Comparative meiosis and cytogenomic analysis in euploid and aneuploid hybrids of Urochloa P. Beauv Abstract The genus Urochloa includes most of the important grasses and hybrids currently used as pastures in the tropical regions. Cytogenetic analyzes have identified some aneuploid hybrids that provide new perspectives for genetic breeding. The objective was to analyze the meiotic behavior in euploid (2n = 4x = 36) and aneuploid (2n = 4x = 36 + 2) hybrids of U. ruziziensis x U. decumbens and U. ruziziensis x U. brizantha. Later, the chromosomes and respective genomes involved in pairing configurations and abnormalities were identified through GISH, with an emphasis on tracking the behavior of the additional chromosomes in the aneuploid hybrid U. ruziziensis x U. decumbens (B1B2B2B2 genomes). The aneuploid U. ruziziensis x U. decumbens shows a higher frequency of univalents, reduction of bivalents, and higher index of irregularities compared with the euploid hybrid. For the aneuploid U. ruziziensis x U. brizantha, there was a reduction in the frequency of univalents, an increase in bivalent and trivalent rates and a lower frequency of abnormalities when compared with the euploid hybrid. The rates of meiotic abnormalities and pairing configurations are parental genotype-dependent and influenced by trisomy. The chromosomes of the B1 and B2 genomes of the aneuploid hybrid (U. ruziziensis x U. decumbens) are involved in the formation of univalents, bivalents, and multivalents in inter-, intra- and inter–intragenomic pairings. In general, the segregation times of chromosomes of the genomes are different, since the chromosomes of the B1 genome segregate more slowly.

Correction to: Implications of sequence variation on the evolution of rRNA The article Implications of sequence variation on the evolution of rRNA, written by Matthew M. Parks, Chad M. Kurylo, Jake E. Batchelder, C. Theresa Vincent and Scott C. Blanchard, was originally published electronically on the publisher’s internet portal (currently SpringerLink).

Fluorescence in situ hybridization in plants: recent developments and future applications Abstract Fluorescence in situ hybridization (FISH) was developed more than 30 years ago and has been the most paradigm-changing technique in cytogenetic research. FISH has been used to answer questions related to structure, mutation, and evolution of not only individual chromosomes but also entire genomes. FISH has served as an important tool for chromosome identification in many plant species. This review intends to summarize and discuss key technical development and applications of FISH in plants since 2006. The most significant recent advance of FISH is the development and application of probes based on synthetic oligonucleotides (oligos). Oligos specific to a repetitive DNA sequence, to a specific chromosomal region, or to an entire chromosome can be computationally identified, synthesized in parallel, and fluorescently labeled. Oligo probes designed from conserved DNA sequences from one species can be used among genetically related species, allowing comparative cytogenetic mapping of these species. The advances with synthetic oligo probes will significantly expand the applications of FISH especially in non-model plant species. Recent achievements and future applications of FISH and oligo-FISH are discussed.

A centromere satellite concomitant with extensive karyotypic diversity across the Peromyscus genus defies predictions of molecular drive Abstract A common feature of eukaryotic centromeres is the presence of large tracts of tandemly arranged repeats, known as satellite DNA. However, these centromeric repeats appear to experience rapid evolution under forces such as molecular drive and centromere drive, seemingly without consequence to the integrity of the centromere. Moreover, blocks of heterochromatin within the karyotype, including the centromere, are hotspots for chromosome rearrangements that may drive speciation events by contributing to reproductive isolation. However, the relationship between the evolution of heterochromatic sequences and the karyotypic dynamics of these regions remains largely unknown. Here, we show that a single conserved satellite DNA sequence in the order Rodentia of the genus Peromyscus localizes to recurrent sites of chromosome rearrangements and heterochromatic amplifications. Peromyscine species display several unique features of chromosome evolution compared to other Rodentia, including stable maintenance of a strict chromosome number of 48 among all known species in the absence of any detectable interchromosomal rearrangements. Rather, the diverse karyotypes of Peromyscine species are due to intrachromosomal variation in blocks of repeated DNA content. Despite wide variation in the copy number and location of repeat blocks among different species, we find that a single satellite monomer maintains a conserved sequence and homogenized tandem repeat structure, defying predictions of molecular drive. The conservation of this satellite monomer results in common, abundant, and large blocks of chromatin that are homologous among chromosomes within one species and among diverged species. Thus, such a conserved repeat may have facilitated the retention of polymorphic chromosome variants within individuals and intrachromosomal rearrangements between species—both factors that have previously been hypothesized to contribute towards the extremely wide range of ecological adaptations that this genus exhibits.

