Distal Region Of Chromosome Research Articles

Structural effects of inosine substitution in telomeric DNA quadruplex.

The telomeric DNA, a distal region of eukaryotic chromosome containing guanine-rich repetitive sequence of (TTAGGG)n, has been shown to adopt higher-order structures, specifically G-quadruplexes (G4s). Previous studies have demonstrated the implication of G4 in tumor inhibition through chromosome maintenance and manipulation of oncogene expression featuring their G-rich promoter regions. Besides higher order structures, several regulatory roles are attributed to DNA epigenetic markers. In this work, we investigated how the structural dynamics of a G-quadruplex, formed by the telomeric sequence, is affected by inosine, a prevalent modified nucleotide. We used the standard (TTAGGG)n telomere repeats with guanosine mutated to inosine at each G position. Sequences (GGG)4, (IGG)4, (GIG)4, (GGI)4, (IGI)4, (IIG)4, (GII)4, and (III)4, bridged by TTA linker, are studied using biophysical experiments and molecular modeling. The effects of metal cations in quadruplex folding were explored in both Na+ and K+ containing buffers using CD and UV-melting studies. Our results show that antiparallel quadruplex topology forms with the native sequence (GGG)4 and the terminal modified DNAs (IGG)4 and (GGI)4 in both Na+ and K+ containing buffers. Specifically, quadruplex hybrid was observed for (GGG)4 in K+ buffer. Among the other modified sequences, (GIG)4, (IGI)4 and (GII)4 show parallel features, while (IIG)4 and (III)4 show no detectable conformation in the presence of either Na+ or K+. Our studies indicate that terminal lesions (IGG)4 and (GGI)4 may induce certain unknown conformations. The folding dynamics become undetectable in the presence of more than one inosine substitution except (IGI)4 in both buffer ions. In addition, both UV melting and CD melting studies implied that in most cases the K+ cation confers more thermodynamic stability compared to Na+. Collectively, our conformational studies revealed the diverse structural polymorphisms of G4 with position dependent G-to-I mutations in different ion conditions.

A single dominant locus restricts retrovirus replication in YBR/Ei mice.

Differential responses to viral infections are influenced by the genetic makeup of the host. Studies of resistance to retroviruses in human populations are complicated due to the inability to conduct proof-of-principle studies. Inbred mouse lines, which have a range of susceptible phenotypes to retroviruses, are an ideal tool to identify and characterize mechanisms of resistance and define their genetic underpinnings. YBR/Ei mice become infected with Mouse Mammary Tumor Virus, a mucosally transmitted murine retrovirus, but eliminate the virus from their pedigrees. Virus elimination correlates with a lack of virus-specific neonatal oral tolerance, which is a major mechanism for blocking the anti-virus response in susceptible mice. Virus control is unrelated to virus-neutralizing antibodies, cytotoxic CD8+ T cells, NK cells, and NK T cells, which are the best characterized mechanisms of resistance to retroviruses. We identified a single, dominant locus that controls the resistance mechanism, which we provisionally named attenuation of virus titers (Avt) and mapped to the distal region of chromosome 18. IMPORTANCE Elucidation of the mechanism that mediates resistance to retroviruses is of fundamental importance to human health, as it will ultimately lead to knowledge of the genetic differences among individuals in susceptibility to microbial infections.

Divergent patterns of meiotic double strand breaks and synapsis initiation dynamics suggest an evolutionary shift in the meiosis program between American and Australian marsupials.

