scispace - formally typeset
Search or ask a question

Showing papers on "Chromosome published in 2015"


Journal ArticleDOI
09 Jul 2015-Nature
TL;DR: This model implies that the DCC reshapes the topology of X chromosomes by forming new TAD boundaries and reinforcing weak boundaries through interactions between its highest-affinity binding sites, and deletion of an endogenous rex site at a DCC-dependent TAD boundary using CRISPR/Cas9 greatly diminished the boundary.
Abstract: The three-dimensional organization of a genome plays a critical role in regulating gene expression, yet little is known about the machinery and mechanisms that determine higher-order chromosome structure. Here we perform genome-wide chromosome conformation capture analysis, fluorescent in situ hybridization (FISH), and RNA-seq to obtain comprehensive three-dimensional (3D) maps of the Caenorhabditis elegans genome and to dissect X chromosome dosage compensation, which balances gene expression between XX hermaphrodites and XO males. The dosage compensation complex (DCC), a condensin complex, binds to both hermaphrodite X chromosomes via sequence-specific recruitment elements on X (rex sites) to reduce chromosome-wide gene expression by half. Most DCC condensin subunits also act in other condensin complexes to control the compaction and resolution of all mitotic and meiotic chromosomes. By comparing chromosome structure in wild-type and DCC-defective embryos, we show that the DCC remodels hermaphrodite X chromosomes into a sex-specific spatial conformation distinct from autosomes. Dosage-compensated X chromosomes consist of self-interacting domains (∼1 Mb) resembling mammalian topologically associating domains (TADs). TADs on X chromosomes have stronger boundaries and more regular spacing than on autosomes. Many TAD boundaries on X chromosomes coincide with the highest-affinity rex sites and become diminished or lost in DCC-defective mutants, thereby converting the topology of X to a conformation resembling autosomes. rex sites engage in DCC-dependent long-range interactions, with the most frequent interactions occurring between rex sites at DCC-dependent TAD boundaries. These results imply that the DCC reshapes the topology of X chromosomes by forming new TAD boundaries and reinforcing weak boundaries through interactions between its highest-affinity binding sites. As this model predicts, deletion of an endogenous rex site at a DCC-dependent TAD boundary using CRISPR/Cas9 greatly diminished the boundary. Thus, the DCC imposes a distinct higher-order structure onto X chromosomes while regulating gene expression chromosome-wide.

738 citations


Journal ArticleDOI
08 May 2015-Science
TL;DR: This work finds that CENP-C affects nucleosome shape and dynamics in a manner analogous to allosteric regulation of enzymes, and is a prime candidate to stabilize centromeric chromatin.
Abstract: Inheritance of each chromosome depends upon its centromere. A histone H3 variant, centromere protein A (CENP-A), is essential for epigenetically marking centromere location. We find that CENP-A is quantitatively retained at the centromere upon which it is initially assembled. CENP-C binds to CENP-A nucleosomes and is a prime candidate to stabilize centromeric chromatin. Using purified components, we find that CENP-C reshapes the octameric histone core of CENP-A nucleosomes, rigidifies both surface and internal nucleosome structure, and modulates terminal DNA to match the loose wrap that is found on native CENP-A nucleosomes at functional human centromeres. Thus, CENP-C affects nucleosome shape and dynamics in a manner analogous to allosteric regulation of enzymes. CENP-C depletion leads to rapid removal of CENP-A from centromeres, indicating their collaboration in maintaining centromere identity.

