Metapolycentromeres consist of multiple sequential domains of centromeric chromatin associated with a centromere-specific variant of histone H3 (CENP-A), functioning collectively as a single centromere. To date, they have been revealed in nine flowering plant, five insect and six vertebrate species. In this paper, we focus on their structure and possible mechanisms of emergence and evolution. The metapolycentromeres may vary in the number of centromeric domains and in their genetic content and epigenetic modifications. However, these variations do not seem to affect their function. The emergence of metapolycentromeres has been attributed to multiple Robertsonian translocations and segmental duplications. Conditions of genomic instability, such as interspecific hybridization and malignant neoplasms, are suggested as triggers for the de novo emergence of metapolycentromeres. Addressing the “centromere paradox” – the rapid evolution of centromeric DNA and proteins despite their conserved cellular function – we explore the centromere drive hypothesis as a plausible explanation for the dynamic evolution of centromeres in general, and in particular the emergence of metapolycentromeres and holocentromeres. Apparently, metapolycentromeres are more common across different species than it was believed until recently. Indeed, a systematic review of the available cytogenetic publications allowed us to identify 27 candidate species with metapolycentromeres. Тhe list of the already established and newly revealed candidate species thus spans 27 species of flowering plants and eight species of gymnosperm plants, five species of insects, and seven species of vertebrates. This indicates an erratic phylogenetic distribution of the species with metapolycentromeres and may suggest an independent emergence of the metapolycentromeres in the course of evolution. However, the current catalog of species with identified and likely metapolycentromeres remains too short to draw reliable conclusions about their evolution, particularly in the absence of knowledge about related species without metapolycentromeres for comparative analysis. More studies are necessary to shed light on the mechanisms of metapolycentromere formation and evolution.

Meiotic recombination is an important source of genetic diversity. Using immunolocalization of several meiotic proteins at the spreads of male pachytene cells, we estimated the number of recombination nodules per cell and their distribution along the macrochromosome 1 of the Common linnet, Eurasian bullfinch, Eurasian siskin, and European goldfinch. The macrochromosomes of the two former species have metapolycentromeres, composed of several centromeric domains. We detected significant interspecies differences in the mean numbers of recombination nodules per genome: 52.9 ± 2.8 in the linnet, 49.5 ± 3.5 in the bullfinch, 61.5 ± 6.3 in the siskin and 52.2 ± 2.7 in the goldfinch. Recombination patterns on macrochromosome 1 were similar across species, with more nodules localized near chromosome ends and fewer around centromeres. The distance from the proximal nodule to the centromere depended on the nodule count per chromosome arm, with more events leading to a closer location. However, species with different centromere types showed no difference in this regard. We propose that the deficiency of recombination sites near centromeres could be due to the sequential occurrence of crossovers starting from the chromosome ends and may not be attributed to any suppressive effect of the centromere itself.

Germline-restricted chromosomes (GRCs) are present in the genomes of germline cells and absent from somatic cells. A GRC is found in all species of the songbirds (Passeri) and in none of the other bird orders studied to date. This indicates that GRC originated in the common ancestor of the songbirds. The germline-restricted chromosome is permanently absent from somatic cells of the songbird, while female germline cells usually contain two copies of GRC and male ones have one copy. In females, GRCs undergo synapsis and restricted recombination in their terminal regions during meiotic prophase. In males, it is almost always eliminated from spermatocytes. Thus, GRC is inherited almost exclusively through the maternal lineage. The germline-restricted chromosome is a necessary genomic element in the germline cells of songbirds. To date, the GRC genetic composition has been studied in four species only. Some GRC genes are actively expressed in female and male gonads, controlling the development of germline cells and synthesis of the proteins involved in the organization of meiotic chromosomes. Songbird species vary in GRC size and genetic composition. The GRC of each bird species consists of amplified and modified copies of genes from the basic genome of that species. The level of homology between GRCs of different species is relatively low, indicating a high rate of genetic evolution of this chromosome. Transmission through the maternal lineage and suppression of the recombination contribute significantly to the accelerated evolution of GRCs. One may suggest that the rapid coordinated evolution between the GRC genes and the genes of the basic genome in the songbirds might be responsible for the explosive speciation and adaptive radiation of this most species-rich and diverse infraorder of birds.