In eutherian mammals, hundreds of programmed DNA double-strand breaks (DSBs) are generated at the onset of meiosis. The DNA damage response is then triggered. Although the dynamics of this response is well studied in eutherian mammals, recent findings have revealed different patterns of DNA damage signaling and repair in marsupial mammals. To better characterize these differences, here we analyzed synapsis and the chromosomal distribution of meiotic DSBs markers in three different marsupial species (Thylamys elegans, Dromiciops gliorides, and Macropus eugenii) that represent South American and Australian Orders. Our results revealed inter-specific differences in the chromosomal distribution of DNA damage and repair proteins, which were associated with differing synapsis patterns. In the American species T. elegans and D. gliroides, chromosomal ends were conspicuously polarized in a bouquet configuration and synapsis progressed exclusively from the telomeres towards interstitial regions. This was accompanied by sparse H2AX phosphorylation, mainly accumulating at chromosomal ends. Accordingly, RAD51 and RPA were mainly localized at chromosomal ends throughout prophase I in both American marsupials, likely resulting in reduced recombination rates at interstitial positions. In sharp contrast, synapsis initiated at both interstitial and distal chromosomal regions in the Australian representative M. eugenii, the bouquet polarization was incomplete and ephemeral, γH2AX had a broad nuclear distribution, and RAD51 and RPA foci displayed an even chromosomal distribution. Given the basal evolutionary position of T. elegans, it is likely that the meiotic features reported in this species represent an ancestral pattern in marsupials and that a shift in the meiotic program occurred after the split of D. gliroides and the Australian marsupial clade. Our results open intriguing questions about the regulation and homeostasis of meiotic DSBs in marsupials. The low recombination rates observed at the interstitial chromosomal regions in American marsupials can result in the formation of large linkage groups, thus having an impact in the evolution of their genomes.

The Arabidopsis Hop1 homolog ASY1 mediates cross-over assurance and interference.

The chromosome axis plays a crucial role in meiotic recombination. Here, we study the function of ASY1, the Arabidopsis homolog of the yeast chromosome axis-associated component Hop1. Specifically, we characterized cross-over (CO) distribution in female and male meiosis by deep sequencing of the progeny of an allelic series of asy1 mutants. Combining data from nearly 1,000 individual plants, we find that reduced ASY1 functionality leads to genomic instability and sometimes drastic genomic rearrangements. We further observed that COs are less frequent and appear in more distal chromosomal regions in plants with no or reduced ASY1 functionality, consistent with previous analyses. However, our sequencing approach revealed that the reduction in CO number is not as dramatic as suggested by cytological analyses. Analysis of double mutants of asy1 with mutants with three other CO factors, MUS81, MSH4, and MSH5, as well as the determination of foci number of the CO regulator MLH1 demonstrates that the majority of the COs in asy1, similar to the situation in the wildtype (WT), largely belong to the class I, which are subject to interference. However, these COs are redistributed in asy1 mutants and typically appear much closer than in the WT. Hence, ASY1 plays a key role in CO interference that spaces COs along a chromosome. Conversely, since a large proportion of chromosomes do not receive any CO, we conclude that CO assurance, the process that ensures the obligatory assignment of one CO per chromosome, is also affected in asy1 mutants.

Epigenomic and structural events preclude recombination in Brassica napus.

Meiotic recombination is a major evolutionary process generating genetic diversity at each generation in sexual organisms. However, this process is highly regulated, with the majority of crossovers lying in the distal chromosomal regions that harbor low DNA methylation levels. Even in these regions, some islands without recombination remain, for which we investigated the underlying causes. Genetic maps were established in two Brassica napus hybrids to detect the presence of such large nonrecombinant islands. The role played by DNA methylation and structural variations in this local absence of recombination was determined by performing bisulfite sequencing and whole genome comparisons. Inferred structural variations were validated using either optical mapping or oligo fluorescence in situ hybridization. Hypermethylated or inverted regions between Brassica genomes were associated with the absence of recombination. Pairwise comparisons of nine B. napus genome assemblies revealed that such inversions occur frequently and may contain key agronomic genes such as resistance to biotic stresses. We conclude that such islands without recombination can have different origins, such as DNA methylation or structural variations in B. napus. It is thus essential to take into account these features in breeding programs as they may hamper the efficient combination of favorable alleles in elite varieties.

Downregulation of Barley Regulator of Telomere Elongation Helicase 1 Alters the Distribution of Meiotic Crossovers.

Programmed meiotic DNA double-strand breaks (DSBs), necessary for proper chromosomal segregation and viable gamete formation, are repaired by homologous recombination (HR) as crossovers (COs) or non-crossovers (NCOs). The mechanisms regulating the number and distribution of COs are still poorly understood. The regulator of telomere elongation helicase 1 (RTEL1) DNA helicase was previously shown to enforce the number of meiotic COs in Caenorhabditis elegans but its function in plants has been studied only in the vegetative phase. Here, we characterised barley RTEL1 gene structure and expression using RNA-seq data previously obtained from vegetative and reproductive organs and tissues. Using RNAi, we downregulated RTEL1 expression specifically in reproductive tissues and analysed its impact on recombination using a barley 50k iSelect SNP Array. Unlike in C. elegans, in a population segregating for RTEL1 downregulated by RNAi, high resolution genome-wide genetic analysis revealed a significant increase of COs at distal chromosomal regions of barley without a change in their total number. Our data reveal the important role of RTEL1 helicase in plant meiosis and control of recombination.