203 citations


Journal ArticleDOI
TL;DR: The rust resistance and novel agronomic traits revealed that the wheat–Thinopyrum substitution lines will be potentially useful for genetic improvement of wheat.
Abstract: Two wheat–Thinopyrum substitution lines X479 and X482 selected from the progenies of wheat “Mianyang26 (MY26)” × wheat–Thinopyrum intermedium ssp. trichophorum partial amphiploid were characterized by seed storage protein electrophoresis, genomic in situ hybridization (GISH), fluorescence in situ hybridization (FISH), and PCR-based molecular markers. Seed storage protein analysis showed that X479 expressed some of Th. intermedium ssp. trichophorum-specific gliadin and glutenin bands. Chromosome counting and GISH probed by Pseudoroegneria spicata genomic DNA indicated that two pairs of Thinopyrum-derived chromosomes (St genome and St–JS translocated chromosomes) substituted for two pairs of wheat chromosomes in both X479 and X482. FISH using pAs1 and pHvG38 as probes showed that chromosomes 1B and 4B, and 4D and 6D were absent in X479 and X482, respectively. Using the newly isolated JS chromosome-specific repetitive sequence pDb12H as a probe, the FISH signals revealed that the translocation of St–JS chromosomes in X479 and X482 occurred in repetitive sequence regions of the short arm. The molecular markers based on wheat–rice colinearity confirmed that the chromosome constitutions of X479 and X482 were 1St (1B) + 4St–4JS (4B) and 4St–JS (4D) + 6St (6D), respectively. The substitution lines were both fully fertile which suggests that the Th. intermedium chromosomes in X479 and X482 substitute well for the corresponding wheat chromosomes. The rust resistance and novel agronomic traits revealed that the substitution lines will be potentially useful for genetic improvement of wheat.

157 citations


Journal ArticleDOI
TL;DR: This fully completed F. graminearum PH-1 genome and manually curated annotation, available at Ensembl Fungi, provides the optimum resource to perform interspecies comparative analyses and gene function studies.
Abstract: Accurate genome assembly and gene model annotation are critical for comparative species and gene functional analyses. Here we present the completed genome sequence and annotation of the reference strain PH-1 of Fusarium graminearum, the causal agent of head scab disease of small grain cereals which threatens global food security. Completion was achieved by combining (a) the BROAD Sanger sequenced draft, with (b) the gene predictions from Munich Information Services for Protein Sequences (MIPS) v3.2, with (c) de novo whole-genome shotgun re-sequencing, (d) re-annotation of the gene models using RNA-seq evidence and Fgenesh, Snap, GeneMark and Augustus prediction algorithms, followed by (e) manual curation. We have comprehensively completed the genomic 36,563,796 bp sequence by replacing unknown bases, placing supercontigs within their correct loci, correcting assembly errors, and inserting new sequences which include for the first time complete AT rich sequences such as centromere sequences, subtelomeric regions and the telomeres. Each of the four F. graminearium chromosomes was found to be submetacentric with respect to centromere positioning. The position of a potential neocentromere was also defined. A preferentially higher frequency of genetic recombination was observed at the end of the longer arm of each chromosome. Within the genome 1529 gene models have been modified and 412 new gene models predicted, with a total gene call of 14,164. The re-annotation impacts upon 69 entries held within the Pathogen-Host Interactions database (PHI-base) which stores information on genes for which mutant phenotypes in pathogen-host interactions have been experimentally tested, of which 59 are putative transcription factors, 8 kinases, 1 ATP citrate lyase (ACL1), and 1 syntaxin-like SNARE gene (GzSYN1). Although the completed F. graminearum contains very few transposon sequences, a previously unrecognised and potentially active gypsy-type long-terminal-repeat (LTR) retrotransposon was identified. In addition, each of the sub-telomeres and centromeres contained either a LTR or MarCry-1_FO element. The full content of the proposed ancient chromosome fusion sites has also been revealed and investigated. Regions with high recombination previously noted to be rich in secretome encoding genes were also found to be rich in tRNA sequences. This study has identified 741 F. graminearum species specific genes and provides the first complete genome assembly for a Sordariomycetes species. This fully completed F. graminearum PH-1 genome and manually curated annotation, available at Ensembl Fungi, provides the optimum resource to perform interspecies comparative analyses and gene function studies.

