition that defines holocentric chromosomes. The review aims to provide an overview of karyomorphological aspects of holocentric chromosomes in animals and plants, as well as an understanding of their origin, evolution and possible adaptive implications of their presence in eukaryotes. The main structural differences between holocentric and monocentric chromosomes concern the kinetochore proteins, the pattern of histone H3 phosphorylation and centromeric satellite DNA. The distribution of holocentric chromosomes in phylogenetic trees evidence their independent emergence numerous times throughout evolution. While many hypotheses have been created to explain the origin of holocentric chromosomes, none have been confirmed or refuted. Although the adaptive advantages generated by their presence are undeniable, especially in clastogenic environments, the typical behavior of chromosomes with diffuse kinetochores seems to be enough to make them the exception rather than the rule among eukaryotes.

Microtubule search-and-capture model evaluates the effect of chromosomal volume conservation on spindle assembly during mitosis.

Variation in the chromosome numbers can arise from the erroneous mitosis or fusion and fission of chromosomes. While the mitotic errors lead to an increase or decrease in the overall chromosomal substance in the daughter cells, fission and fusion keep this conserved. Variations in chromosome numbers are assumed to be a crucial driver of speciation. For example, the members of the muntjac species are known to have very different karyotypes with the chromosome numbers varying from 2n=70+3B in the brown brocket deer to 2n=46 in the Chinese muntjac and 2n=6/7 in the Indian muntjac. The chromosomal content in the nucleus of these closely related mammals is roughly the same and various chromosome fusion and fission pathways have been suggested as the evolution process of these karyotypes. Similar trends can also be found in lepidoptera and yeast species which show a wide variation of chromosome numbers. The effect of chromosome number variation on the spindle assembly time and accuracy is still not properly addressed. We computationally investigate the effect of conservation of the total chromosomal substance on the spindle assembly during prometaphase. Our results suggest that chromosomal fusion pathways aid the microtubule-driven search and capture of the kinetochore in cells with monocentric chromosomes. We further report a comparative analysis of the site and percentage of amphitelic captures, dependence on cell shape, and position of the kinetochore in respect to chromosomal volume partitioning.

Lepidopteran Synteny Units reveal deep chromosomal conservation in butterflies and moths.

DNA is compacted into individual particles or chromosomes that form the basic units of inheritance. However, different animals and plants have widely different numbers of chromosomes. This means that we cannot readily tell which chromosomes are related to which. Here, we describe a simple technique that looks at the similarity of genes on each chromosome and thus gives us a true picture of their homology or similarity through evolutionary time. We use this new system to look at the chromosomes of butterflies and moths or Lepidoptera. We term the associated synteny units, Lepidopteran Synteny Units (LSUs). Using a sample of butterfly and moth genomes from across evolutionary time, we show that LSUs form a simple and reliable method of tracing chromosomal homology back through time. Surprisingly, this technique reveals that butterfly and moth chromosomes show conserved blocks dating back to their sister group the Trichoptera. As Lepidoptera have holocentric chromosomes, it will be interesting to see if similar levels of synteny are shown in groups of animals with monocentric chromosomes. The ability to define homology via LSU analysis makes it considerably easier to approach many questions in chromosomal evolution.

Holocentromeres can consist of merely a few megabase-sized satellite arrays

The centromere is the chromosome region where microtubules attach during cell division. In contrast to monocentric chromosomes with one centromere, holocentric species usually distribute hundreds of centromere units along the entire chromatid. We assembled the chromosome-scale reference genome and analyzed the holocentromere and (epi)genome organization of the lilioid Chionographis japonica. Remarkably, each of its holocentric chromatids consists of only 7 to 11 evenly spaced megabase-sized centromere-specific histone H3-positive units. These units contain satellite arrays of 23 and 28 bp-long monomers capable of forming palindromic structures. Like monocentric species, C. japonica forms clustered centromeres in chromocenters at interphase. In addition, the large-scale eu- and heterochromatin arrangement differs between C. japonica and other known holocentric species. Finally, using polymer simulations, we model the formation of prometaphase line-like holocentromeres from interphase centromere clusters. Our findings broaden the knowledge about centromere diversity, showing that holocentricity is not restricted to species with numerous and small centromere units.

Evolution of satDNAs on holocentric chromosomes: insights from hemipteran insects of the genus Mahanarva.