Aegilops tauschii genome assembly Aet v5.0 features greater sequence contiguity and improved annotation.

Aegilops tauschii is the donor of the D subgenome of hexaploid wheat and an important genetic resource. The reference-quality genome sequence Aet v4.0 for Ae. tauschii acc. AL8/78 was therefore an important milestone for wheat biology and breeding. Further advances in sequencing acc. AL8/78 and release of the Aet v5.0 sequence assembly are reported here. Two new optical maps were constructed and used in the revision of pseudomolecules. Gaps were closed with Pacific Biosciences long-read contigs, decreasing the gap number by 38,899. Transposable elements and protein-coding genes were reannotated. The number of annotated high-confidence genes was reduced from 39,635 in Aet v4.0 to 32,885 in Aet v5.0. A total of 2245 biologically important genes, including those affecting plant phenology, grain quality, and tolerance of abiotic stresses in wheat, was manually annotated and disease-resistance genes were annotated by a dedicated pipeline. Disease-resistance genes encoding nucleotide-binding site domains, receptor-like protein kinases, and receptor-like proteins were preferentially located in distal chromosome regions, whereas those encoding transmembrane coiled-coil proteins were dispersed more evenly along the chromosomes. Discovery, annotation, and expression analyses of microRNA (miRNA) precursors, mature miRNAs, and phasiRNAs are reported, including miRNA target genes. Other small RNAs, such as hc-siRNAs and tRFs, were characterized. These advances enhance the utility of the Ae. tauschii genome sequence for wheat genetics, biotechnology, and breeding.

The Landscape of the Genomic Distribution and the Expression of the F-Box Genes Unveil Genome Plasticity in Hexaploid Wheat during Grain Development and in Response to Heat and Drought Stress

FBX proteins are subunits of the SCF complex (Skp1–cullin–FBX) belonging to the E3 ligase family, which is involved in the ubiquitin–proteasome 26S (UPS) pathway responsible for the post-translational protein turnover. By targeting, in a selective manner, key regulatory proteins for ubiquitination and 26S proteasome degradation, FBX proteins play a major role in plant responses to diverse developmental and stress conditions. Although studies on the genomic organization of the FBX gene family in various species have been reported, knowledge related to bread wheat (Triticum aestivum) is scarce and needs to be broadened. Using the latest assembly of the wheat genome, we identified 3670 TaFBX genes distributed non-homogeneously within the three subgenomes (A, B and D) and between the 21 chromosomes, establishing it as one of the richest gene families among plant species. Based on the presence of the five different chromosomal regions previously identified, the present study focused on the genomic distribution of the TaFBX family and the identification of differentially expressed genes during the embryogenesis stages and in response to heat and drought stress. Most of the time, when comparing the expected number of genes (taking into account the formal gene distribution on the entire wheat genome), the TaFBX family harbors a different pattern at the various stratum of observation (subgenome, chromosome, chromosomal regions). We report here that the local gene expansion of the TaFBX family must be the consequence of multiple and complex events, including tandem and small-scale duplications. Regarding the differentially expressed TaFBX genes, while the majority of the genes are localized in the distal chromosomal regions (R1 and R3), differentially expressed genes are more present in the interstitial regions (R2a and R2b) than expected, which could be an indication of the preservation of major genes in those specific chromosomal regions.

Distal Bias of Meiotic Crossovers in Hexaploid Bread Wheat Reflects Spatio-Temporal Asymmetry of the Meiotic Program.