157 citations


Journal ArticleDOI
01 Jul 2015-Genetics
TL;DR: An oligonucleotide (oligo)-based chromosome painting technique in cucumber that will be applicable in any plant species with a sequenced genome is developed and precisely map the pairing between cucumber chromosome 7 and chromosome 1 of Cucumis hystrix in a F1 hybrid.
Abstract: Chromosome-specific painting is a powerful technique in molecular cytogenetic and genome research. We developed an oligonucleotide (oligo)-based chromosome painting technique in cucumber (Cucumis sativus) that will be applicable in any plant species with a sequenced genome. Oligos specific to a single chromosome of cucumber were identified using a newly developed bioinformatic pipeline and then massively synthesized de novo in parallel. The synthesized oligos were amplified and labeled with biotin or digoxigenin for use in fluorescence in situ hybridization (FISH). We developed three different probes with each containing 23,000-27,000 oligos. These probes spanned 8.3-17 Mb of DNA on targeted cucumber chromosomes and had the densities of 1.5-3.2 oligos per kilobases. These probes produced FISH signals on a single cucumber chromosome and were used to paint homeologous chromosomes in other Cucumis species diverged from cucumber for up to 12 million years. The bulked oligo probes allowed us to track a single chromosome in early stages during meiosis. We were able to precisely map the pairing between cucumber chromosome 7 and chromosome 1 of Cucumis hystrix in a F1 hybrid. These two homeologous chromosomes paired in 71% of prophase I cells but only 25% of metaphase I cells, which may provide an explanation of the higher recombination rates compared to the chiasma frequencies between homeologous chromosomes reported in plant hybrids.

148 citations


Journal ArticleDOI
TL;DR: This review discusses the current knowledge of centromere establishment, maintenance, composition, structure, and function in mitosis and presents a brief history ofcentromere research.
Abstract: A fundamental challenge for the survival of all organisms is maintaining the integrity of the genome in all cells. Cells must therefore segregate their replicated genome equally during each cell division. Eukaryotic organisms package their genome into a number of physically distinct chromosomes, which replicate during S phase and condense during prophase of mitosis to form paired sister chromatids. During mitosis, cells form a physical connection between each sister chromatid and microtubules of the mitotic spindle, which segregate one copy of each chromatid to each new daughter cell. The centromere is the DNA locus on each chromosome that creates the site of this connection. In this review, we present a brief history of centromere research and discuss our current knowledge of centromere establishment, maintenance, composition, structure, and function in mitosis.

127 citations


Journal ArticleDOI
TL;DR: A draft assembly of the non-recombining region of the collared flycatcher W chromosome is presented, containing 46 genes without evidence of female-specific functional differentiation, demonstrating evolutionary stable matrilineal inheritance of this nuclear–cytonuclear pair of chromosomes.
Abstract: The typically repetitive nature of the sex-limited chromosome means that it is often excluded from or poorly covered in genome assemblies, hindering studies of evolutionary and population genomic processes in non-recombining chromosomes. Here, we present a draft assembly of the non-recombining region of the collared flycatcher W chromosome, containing 46 genes without evidence of female-specific functional differentiation. Survival of genes during W chromosome degeneration has been highly non-random and expression data suggest that this can be attributed to selection for maintaining gene dose and ancestral expression levels of essential genes. Re-sequencing of large population samples revealed dramatically reduced levels of within-species diversity and elevated rates of between-species differentiation (lineage sorting), consistent with low effective population size. Concordance between W chromosome and mitochondrial DNA phylogenetic trees demonstrates evolutionary stable matrilineal inheritance of this nuclear–cytonuclear pair of chromosomes. Our results show both commonalities and differences between W chromosome and Y chromosome evolution.