The classical hypothesis proposes that the lack of recombination on sex chromosomes arises due to selection for linkage between a sex-determining locus and sexually antagonistic loci, primarily facilitated by inversions. However, cessation of recombination on sex chromosomes could be attributed also to neutral processes, connected with other chromosome rearrangements or can reflect sex-specific recombination patterns existing already before sex chromosome differentiation. Three Coleonyx gecko species share a complex X1X1X2X2/X1X2Y system of sex chromosomes evolved via a fusion of the Y chromosome with an autosome. We analyzed synaptonemal complexes and sequenced flow-sorted sex chromosomes to investigate the effect of chromosomal rearrangement on recombination and differentiation of these sex chromosomes. The gecko sex chromosomes evolved from syntenic regions that were also co-opted also for sex chromosomes in other reptiles. We showed that in male geckos, recombination is less prevalent in the proximal regions of chromosomes and is even further drastically reduced around the centromere of the neo-Y chromosome. We highlight that pre-existing recombination patterns and Robertsonian fusions can be responsible for the cessation of recombination on sex chromosomes and that such processes can be largely neutral.

Negative heterosis can occur on different economically important traits, but the exact biological mechanisms of this phenomenon are still unknown. The present study focuses on determining the genetic factors associated with negative heterosis in interspecific hybrids between domestic sheep (Ovis aries) and argali (Ovis ammon). One locus (rs417431015) associated with viability and two loci (rs413302370, rs402808951) associated with meat productivity were identified. One gene (ARAP2) was prioritized for viability and three for meat productivity (PDE2A, ARAP1, and PCDH15). The loci associated with meat productivity were demonstrated to fit the overdominant inheritance model and could potentially be involved int negative heterosis mechanisms.

Closely related mammalian species often have differences in chromosome number and morphology, but there is still a debate about how these differences relate to reproductive isolation. To study the role of chromosome rearrangements in speciation, we used the gray voles in the Alexandromys genus as a model. These voles have a high level of chromosome polymorphism and substantial karyotypic divergence. We investigated testis histology and meiotic chromosome behavior in the captive-bred colonies of Alexandromys maximowiczii, Alexandromys mujanensis, two chromosome races of Alexandromys evoronensis, and their interracial and interspecies hybrids, to explore the relationship between karyotypic differences and male hybrid sterility. We found that the seminiferous tubules of the males of the parental species and the interracial hybrids, which were simple heterozygotes for one or more chromosome rearrangements, contained germ cells at all stages of spermatogenesis, indicating their potential fertility. Their meiotic cells displayed orderly chromosome synapsis and recombination. In contrast, all interspecies male hybrids, which were complex heterozygotes for a series of chromosome rearrangements, showed signs of complete sterility. Their spermatogenesis was mainly arrested at the zygotene- or pachytene-like stages due to the formation of complex multivalent chains, which caused extended chromosome asynapsis. The asynapsis led to the silencing of unsynapsed chromatin. We suggest that chromosome asynapsis is the main cause of meiotic arrest and male sterility in the interspecies hybrids of East Asian voles.

Karyotypes of less than 10% of bird species are known. Using immunolocalization of the synaptonemal complex, the core structure of meiotic chromosomes at the pachytene stage, and centromere proteins, we describe male pachytene karyotypes of 17 species of birds. This method enables higher resolution than the conventional analyses of metaphase chromosomes. We provide the first descriptions of the karyotypes of 3 species (rook, Blyth's reed warbler, and European pied flycatcher), correct the published data on the karyotypes of 10 species, and confirm them for 4 species. All passerine species examined have highly conservative karyotypes, 2n = 80-82 with 7 pairs of macrochromosomes (including the ZZ sex chromosome pair which was not unambiguously distinguished from other macrochromosomes in most species) and 33-34 pairs of microchromosomes. In all of them, but not in the common cuckoo, we revealed single copies of the germline-restricted chromosomes varying in size and morphology even between closely related species. This indicates a fast evolution of this additional chromosome. The interspecies differences concern the sizes of the macrochromosomes, morphology of the microchromosomes, and sizes of the centromeres. The pachytene cells of the gouldian finch, brambling, and common linnet contain heteromorphic synaptonemal complexes indicating heterozygosity for inversions or centromere shifts. The European pied flycatcher, gouldian finch, and domestic canary have extended centromeres in several macro- and microchromosomes.