Meiotic recombination generates genetic variation and provides physical links between homologous chromosomes (crossovers) essential for accurate segregation. In cereals the distribution of crossovers, cytologically evident as chiasmata, is biased toward the distal regions of chromosomes. This creates a bottleneck for plant breeders in the development of varieties with improved agronomic traits, as genes situated in the interstitial and centromere proximal regions of chromosomes rarely recombine. Recent advances in wheat genomics and genome engineering combined with well-developed wheat cytogenetics offer new opportunities to manipulate recombination and unlock genetic variation. As a basis for these investigations we have carried out a detailed analysis of meiotic progression in hexaploid wheat (Triticum aestivum) using immunolocalization of chromosome axis, synaptonemal complex and recombination proteins. 5-Bromo-2′-deoxyuridine (BrdU) labeling was used to determine the chronology of key events in relation to DNA replication. Axis morphogenesis, synapsis and recombination initiation were found to be spatio-temporally coordinated, beginning in the gene-dense distal chromosomal regions and later occurring in the interstitial/proximal regions. Moreover, meiotic progression in the distal regions was coordinated with the conserved chromatin cycles that are a feature of meiosis. This mirroring of the chiasma bias was also evident in the distribution of the gene-associated histone marks, H3K4me3 and H3K27me3; the repeat-associated mark, H3K27me1; and H3K9me3. We believe that this study provides a cytogenetic framework for functional studies and ongoing initiatives to manipulate recombination in the wheat genome.

Meiotic chromosome axis remodelling is critical for meiotic recombination in Brassica rapa.

Meiosis generates genetic variation through homologous recombination (HR) that is harnessed during breeding. HR occurs in the context of meiotic chromosome axes and the synaptonemal complex. To study the role of axis remodelling in crossover (CO) formation in a crop species, we characterized mutants of the axis-associated protein ASY1 and the axis-remodelling protein PCH2 in Brassica rapa. asy1 plants form meiotic chromosome axes that fail to synapse. CO formation is almost abolished, and residual chiasmata are proportionally enriched in terminal chromosome regions, particularly in the nucleolar organizing region (NOR)-carrying chromosome arm. pch2 plants show impaired ASY1 loading and remodelling, consequently achieving only partial synapsis, which leads to reduced CO formation and loss of the obligatory CO. PCH2-independent chiasmata are proportionally enriched towards distal chromosome regions. Similarly, in Arabidopsis pch2, COs are increased towards telomeric regions at the expense of (peri-) centromeric COs compared with the wild type. Taken together, in B. rapa, axis formation and remodelling are critical for meiotic fidelity including synapsis and CO formation, and in asy1 and pch2 CO distributions are altered. While asy1 plants are sterile, pch2 plants are semi-sterile and thus PCH2 could be an interesting target for breeding programmes.

Genetic structure of a germplasm for hybrid breeding in rye (Secale cereale L.).

Rye (Secale cereale L.) responds strongly to changes in heterozygosity with hybrids portraying strong heterosis effect on all developmental and yielding characteristics. In order to achieve the highest potential heterosis effect parental lines must originate from genetically distinct gene pools. Here we report the first comprehensive SNP-based population study of an elite germplasm using fertilization control system for hybrid breeding in rye that is genetically different to the predominating P-type. In total 376 inbred lines from Nordic Seed Germany GmbH were genotyped for 4419 polymorphic SNPs. The aim of this study was to confirm and quantify the genetic separation of parental populations, unveil their genetic characteristics and investigate underlying population structures. Through a palette of complimenting analysis, we confirmed a strong genetic differentiation (FST = 0.332) of parental populations validating the germplasms suitability for hybrid breeding. These were, furthermore, found to diverge considerably in several features with the maternal population portraying a strong population structure characterized by a narrow genetic profile, small effective population size and high genome-wise linkage disequilibrium. We propose that the employed male-sterility system putatively constitutes a population determining parameter by influencing the rate of introducing novel genetic variation to the parental populations. Functional analysis of linkage blocks led to identification of a conserved segment on the distal 4RL chromosomal region annotated to the Rfp3 male-fertility restoration 'Pampa' type gene. Findings of our study emphasized the immediate value of comprehensive population studies on elite breeding germplasms as a pre-requisite for application of genomic-based breeding techniques, introgression of novel material and to support breeder decision-making.