Satellite DNAs (satDNAs) constitute one of the main components of eukaryote genomes and are involved in chromosomal organization and diversification. Although largely studied, little information was gathered about their evolution on holocentric species, i.e., diffuse centromeres, which, due to differences in repeat organization, could result in different evolutionary patterns. Here, we combined bioinformatics and cytogenetic approaches to evaluate the evolution of the satellitomes in Mahanarva holocentric insects. In two species, de novo identification revealed a high number of satDNAs, 110 and 113, with an extreme monomer length range of 18-4228 bp. The overall abundance of satDNAs was observed to be 6.67% in M. quadripunctata and 1.98% in M. spectabilis, with different abundances for the shared satDNAs. Chromosomal mapping of the most abundant repeats of M. quadripunctata and M. spectabilis on other Mahanarva reinforced the dynamic nature of satDNAs. Variable patterns of chromosomal distribution for the satDNAs were noticed, with the occurrence of clusters on distinct numbers of chromosomes and at different positions and the occurrence of scattered signals or nonclustered satDNAs. Altogether, our data demonstrated the high dynamism of satDNAs in Mahanarva with the involvement of this genomic fraction in chromosome diversification of the genus. The general characteristics and patterns of evolution of satDNAs are similar to those observed on monocentric chromosomes, suggesting that the differential organization of genome compartments observed on holocentric chromosomes compared with monocentric chromosomes does not have a large impact on the evolution of satDNAs. Analysis of the satellitomes of other holocentric species in a comparative manner will shed light on this issue.

The first chromosome study of the genera Calchas Birula, 1899 and Neocalchas Yağmur, Soleglad, Fet & Kovařík, 2013 (Scorpiones: Iuridae)

The chromosomes of two species belonging to the scorpion family Iuridae are studied for the first time. Both analysed species displayed achiasmatic meiosis and no morphologically differentiated sex chromosomes; these features are typical for all scorpions. Both species possessed monocentric chromosomes, a characteristic specific to the entire parvorder Iurida. The karyotype of Calchas nordmanni Birula, 1899 is composed of 82 chromosomes, and the diploid number of chromosomes in Neocalchas gruberi is 40. Despite this conspicuous difference in 2n chromosome morphology is mainly acrocentric. Both species possess two pairs of the 18S rDNA clusters identified by FISH. A higher number of rDNA loci may represent an ancestral state for the parvorder Iurida.

Chromosome diversity in Buthidae and Chactidae scorpions from Brazilian fauna: Diploid number and distribution of repetitive DNA sequences.

In this work, we analyzed cytogenetically eight Chactidae and Buthidae, including the localization of repetitive DNA sequences. The chactids possess monocentric chromosomes and the highest diploid numbers (2n=50 in Brotheas amazonicus, 2n=36 in Chactopsis amazonica, 2n=30 in Neochactas sp.) when compared with buthids (2n=10 in Tityus bahiensis, 2n=14 in Tityus apiacas and Tityus metuendus, 2n=18 in Tityus aba, 2n=26 in Ischnotelson peruassu). The localization of rDNA genes and (TTAGG)n sequences exhibited a conserved pattern of two terminal/subterminal ribosomal cistrons and terminal telomere signals. However, the comparison between the data of C-banding, DAPI after FISH and Cot-DNA fraction indicated a variable quantity and distribution of these regions, as follow: (i) positive heterochromatin and Cot-DNA signals (B. amazonicus and I. peruassu), (ii) small blocks of heterochromatin with large Cot-DNA signals (T. metuendus), (iii) positive heterochromatic regions and absence of Cot-DNA signals (T. aba and T. apiacas), and (iv) negative heterochromatin and Cot-DNA signals (T. bahiensis). Therefore, our results revealed that there still is not a clear relation between quantity of heterochromatin and presence of monocentric or holocentric chromosomes and occurrence of chromosomal rearrangements, indicating that repetitive regions in scorpions must be analyzed using different cytogenetic approaches.

Only the Rye Derived Part of the 1BL/1RS Hybrid Centromere Incorporates CENH3 of Wheat.

The precise assembly of the kinetochore complex at the centromere is epigenetically determined by substituting histone H3 with the centromere-specific histone H3 variant CENH3 in centromeric nucleosomes. A chromosome 1B reconstructed in wheat by centric misdivision from two wheat-rye centric translocations is known to carry a hybrid wheat-rye centromere. The resulting hybrid (dicentric)centromere is composed of both wheat and rye centromeric repeats. As CENH3 is a marker for centromere activity, we applied Immuno-FISH followed by ultrastructural super-resolution microscopy to address whether both or only parts of the hybrid centromere are active. Our study demonstrates that only the rye-derived centromere part incorporates CENH3 of wheat in the 1BL/1RS hybrid centromere. This finding supports the notion that one centromere part of a translocated chromosome undergoes inactivation, creating functional monocentric chromosomes to maintain chromosome stability.