Hybrids between Brassica napus and B. nigra show frequent pairing between the B and A/C genomes and resistance to blackleg Abstract High frequencies of homoeologous and even non-homologous chromosome recombination in Brassica hybrids can transfer useful traits between genomes, but also destabilise synthetic allopolyploids. We produced triploid hybrids (2n = 3x = ABC) from the cross B. napus (rapeseed, 2n = 4x = AACC) × B. nigra (black mustard, 2n = 2x = BB) by embryo rescue and allohexaploid hybrids (2n = 6x = AABBCC = 54) by chromosome doubling of the triploids. These hybrids demonstrated resistance to blackleg disease (causal agent: Leptosphaeria maculans) inherited from their B. nigra parent. In order to assess the possibility of transfer of this resistance between the B genome and the A and C subgenomes of B. napus, as well as to assess the genomic stability of allohexaploids from the cross B. napus × B. nigra, frequencies of non-homologous chromosome pairing in these hybrids were assessed using classical cytogenetics and genomic in-situ hybridization. Meiosis was highly irregular, and non-homologous chromosome pairing between the B genome and the A/C genomes was common in both triploid hybrids (observed in 38% of pollen mother cells) and allohexaploid hybrids (observed in 15% of pollen mother cells). Our results suggest that introgression of blackleg resistance from the B genome into the A or C genomes should be possible, but that allohexaploids from this genome combination are likely unstable.

MiR-153 regulates cardiomyocyte apoptosis by targeting Nrf2/HO-1 signaling Abstract MicroRNAs (miRNAs) play various roles in the regulation of human disease, including cardiovascular diseases. MiR-153 has been previously shown to be involved in regulating neuron survival during cerebral ischemia/reperfusion (I/R) injury. However, whether miR-153 is involved in I/R-induced cardiomyocyte apoptosis remains to be elucidated. In this study, we aimed to explore the role of miR-153 in the regulation of I/R-induced cardiomyocyte apoptosis and to investigate the miR-153-mediated molecular signaling pathway responsible for its effect on cardiomyocytes using an oxygen-glucose deprivation and reoxygenation (OGD/R) cellular model. We found that OGD/R treatment induced significant upregulation of miR-153 in cardiomyocytes causing reactive oxygen species (ROS) production and cell apoptosis signaling activation and subsequently leading to cardiomyocyte apoptosis. Suppression of miR-153 protected cardiomyocytes against OGD/R treatment. We further identified that nuclear factor-like 2 (Nrf2) is a functional target of miR-153. Nrf2/ heme oxygenase-1 (HO-1) signaling plays a critical role in miR-153 regulated OGD/R-induced cardiomyocyte apoptosis. Our study indicates that the inhibition of miR-153 or restoration of Nrf2 may serve as a potential therapeutic strategy for ischemia/reperfusion injury prevention.

Genome-wide DNA copy number analysis and targeted transcriptional analysis of canine histiocytic malignancies identifies diagnostic signatures and highlights disruption of spindle assembly complex Abstract Canine histiocytic malignancies (HM) are rare across the general dog population, but overrepresented in certain breeds, such as Bernese mountain dog and flat-coated retriever. Accurate diagnosis relies on immunohistochemical staining to rule out histologically similar cancers with different prognoses and treatment strategies (e.g., lymphoma and hemangiosarcoma). HM are generally treatment refractory with overall survival of less than 6 months. A lack of understanding regarding the mechanisms of disease development and progression hinders development of novel therapeutics. While the study of human tumors can benefit veterinary medicine, the rarity of the suggested orthologous disease (dendritic cell sarcoma) precludes this. This study aims to improve the understanding of underlying disease mechanisms using genome-wide DNA copy number and gene expression analysis of spontaneous HM across several dog breeds. Extensive DNA copy number disruption was evident, with losses of segments of chromosomes 16 and 31 detected in 93% and 72% of tumors, respectively. Droplet digital PCR (ddPCR) evaluation of these regions in numerous cancer specimens effectively discriminated HM from other common round cell tumors, including lymphoma and hemangiosarcoma, resulting in a novel, rapid diagnostic aid for veterinary medicine. Transcriptional analysis demonstrated disruption of the spindle assembly complex, which is linked to genomic instability and reduced therapeutic impact in humans. A key signature detected was up-regulation of Matrix Metalloproteinase 9 (MMP9), supported by an immunohistochemistry-based assessment of MMP9 protein levels. Since MMP9 has been linked with rapid metastasis and tumor aggression in humans, the data in this study offer a possible mechanism of aggression in HM.