PacBio genome sequencing reveals new insights into the genomic organisation of the multi-copy ToxB gene of the wheat fungal pathogen Pyrenophora tritici-repentis

BackgroundNecrotrophic effector proteins secreted by fungal pathogens are important virulence factors that mediate the development of disease in wheat. Pyrenophora tritici-repentis (Ptr), the causal agent of wheat tan spot, has a race structure dependent on the combination of effectors. In Ptr, ToxA and ToxB are known proteinaceous effectors responsible for necrosis and chlorosis respectively. While Ptr ToxA is encoded by the single gene ToxA, ToxB has multiple loci in the Ptr genome, which is postulated to be directly related to the level of ToxB production and leaf chlorosis. Although previous analysis has indicated that the majority of the ToxB loci lie on a single chromosome, the exact number and chromosomal locations for all the ToxB loci have not been fully identified.ResultsIn this study, we have sequenced the genome of a race 5 ToxB-producing isolate (DW5), using PacBio long read technology, and found that ToxB duplications are nested in the complex subtelomeric chromosomal regions. A total of ten identical ToxB gene copies were identified and based on flanking sequence identity, nine loci appeared associated with chromosome 11 and a single copy with chromosome 5. Chromosome 11 multiple ToxB gene loci were separated by large sequence regions between 31 and 66 kb within larger segmental duplications in an alternating pattern related to loci strand, and flanked by transposable elements.ConclusionThis work provides for the first time the full accompaniment of ToxB loci and surrounding regions, and identifies the organization and distribution of ten ToxB loci to subtelomeric regions. To our knowledge, this is the first report of an interwoven strand-related duplication pattern event. This study further highlights the importance of resolving the highly complex distal chromosomal regions, that remain difficult to assemble, and can harbour important effectors and virulence factors.

Partial trisomy 4q and monosomy 5p inherited from a maternal translocationt(4;5)(q33; p15) in three adverse pregnancies

BackgroundCarriers of balanced reciprocal chromosomal translocations are at known reproductive risk for offspring with unbalanced genotypes and resultantly abnormal phenotypes. Once fertilization of a balanced translocation gamete with a normal gamete, the partial monosomy or partial trisomy embryo will undergo abortion, fetal arrest or fetal malformations. We reported a woman with chromosomal balanced translocation who had two adverse pregnancies. Prenatal diagnosis was made for her third pregnancy to provide genetic counseling and guide her fertility.Case presentationWe presented a woman with chromosomal balanced translocation who had three adverse pregnancies. Routine G banding and CNV-seq were used to analyze the chromosome karyotypes and copy number variants of amniotic fluid cells and peripheral blood. The karyotype of the woman was 46,XX,t(4;5)(q33;p15). During her first pregnancy, odinopoeia was performed due to fetal edema and abdominal fluid. The umbilical cord tissue of the fetus was examined by CNV-seq. The results showed a genomic gain of 24.18 Mb at 4q32.3-q35.2 and a genomic deletion of 10.84 Mb at 5p15.2-p15.33 and 2.36 Mb at 15q11.1-q11.2. During her second pregnancy, she did not receive a prenatal diagnosis because a routine prenatal ultrasound examination found no abnormalities. In 2016, she gave birth to a boy. The karyotype the of the boy was 46,XY,der(5)t(4;5)(q33;p15)mat. The results of CNV-seq showed a deletion of short arm of chromosome 5 capturing regions 5p15.2-p15.33, a copy gain of the distal region of chromosome 4 at segment 4q32.3q35.2, a duplication of chromosome 1 at segment 1q41q42.11 and a duplication of chromosome 17 at segment 17p12. During her third pregnancy, she underwent amniocentesis at 17 weeks of gestation. Chromosome karyotype hinted 46,XY,der(5)t(4;5)(q33;p15)mat. Results of CNV-seq showed a deletion of short arm (p) of chromosome 5 at the segment 5p15.2p15.33 and a duplication of the distal region of chromosome 4 at segment 4q32.3q35.2.ConclusionsChromosomal abnormalities in three pregnancies were inherited from the mother. Preimplantation genetic diagnosis is recommended to prevent the birth of children with chromosomal abnormalities.

ATR-16 syndrome: mechanisms linking monosomy to phenotype

BackgroundDeletions removing 100s–1000s kb of DNA, and variable numbers of poorly characterised genes, are often found in patients with a wide range of developmental abnormalities. In such cases, understanding the...