A simple model explains the cell cycle-dependent assembly of centromeric nucleosomes in holocentric species.

Centromeres are essential for chromosome movement. In independent taxa, species with holocentric chromosomes exist. In contrast to monocentric species, where no obvious dispersion of centromeres occurs during interphase, the organization of holocentromeres differs between condensed and decondensed chromosomes. During interphase, centromeres are dispersed into a large number of CENH3-positive nucleosome clusters in a number of holocentric species. With the onset of chromosome condensation, the centromeric nucleosomes join and form line-like holocentromeres. Using polymer simulations, we propose a mechanism relying on the interaction between centromeric nucleosomes and structural maintenance of chromosomes (SMC) proteins. Different sets of molecular dynamic simulations were evaluated by testing four parameters: (i) the concentration of Loop Extruders (LEs) corresponding to SMCs, (ii) the distribution and number of centromeric nucleosomes, (iii) the effect of centromeric nucleosomes on interacting LEs and (iv) the assembly of kinetochores bound to centromeric nucleosomes. We observed the formation of a line-like holocentromere, due to the aggregation of the centromeric nucleosomes when the chromosome was compacted into loops. A groove-like holocentromere structure formed after a kinetochore complex was simulated along the centromeric line. Similar mechanisms may also organize a monocentric chromosome constriction, and its regulation may cause different centromere types during evolution.

A new opilioacarid species (Parasitiformes: Opilioacarida) from Crete (Greece) with notes on its karyotype

A new species of opilioacarid mite, Opilioacarus thaleri n. sp., is described from mid-level elevation close to the seashore in Crete (Greece). It was found on moister sites, under stones or inside stone accumulations in phrygana vegetation. The peak of activity was found to be in early May. The diagnostic morphological characters include 7–8 eugenital setae (female), 5/7 setae on each sclerite at pregenital and genital areas, 11 setae on genital area (male), anterior dorsal shield with two pairs of eyes and 60–70 stout, ribbed setae, 5 (female) or 6 (male) leaf setae plus one pectinate seta on the palp tarsus, and the "simple" type of ovipositor with two pairs of glands, a pair of ducts plus a basal structure. The female karyotype comprises 16 monocentric chromosomes, predominantly with acrocentric morphology. Comparison with data of the ixodids and mesostigmatids indicates that a low number of chromosomes and predomination of acrocentric chromosomes could be plesiomorphies of the parasitiform mites.

Meiosis Progression and Recombination in Holocentric Plants: What Is Known?

Differently from the common monocentric organization of eukaryotic chromosomes, the so-called holocentric chromosomes present many centromeric regions along their length. This chromosomal organization can be found in animal and plant lineages, whose distribution suggests that it has evolved independently several times. Holocentric chromosomes present an advantage: even broken chromosome parts can be correctly segregated upon cell division. However, the evolution of holocentricity brought about consequences to nuclear processes and several adaptations are necessary to cope with this new organization. Centromeres of monocentric chromosomes are involved in a two-step cohesion release during meiosis. To deal with that holocentric lineages developed different adaptations, like the chromosome remodeling strategy in Caenorhabditis elegans or the inverted meiosis in plants. Furthermore, the frequency of recombination at or around centromeres is normally very low and the presence of centromeric regions throughout the entire length of the chromosomes could potentially pose a problem for recombination in holocentric organisms. However, meiotic recombination happens, with exceptions, in those lineages in spite of their holocentric organization suggesting that the role of centromere as recombination suppressor might be altered in these lineages. Most of the available information about adaptations to meiosis in holocentric organisms is derived from the animal model C. elegans. As holocentricity evolved independently in different lineages, adaptations observed in C. elegans probably do not apply to other lineages and very limited research is available for holocentric plants. Currently, we still lack a holocentric model for plants, but good candidates may be found among Cyperaceae, a large angiosperm family. Besides holocentricity, chiasmatic and achiasmatic inverted meiosis are found in the family. Here, we introduce the main concepts of meiotic constraints and adaptations with special focus in meiosis progression and recombination in holocentric plants. Finally, we present the main challenges and perspectives for future research in the field of chromosome biology and meiosis in holocentric plants.