Heterochromatic regions in Japanese quail chromosomes: comprehensive molecular-cytogenetic characterization and 3D mapping in interphase nucleus Abstract Chromosomes of Japanese quail (Coturnix coturnix japonica, 2n=78), a galliform domestic species closely related to chicken, possess multiple heterochromatic segments. Due to the difficulties in careful analysis of such heterochromatic regions, there is a lack of data on their DNA composition, epigenetic status, as well as spatial distribution in interphase nucleus. In the present study, we applied giant lampbrush chromosome (LBC) microdissection for high-resolution analysis of quail centromeric regions of macrochromosomes and polymorphic short arms of submetacentric microchromosomes. FISH with the dissected material on mitotic and meiotic chromosomes indicated that in contrast to centromeres of chicken macrochromosomes, which are known to harbor chromosome-specific and, in some cases, tandem repeat-free sequences, centromeres of quail macroautosomes (CCO1–CCO11) have canonical organization. CCO1–CCO11 centromeres possess massive blocks of common DNA repeats demonstrating transcriptional activity at LBC stage. These repeats seem to have been subjected to chromosome size-correlated homogenization previously described primarily for avian microchromosomes. In addition, comparative FISH on chicken chromosomes supported the previous data on centromere repositioning events during galliform karyotype evolution. In interphase nucleus of different cell types, repetitive elements specific for microchromosome short arms constitute the material of prominent centrally located chromocenters enriched with markers of constitutive heterochromatin and rimmed with clusters of microchromosomal centromeric BglII-repeat. Thus, clustering of such repeats is responsible for the peculiar architecture of quail interphase nucleus. In contrast, centromere repeats of the largest macrochromosomes (CCO1 and CCO2) are predominantly localized in perinuclear heterochromatin. The possible involvement of the isolated repeats in radial genome organization is discussed.

The importance of the nuclear positioning of the PPARG gene for its expression during porcine in vitro adipogenesis Abstract Proper expression of the PPARG gene, which encodes a key transcription factor of adipogenesis, is indispensable in the formation of mature adipocytes. The positioning of a gene within the nuclear space has been implicated in gene regulation. We here report on the significance of the PPARG gene’s nuclear positioning for its activity during in vitro adipogenesis in the pig. We used an established system of differentiation of mesenchymal stem cells derived from bone marrow and adipose tissue into adipocytes. The differentiation process was carried out for 7 days, and the cells were examined using the 3D DNA/immuno-FISH and RNA/DNA-FISH approaches. PPARG transcript level was measured using real-time PCR, and PPARγ activity was detected with colorimetric assay. Changes in the nuclear location of the PPARG gene were observed when we compared undifferentiated mesenchymal stem cells with mature adipocytes. The gene moved from the nuclear periphery to the nuclear center as its transcriptional activity increased. The RNA/DNA-FISH approach shows that differences in primary transcript production correlated with the allele’s nuclear positioning. Transcriptionally active alleles preferentially occupy the central part of the nucleus, while inactive alleles are found on the nuclear periphery. We also show that transcription of PPARG begins with one allele, but that both alleles are active in later stages of differentiation. Our results provide evidence that functionally distinct alleles of the PPARG gene are positioned in different parts of the cell nucleus. This confirms the importance of nuclear architecture to the regulation of PPARG gene transcription, and thus to the fate of the adipose cell.

The mobilome of Drosophila incompta , a flower-breeding species: comparison of transposable element landscapes among generalist and specialist flies Abstract The Drosophila genus is one of the main model organisms in evolutionary studies, including those investigating the role of transposable elements (TE) in genomic evolution both at the nucleotide and chromosome levels. D. incompta is a species with restricted ecology, using Cestrum (Solanaceae) flowers as unique sources for oviposition, feeding and development. In the present study, we deeply characterise the D. incompta mobilome and generate a curated dataset. A total of 277 elements were identified, corresponding to approximately 14% of the genome, and 164 of these elements are new, of which 32.62% are putatively autonomous and 8.9% are transcriptionally active in adult flies. The restricted ecology does not seem to influence the dynamics of TE in this fly, since the proportion and diversity of TEs in its genome are similar to that of other Drosophila species. This result is reinforced by the absence of a clear pattern when comparing the TE landscape between generalist and specialist flies. Using 32 available Drosophila genomes—24 ecologically generalist species and 8 specialist species—no difference was found between their TE landscape patterns. However, differences were found between species of the Sophophora and Drosophila subgenus, indicating there are lineage-specific factors shaping TE landscapes.