Two large reciprocal translocations characterized in the disease resistance-rich burmannica genetic group of Musa acuminata

Background and AimsBanana cultivars are derived from hybridizations involving Musa acuminata subspecies. The latter diverged following geographical isolation in distinct South-east Asian continental regions and islands. Observation of chromosome pairing irregularities in meiosis of hybrids between these subspecies suggested the presence of large chromosomal structural variations. The aim of this study was to characterize such rearrangements.MethodsMarker (single nucleotide polymorphism) segregation in a self-progeny of the ‘Calcutta 4’ accession and mate-pair sequencing were used to search for chromosomal rearrangements in comparison with the M. acuminata ssp. malaccensis genome reference sequence. Signature segment junctions of the revealed chromosome structures were identified and searched in whole-genome sequencing data from 123 wild and cultivated Musa accessions.Key ResultsTwo large reciprocal translocations were characterized in the seedy banana M. acuminata ssp. burmannicoides ‘Calcutta 4’ accession. One consisted of an exchange of a 240 kb distal region of chromosome 2 with a 7.2 Mb distal region of chromosome 8. The other involved an exchange of a 20.8 Mb distal region of chromosome 1 with a 11.6 Mb distal region of chromosome 9. Both translocations were found only in wild accessions belonging to the burmannicoides/burmannica/siamea subspecies. Only two of the 87 cultivars analysed displayed the 2/8 translocation, while none displayed the 1/9 translocation.ConclusionTwo large reciprocal translocations were identified that probably originated in the burmannica genetic group. Accurate characterization of these translocations should enhance the use of this disease resistance-rich burmannica group in breeding programmes.

The Role of Telomerase and Telomeres in Interstitial Lung Diseases: From Molecules to Clinical Implications.

Telomeres are distal chromosome regions associated with specific protein complexes that protect the chromosome against degradation and aberrations. Telomere maintenance capacity is an essential indication of healthy cell populations, and telomere damage is observed in processes such as malignant transformation, apoptosis, or cell senescence. At a cellular level, telomere damage may result from genotoxic stress, decreased activity of telomerase enzyme complex, dysfunction of shelterin proteins, or changes in expression of telomere-associated RNA such as TERRA. Clinical evidence suggests that mutation of telomerase genes (Tert/Terc) are associated with increased risk of congenital as well as age-related diseases (e.g., pneumonitis, idiopathic pulmonary fibrosis (IPF), dyskeratosis congenita, emphysema, nonspecific interstitial pneumonia, etc.). Thus, telomere length and maintenance can serve as an important prognostic factor as well as a potential target for new strategies of treatment for interstitial lung diseases (ILDs) and associated pulmonary pathologies.

Population genomics reveals a fine-scale recombination landscape for genetic improvement of cotton.

Recombination breaks up ancestral linkage disequilibrium, creates combinations of alleles, affects the efficiency of natural selection, and plays a major role in crop domestication and improvement. However, there is little knowledge regarding the variation in the population-scaled recombination rate in cotton. We constructed recombination maps and characterized the difference in the genomic landscape of the population-scaled recombination rate between Gossypium hirsutum and G.arboreum and sub-genomes based on the 381 sequenced G.hirsutum and 215G.arboreum accessions. Comparative genomics identified large structural variations and syntenic genes in the recombination regions, suggesting that recombination was related to structural variation and occurred preferentially in the distal chromosomal regions. Correlation analysis indicated that recombination was only slightly affected by geographical distribution and breeding period. A genome-wide association study (GWAS) was performed with 15 agronomic traits using 267 cotton accessions and identified 163 quantitative trait loci (QTL) and an important candidate gene (Ghir_COL2) for early maturity traits. Comparative analysis of recombination and a GWAS revealed that the QTL of fibre quality traits tended to be more common in high-recombination regions than were those of yield and early maturity traits. These results provide insights into the population-scaled recombination landscape, suggesting that recombination contributed to the domestication and improvement of cotton, which provides a useful reference for studying recombination in other species.