Karyotype evolution and preliminary molecular assessment of genera in the family Scorpiopidae (Arachnida: Scorpiones)

The scorpions represent an ancient and morphologically conserved order of arachnids. Despite that, their karyotypes may differ considerably even among closely related species. In this study, we identify the trends of the karyotype evolution in the family Scorpiopidae based on integrating cytogenetic data and multi-locus molecular phylogenetic approaches. We detected considerable variability in diploid numbers of chromosomes (from 48 to 147), 18S rRNA gene cluster positions (from terminal to pericentromeric) at the interspecific level. Moreover, we identified independent fusions, fissions and inversions in the evolution of the family Scorpiopidae, leading to a remarkable diversification of the karyotypes. The dynamic system of the karyotype changes in this group is further documented by the presence of interstitial telomeric sequences (ITS) in two species. The cytogenetic differences observed among the analyzed species highlight the potential of this type of data for species-level taxonomy in scorpion lineages with monocentric chromosomes. Additionally, the results of our phylogenetic analyses support the monophyly of the family Scorpiopidae, but rendered several genera para- or polyphyletic.

CenH3 distribution reveals extended centromeres in the model beetle Tribolium castaneum.

Centromeres are chromosomal domains essential for kinetochore assembly and correct chromosome segregation. Inconsistent in their underlying DNA sequences, centromeres are defined epigenetically by the presence of the centromere-specific histone H3 variant CenH3. Most of the analyzed eukaryotes have monocentric chromosomes in which CenH3 proteins deposit into a single, primary constriction visible at metaphase chromosomes. Contrary to monocentrics, evolutionary sporadic holocentric chromosomes lack a primary constriction and have kinetochore activity distributed along the entire chromosome length. In this work, we identified cCENH3 protein, the centromeric H3 histone of the coleopteran model beetle Tribolium castaneum. By ChIP-seq analysis we disclosed that cCENH3 chromatin assembles upon a repertoire of repetitive DNAs. cCENH3 in situ mapping revealed unusually elongated T. castaneum centromeres that comprise approximately 40% of the chromosome length. Being the longest insect regional centromeres evidenced so far, T. castaneum centromeres are characterized by metapolycentric structure composed of several individual cCENH3-containing domains. We suggest that the model beetle T. castaneum with its metapolycentromeres could represent an excellent model for further studies of non-canonical centromeres in insects.

Chromosome number evolves at equal rates in holocentric and monocentric clades.

Despite the fundamental role of centromeres two different types are observed across plants and animals. Monocentric chromosomes possess a single region that function as the centromere while in holocentric chromosomes centromere activity is spread across the entire chromosome. Proper segregation may fail in species with monocentric chromosomes after a fusion or fission, which may lead to chromosomes with no centromere or multiple centromeres. In contrast, species with holocentric chromosomes should still be able to safely segregate chromosomes after fusion or fission. This along with the observation of high chromosome number in some holocentric clades has led to the hypothesis that holocentricity leads to higher rates of chromosome number evolution. To test for differences in rates of chromosome number evolution between these systems, we analyzed data from 4,393 species of insects in a phylogenetic framework. We found that insect orders exhibit striking differences in rates of fissions, fusions, and polyploidy. However, across all insects we found no evidence that holocentric clades have higher rates of fissions, fusions, or polyploidy than monocentric clades. Our results suggest that holocentricity alone does not lead to higher rates of chromosome number changes. Instead, we suggest that other co-evolving traits must explain striking differences between clades.

Analysis of Holhymenia histrio genome provides insight into the satDNA evolution in an insect with holocentric chromosomes.

Satellite DNAs (satDNA) are fast-evolving repetitive sequences organized in large tandem arrays, with characteristic enrichment in heterochromatin. Knowledge about evolutionary dynamics of this genome fraction is mostly restricted to its characterization in species with monocentric chromosomes, i.e., localized centromeres. In holocentric species, with non-localized centromeres, satDNAs have been largely ignored. Here we advance the understanding of satDNA evolution among holocentric species by characterization of the most abundant satDNAs in the hemipteran Holhymenia histrio, integrating genomic and chromosomal analyses. High plasticity at chromosomal and molecular levels was noticed for 34 satDNAs populating H. histrio genome. One satDNA family in particular (HhiSat01-184) was highly amplified on multiple chromosomes and also highly polymorphic. Our data support the emergence of a new satDNA family from this abundant satDNA, confined to a single chromosome. Moreover, we present new information about composition of a peculiar chromosome in Coreidae, the m-chromosome, and of the X chromosome. Overall, the molecular and chromosomal patterns for satDNAs in the holocentric species H. histrio seem to be similar to those observed in monocentric species.