In recent years, the number of genome-wide association studies (GWAS) carried out for various economically important animal traits has been increasing. GWAS discoveries provide summary statistics that can be used both for targeted marker-oriented selection and for studying the genetic control of economically important traits of farm animals. In contrast to research in human genetics, GWAS on farm animals often does not meet generally accepted standards (availability of information about effect and reference alleles, the size and direction of the effect, etc.). This greatly complicates the use of GWAS results for breeding needs. Within the framework of human genetics, there are several technological solutions for researching the harmonized results of GWAS, including one of the largest, the GWAS-MAP platform. For other types of living organisms, including economically important agricultural animals, there are no similar solutions. To our knowledge, no similar solution has been proposed to date for any of the species of economically important animals. As part of this work, we focused on creating a platform similar to GWAS-MAP for working with the results of GWAS of sheep, since sheep breeding is one of the most important branches of agriculture. By analogy with the GWAS-MAP platform for storing, unifying and analyzing human GWAS, we have created the GWAS-MAP|ovis platform. The platform currently contains information on more than 34 million associations between genomic sequence variants and traits of meat production in sheep. The platform can also be used to conduct colocalization analysis, a method that allows one to determine whether the association of a particular locus with two different traits is the result of pleiotropy or whether these traits are associated with different variants that are in linkage disequilibrium. This platform will be useful for breeders to select promising markers for breeding, as well as to obtain information for the introduction of genomic breeding and for scientists to replicate the results obtained.

The last decade was marked by a steep rise in avian studies at genomic and cellular levels. Cell lines are important tools for in vitro studies in cell biology and cytogenetics. We developed a simple method of primary somatic cell culture establishment from the ovaries of the great tits (Parus major) and testes of ten Passerine species, characterized the cellular composition of the ovary-derived lines using RT-PCR and immunolocalization of the tissue-specific markers and tested the efficiency of two methods of genetic transformation of the ovary-derived cell line. We found that the ovary-derived cell cultures of the great tit were composed of fibroblasts mainly, but also contained interstitial and granulosa cells. They were cultivated until the 10th passage without any noticeable decrease in their proliferative activity. The testis-derived cell cultures had lower proliferative potential. However, both ovary- and testis-derived cell cultures provided enough material for high quality mitotic metaphase chromosome preparations. The efficiency of its transduction with lentivirus containing a GFP reporter was very low, while electroporation with episomal vectors expressing GFP resulted in a high yield of GFP-positive cells. The proposed method could be used for the generation of high quality material for various cytogenetic and genomic studies.

Germline-restricted chromosomes (GRCs) are accessory chromosomes that occur only in germ cells. They are eliminated from somatic cells through programmed DNA elimination during embryo development. GRCs have been observed in several unrelated animal taxa and show peculiar modes of non-Mendelian inheritance and within-individual elimination. Recent cytogenetic and phylogenomic evidence suggests that a GRC is present across the species-rich songbirds, but absent in non-passerine birds, implying that over half of all 10,500 bird species have extensive germline/soma genome differences. Here, we review recent insights gained from genomic, transcriptomic, and cytogenetic approaches with regard to the genetic content, phylogenetic distribution, and inheritance of the songbird GRC. While many questions remain unsolved in terms of GRC inheritance, elimination, and function, we discuss plausible scenarios and future directions for understanding this widespread form of programmed DNA elimination.

Karyotypes of less than 10% of bird species are known. Using immunolocalization of the synaptonemal complex, the core structure of meiotic chromosomes at the pachytene stage, and centromere proteins we described male pachytene karyotypes of seventeen species of birds. This method enables higher resolution than the conventional analyses of metaphase chromosomes. We provided the first descriptions of the karyotypes of three species (Rook, Blyth’s reed warbler and European pied flycatcher), corrected the published data on the karyotypes of ten species and confirmed them for four species. All passerine species examined have highly conservative karyotypes, 2n=80-82 with seven pairs of macrochromosomes and 33-34 pairs of microchromosomes. In all of them but not in the Common cuckoo we revealed single copies of the germline restricted chromosomes varying in size and morphology even between closely related species. This indicates a fast evolution of this additional chromosome. The interspecies differences concern the sizes of the macrochromosomes, morphology of the microchromosomes and sizes of the centromeres. The pachytene cells of the Gouldian finch, Brambling and Common linnet contained heteromorphic synaptonemal complexes indicating heterozygosity for inversions or centromere shifts. The European pied flycatcher, Gouldian finch and Domestic canary have extended centromeres in several macro- and microchromosomes.

Heteromorphic W and Y sex chromosomes often experience gene loss and heterochromatinization, which is frequently viewed as their "degeneration". However, the evolutionary trajectories of the heterochromosomes are in fact more complex since they may not only lose but also acquire new sequences. Previously, we found that the heterochromatic W chromosome of a lizard Eremias velox (Lacertidae) is decondensed and thus transcriptionally active during the lampbrush stage. To determine possible sources of this transcription, we sequenced DNA from a microdissected W chromosome sample and a total female DNA sample and analyzed the results of reference-based and de novo assembly. We found a new repetitive sequence, consisting of fragments of an autosomal protein-coding gene ATF7IP2, several SINE elements, and sequences of unknown origin. This repetitive element is distributed across the whole length of the W chromosome, except the centromeric region. Since it retained only 3 out of 10 original ATF7IP2 exons, it remains unclear whether it is able to produce a protein product. Subsequent studies are required to test the presence of this element in other species of Lacertidae and possible functionality. Our results provide further evidence for the view of W and Y chromosomes as not just "degraded" copies of Z and X chromosomes but independent genomic segments in which novel genetic elements may arise.