124 citations


Journal ArticleDOI
TL;DR: It is shown that a single-point amino acid exchange in the centromere-targeting domain of CENH3 leads to reduced centRomere loading of C ENH3 in barley, sugar beet, and Arabidopsis thaliana.
Abstract: The chromosomal position of the centromere-specific histone H3 variant CENH3 (also called “CENP-A”) is the assembly site for the kinetochore complex of active centromeres. Any error in transcription, translation, modification, or incorporation can affect the ability to assemble intact CENH3 chromatin and can cause centromere inactivation [Allshire RC, Karpen GH (2008) Nat Rev Genet 9 (12):923–937]. Here we show that a single-point amino acid exchange in the centromere-targeting domain of CENH3 leads to reduced centromere loading of CENH3 in barley, sugar beet, and Arabidopsis thaliana. Haploids were obtained after cenh3 L130F-complemented cenh3-null mutant plants were crossed with wild-type A. thaliana. In contrast, in a noncompeting situation (i.e., centromeres possessing only mutated or only wild-type CENH3), no uniparental chromosome elimination occurs during early embryogenesis. The high degree of evolutionary conservation of the identified mutation site offers promising opportunities for application in a wide range of crop species in which haploid technology is of interest.

121 citations


Journal ArticleDOI
TL;DR: These findings suggest that ectopic CENP-A nucleosomes could alter the state of the chromatin fiber, potentially impacting gene regulation and chromosome fragility.
Abstract: Background: The histone H3 variant CENP-A is normally tightly regulated to ensure only one centromere exists per chromosome. Native CENP-A is often found overexpressed in human cancer cells and a range of human tumors. Consequently, CENP-A misregulation is thought to contribute to genome instability in human cancers. However, the consequences of such overexpression have not been directly elucidated in human cancer cells. Results: To investigate native CENP-A overexpression, we sought to uncover CENP-A-associated defects in human cells. We confirm that CENP-A is innately overexpressed in several colorectal cancer cell lines. In such cells, we report that a subset of structurally distinct CENP-A-containing nucleosomes associate with canonical histone H3, and with the transcription-coupled chaperones ATRX and DAXX. Furthermore, such hybrid CENP-A nucleosomes localize to DNase I hypersensitive and transcription factor binding sites, including at promoters of genes across the human genome. A distinct class of CENP-A hotspots also accumulates at subtelomeric chromosomal locations, including at the 8q24/Myc region long-associated with genomic instability. We show this 8q24 accumulation of CENP-A can also be seen in early stage primary colorectal tumors. Conclusions: Our data demonstrate that excess CENP-A accumulates at noncentromeric locations in the human cancer genome. These findings suggest that ectopic CENP-A nucleosomes could alter the state of the chromatin fiber, potentially impacting gene regulation and chromosome fragility.

120 citations


Journal ArticleDOI
TL;DR: It is concluded that features of open chromatin are remarkably stable through mitosis, but are modulated at the level of individual genes and regulatory elements.
Abstract: Mitosis entails global alterations to chromosome structure and nuclear architecture, concomitant with transient silencing of transcription. How cells transmit transcriptional states through mitosis remains incompletely understood. While many nuclear factors dissociate from mitotic chromosomes, the observation that certain nuclear factors and chromatin features remain associated with individual loci during mitosis originated the hypothesis that such mitotically retained molecular signatures could provide transcriptional memory through mitosis. To understand the role of chromatin structure in mitotic memory, we performed the first genome-wide comparison of DNase I sensitivity of chromatin in mitosis and interphase, using a murine erythroblast model. Despite chromosome condensation during mitosis visible by microscopy, the landscape of chromatin accessibility at the macromolecular level is largely unaltered. However, mitotic chromatin accessibility is locally dynamic, with individual loci maintaining none, some, or all of their interphase accessibility. Mitotic reduction in accessibility occurs primarily within narrow, highly DNase hypersensitive sites that frequently coincide with transcription factor binding sites, whereas broader domains of moderate accessibility tend to be more stable. In mitosis, proximal promoters generally maintain their accessibility more strongly, whereas distal regulatory elements tend to lose accessibility. Large domains of DNA hypomethylation mark a subset of promoters that retain accessibility during mitosis and across many cell types in interphase. Erythroid transcription factor GATA1 exerts site-specific changes in interphase accessibility that are most pronounced at distal regulatory elements, but has little influence on mitotic accessibility. We conclude that features of open chromatin are remarkably stable through mitosis, but are modulated at the level of individual genes and regulatory elements.