Global Characterization of X Chromosome Inactivation in Human Pluripotent Stem Cells

Dosage compensation of sex-chromosome gene expression between male and female mammals is achieved via X chromosome inactivation (XCI) by employing epigenetic modifications to randomly silence one X chromosome during early embryogenesis. Human pluripotent stem cells (hPSCs) were reported to present various states of XCI that differ according to the expression of the long non-coding RNA XIST and the degree of X chromosome silencing. To obtain a comprehensive perspective on XCI in female hPSCs, we performed a large-scale analysis characterizing different XCI parameters in more than 700 RNA high-throughput sequencing samples. Our findings suggest differences in XCI status between most published samples of embryonic stem cells (ESCs) and induced PSCs (iPSCs). While the majority of iPSC lines maintain an inactive X chromosome, ESC lines tend to silence the expression of XIST and upregulate distal chromosomal regions. Our study highlights significant epigenetic heterogeneity within hPSCs, which may bear implications for their use in research and regenerative therapy.

Organization and evolution of the chalcone synthase gene family in bread wheat and relative species

BackgroundFlavonoid compounds are secondary plant metabolites, having a functional importance in plant development, protection from pathogens and unfavorable environmental factors. Chalcone synthase (CHS) is a key enzyme in the biosynthesis of flavonoids; it is involved in biosynthesis of all classes of flavonoid compounds. Nevertheless, the Chs gene family in bread wheat (Triticum aestivum L.) has been not characterized yet. The aim of the current study was to investigate structural and functional organization of the Chs genes and evolution of this gene family in bread wheat and relative species.ResultsThe nucleotide sequences of the eight Chs copies in T. aestivum were identified. Among them, two homoeologous sets of the Chs genes were located on the short (Chs-A1, −B1, −D1) and the long (Chs-A4, −B4, −D4) arms of homoeologous group 2 chromosomes. Two paralogous gene copies in the B-genome (Chs-B2, −B3) were located in the distal regions of 2BS chromosome. To clarify the origin of Chs duplications in the B-genome the phylogenetic analysis with the Chs sequences of Triticum and Aegilops species carrying ancestral genomes was conducted. It was estimated that the first duplication event occurred in the genome of the common ancestor of Triticum and Aegilops genera about 10–12 million years ago (MYA), then another copy was formed in the ancestor of the B-genome about 6–7 MYA. A homology modeling revealed high sequence similarity of bread wheat CHS enzymes. A number of short deletions in coding regions of some Chs sequences are not expected to have any significant functional effects. Estimation of transcriptional activity of the Chs copies along with a comparative analysis of their promoters structure suggested their functional specialization, which likely contributed to the maintaining of the duplicated Chs genes in wheat genome.ConclusionsFrom possible ten Chs copies in bread wheat genome, eight members of this family retained their intact structure and activity, while two copies appear to be lost at the level of diploid and tetraploid ancestors. Transcriptional assay along with a comparative analysis of the cis-regulatory elements revealed their functional diversification. The multiple functions supported by the Chs family are assumed to be a driving force for duplications of the Chs gene and their retention in plant genome.

Dissecting the chromosomal composition of mutagen-induced micronuclei in Brachypodium distachyon using multicolour FISH.

Background and Aims Brachypodium distachyon (Brachypodium) is a model species for temperate cereals and other economically important grasses. Its favourable cytogenetic features and advanced molecular infrastructure make it a good model for understanding the mechanisms of instability of plant genomes after mutagenic treatment. The aim of this study was to qualitatively and quantitatively assess the composition and origin of micronuclei arising from genomic fracture, and to detect possible ‘hot spots’ for mutagen-induced DNA breaks.MethodsSeeds of Brachypodium were treated with maleic hydrazide (MH) or X-rays. The structure of mutagen-induced micronuclei was analysed in root-tip meristematic cells using multicolour fluorescence in situ hybridization (mcFISH) with various repetitive (5S rDNA, 25S rDNA, telomeric, centromeric) and low-repeat [small and large pools of bacterial artificial chromosome (BAC) clones specific for chromosome Bd1] DNA sequences.Key ResultsThe majority of micronuclei derive from large, acentric fragments. X-rays caused more interstitial DNA breaks than MH. Double-strand breaks rarely occurred in distal chromosome regions. Bd1 contributed to the formation of more mutagen-induced micronuclei than expected from random chromosome involvement.ConclusionsmcFISH with chromosome-specific BAC clones offers insight into micronuclei composition, in so far as it allows their origin and formation to be determined more specifically. A reliable assay for micronuclei composition is crucial for the development of modern genotoxicity tests using plant cells. The combination of mutagenic treatments and well-developed cytomolecular resources in Brachypodium make this model species very promising for plant mutagenesis research.