Amplified sequences constitute a large part of mammalian genomes. A chromosome 1 containing 2 large (up to 50 Mb) homogeneously staining regions (HSRs) separated by a small inverted euchromatic region is present in many natural populations of the house mouse (Mus musculus musculus). The HSRs are composed of a long-range repeat cluster, Sp100-rs, with a repeat length of 100 kb. In order to understand the organization and function of HSRs in meiotic chromosomes, we examined synapsis and recombination in male mice hetero- and homozygous for the HSR-carrying chromosome using FISH with an HSR-specific DNA probe and immunolocalization of the key meiotic proteins. In all homozygous and heterozygous pachytene nuclei, we observed fully synapsed linear homomorphic bivalents 1 marked by the HSR FISH probe. The synaptic adjustment in the heterozygotes was bilateral: the HSR-carrying homolog was shortened and the wild-type homolog was elongated. The adjustment was reversible: desynapsis at diplotene was accompanied by elongation of the HSRs. Immunolocalization of H3K9me2/3 indicated that the HSRs in the meiotic chromosome retained the epigenetic modification typical for C-heterochromatin in somatic cells. MLH1 foci, marking mature recombination nodules, were detected in the proximal HSR band in heterozygotes and in both HSR bands of homozygotes. Unequal crossing over within the long-range repeat cluster can cause variation in size of the HSRs, which has been detected in the natural populations of the house mouse.

The formation of hybrid sterility is an important stage of speciation. The voles of the genus Microtus , which is the most speciose genus of rodents, provide a good model for studying the cytological mechanisms of hybrid sterility. The voles of the “ mystacinus ” group of the subgenus Microtus (2 n = 54) comprising several recently diverged forms with unclear taxonomic status are especially interesting. To resolve the taxonomic status of Microtus mystacinus and Microtus kermanensis , we crossed both with Microtus rossiaemeridionalis , and M. kermanensis alone with Microtus arvalis “obscurus” and M. transcaspicus and examined the reproductive performance of their F1 hybrids. All interspecies male hybrids were sterile. Female M. kermanensis × M. arvalis and M. kermanensis × M. transcaspicus hybrids were sterile as well. Therefore, M. mystacinus , M. kermanensis , and M. rossiaemeridionalis could be considered valid species. To gain an insight into the cytological mechanisms of male hybrid sterility, we carried out a histological analysis of spermatogenesis and a cytological analysis of chromosome synapsis, recombination, and epigenetic chromatin modifications in the germ cells of the hybrids using immunolocalization of key meiotic proteins. The hybrids showed wide variation in the onset of spermatogenesis arrest stage, from mature (although abnormal) spermatozoa to spermatogonia only. Chromosome asynapsis was apparently the main cause of meiotic arrest. The degree of asynapsis varied widely across cells, individuals, and the crosses—from partial asynapsis of several small bivalents to complete asynapsis of all chromosomes. The asynapsis was accompanied by a delayed repair of DNA double-strand breaks marked by RAD51 antibodies and silencing of unpaired chromatin marked by γH2A.X antibodies. Overall, the severity of disturbances in spermatogenesis in general and in chromosome synapsis in particular increased in the hybrids with an increase in the phylogenetic distance between their parental species.

All songbirds studied so far have a germline-restricted chromosome (GRC), which is present in the germ cells and absent in the somatic cells. It shows a wide variation in size, morphology, and genetic content between the songbird species. In this paper, we analyzed GRC behavior in female and male meiosis of the great tit, using immunolocalization of meiotic proteins and FISH with GRC-derived DNA probes. We found that, despite dozens of million years of independent evolution, the great tit GRC displays a striking similarity with the GRCs of two species of martins and two species of estrildid finches examined earlier. It was usually present in two copies in females forming recombining bivalent and in one copy in males forming a condensed heterochromatic body with dotted-like axial elements of the synaptonemal complex. We observed mosaicism for the GRC copy number in the female and male great tit. This indicates that one of the GRC copies might be passively lost during premeiotic germ cell divisions. After the meiotic prophase, the GRC was ejected from most male germ cells. The reverse and interspecies FISH with GRC-specific microdissected DNA probes indicates that GRCs of the great tit, pale martin, and zebra finch differ substantially in their genetic content despite similarities in the meiotic behavior.

Benefits and costs of meiotic recombination are a matter of discussion. Because recombination breaks allele combinations already tested by natural selection and generates new ones of unpredictable fitness, a high recombination rate is generally beneficial for the populations living in a fluctuating or a rapidly changing environment and costly in a stable environment. Besides genetic benefits and costs, there are cytological effects of recombination, both positive and negative. Recombination is necessary for chromosome synapsis and segregation. However, it involves a massive generation of double-strand DNA breaks, erroneous repair of which may lead to germ cell death or various mutations and chromosome rearrangements. Thus, the benefits of recombination (generation of new allele combinations) would prevail over its costs (occurrence of deleterious mutations) as long as the population remains sufficiently heterogeneous. Using immunolocalization of MLH1, a mismatch repair protein, at the synaptonemal complexes, we examined the number and distribution of recombination nodules in spermatocytes of two chicken breeds with high (Pervomai) and low (Russian Crested) recombination rates and their F1 hybrids and backcrosses. We detected negative heterosis for recombination rate in the F1 hybrids. Backcrosses to the Pervomai breed were rather homogenous and showed an intermediate recombination rate. The differences in overall recombination rate between the breeds, hybrids and backcrosses were mainly determined by the differences in the crossing over number in the seven largest macrochromosomes. The decrease in recombination rate in F1 is probably determined by difficulties in homology matching between the DNA sequences of genetically divergent breeds. The suppression of recombination in the hybrids may impede gene flow between parapatric populations and therefore accelerate their genetic divergence.

Despite their economic value, sheep remain relatively poorly studied animals in terms of the number of known loci and genes associated with commercially important traits. This gap in our knowledge can be filled in by performing new genome-wide association studies (GWAS) or by re-analyzing previously documented data using novel powerful statistical methods. This study is focused on the search for new loci associated with meat productivity and carcass traits in sheep. With a multivariate approach applied to publicly available GWAS results, we identified eight novel loci associated with the meat productivity and carcass traits in sheep. Using an in silico follow-up approach, we prioritized 13 genes in these loci. One of eight novel loci near the FAM3C and WNT16 genes has been replicated in an independent sample of Russian sheep populations (N = 108). The novel loci were added to our regularly updated database increasing the number of known loci to more than 140.

Whole-chromosome fusions play a major role in the karyotypic evolution of reptiles. It has been suggested that certain chromosomes tend to fuse with sex chromosomes more frequently than others. However, the comparative genomic synteny data are too scarce to draw strong conclusions. We obtained and sequenced chromosome-specific DNA pools of Sceloporus malachiticus, an iguanian species which has experienced many chromosome fusions. We found that four of seven lineage-specific fusions involved sex chromosomes, and that certain syntenic blocks which constitute the sex chromosomes, such as the homologues of the Anolis carolinensis chromosomes 11 and 16, are repeatedly involved in sex chromosome formation in different squamate species. To test the hypothesis that the karyotypic shift could be associated with changes in recombination patterns, we performed a synaptonemal complex analysis in this species and in Sceloporus variabilis (2n = 34). It revealed that the sex chromosomes in S. malachiticus had two distal pseudoautosomal regions and a medial differentiated region. We found that multiple fusions little affected the recombination rate in S. malachiticus. Our data confirm more frequent involvement of certain chromosomes in sex chromosome formation, but do not reveal a connection between the gonosome-autosome fusions and the evolution of recombination rate. This article is part of the theme issue 'Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part II)'.

We analyzed the synapsis and recombination between Z and W chromosomes in the oocytes of nine neognath species: domestic chicken Gallus gallus domesticus, grey goose Anser anser, black tern Chlidonias niger, common tern Sterna hirundo, pale martin Riparia diluta, barn swallow Hirundo rustica, European pied flycatcher Ficedula hypoleuca, great tit Parus major and white wagtail Motacilla alba using immunolocalization of SYCP3, the main protein of the lateral elements of the synaptonemal complex, and MLH1, the mismatch repair protein marking mature recombination nodules. In all species examined, homologous synapsis occurs in a short region of variable size at the ends of Z and W chromosomes, where a single recombination nodule is located. The remaining parts of the sex chromosomes undergo synaptic adjustment and synapse non-homologously. In 25% of ZW bivalents of white wagtail, synapsis and recombination also occur at the secondary pairing region, which probably resulted from autosome−sex chromosome translocation. Using FISH with a paint probe specific to the germline-restricted chromosome (GRC) of the pale martin on the oocytes of the pale martin, barn swallow and great tit, we showed that both maternally inherited songbird chromosomes (GRC and W) share common sequences.