106 citations


Journal ArticleDOI
21 May 2015-Cell
TL;DR: In this paper, a 4D live-cell and snapshot deconvolution fluorescence imaging was used to analyze mitotic chromosome morphogenesis, showing that chromosomes expand, axes split and straighten, and chromatin loops transit to a radial disposition around now-central axes.

Journal ArticleDOI
TL;DR: A non-monotonic relationship between gene expression and structure is revealed and leads to the hypothesis that gene structure is determined by its function, whereas gene expression is subject to energetic cost.
Abstract: Because of its size, allohexaploid nature, and high repeat content, the bread wheat genome is a good model to study the impact of the genome structure on gene organization, function, and regulation. However, because of the lack of a reference genome sequence, such studies have long been hampered and our knowledge of the wheat gene space is still limited. The access to the reference sequence of the wheat chromosome 3B provided us with an opportunity to study the wheat transcriptome and its relationships to genome and gene structure at a level that has never been reached before. By combining this sequence with RNA-seq data, we construct a fine transcriptome map of the chromosome 3B. More than 8,800 transcription sites are identified, that are distributed throughout the entire chromosome. Expression level, expression breadth, alternative splicing as well as several structural features of genes, including transcript length, number of exons, and cumulative intron length are investigated. Our analysis reveals a non-monotonic relationship between gene expression and structure and leads to the hypothesis that gene structure is determined by its function, whereas gene expression is subject to energetic cost. Moreover, we observe a recombination-based partitioning at the gene structure and function level. Our analysis provides new insights into the relationships between gene and genome structure and function. It reveals mechanisms conserved with other plant species as well as superimposed evolutionary forces that shaped the wheat gene space, likely participating in wheat adaptation.

Journal ArticleDOI
15 May 2015-eLife
TL;DR: Extreme chromosomal restructuring during genome elimination, a process resulting from hybridization of Arabidopsis plants expressing different centromere histones H3, is described, providing a tractable, natural system towards investigating the causes and mechanisms of complex genomic rearrangements similar to those associated with several human disorders.
Abstract: Genome instability is associated with mitotic errors and cancer. This phenomenon can lead to deleterious rearrangements, but also genetic novelty, and many questions regarding its genesis, fate and evolutionary role remain unanswered. Here, we describe extreme chromosomal restructuring during genome elimination, a process resulting from hybridization of Arabidopsis plants expressing different centromere histones H3. Shattered chromosomes are formed from the genome of the haploid inducer, consistent with genomic catastrophes affecting a single, laggard chromosome compartmentalized within a micronucleus. Analysis of breakpoint junctions implicates breaks followed by repair through non-homologous end joining (NHEJ) or stalled fork repair. Furthermore, mutation of required NHEJ factor DNA Ligase 4 results in enhanced haploid recovery. Lastly, heritability and stability of a rearranged chromosome suggest a potential for enduring genomic novelty. These findings provide a tractable, natural system towards investigating the causes and mechanisms of complex genomic rearrangements similar to those associated with several human disorders.

Journal ArticleDOI
TL;DR: Using genetic approaches, it is proved the existence of three types of sex chromosomes, defining three kinds of males (YZ, YW, and ZZ) and two kinds of females (ZW and WW) in Xenopus tropicalis.
Abstract: Homomorphic sex chromosomes and rapid turnover of sex-determining genes can complicate establishing the sex chromosome system operating in a given species. This difficulty exists in Xenopus tropicalis, an anuran quickly becoming a relevant model for genetic, genomic, biochemical, and ecotoxicological research. Despite the recent interest attracted by this species, little is known about its sex chromosome system. Direct evidence that females are the heterogametic sex, as in the related species Xenopus laevis, has yet to be presented. Furthermore, X. laevis’ sex-determining gene, DM-W, does not exist in X. tropicalis, and the sex chromosomes in the two species are not homologous. Here we identify X. tropicalis’ sex chromosome system by integrating data from (i) breeding sex-reversed individuals, (ii) gynogenesis, (iii) triploids, and (iv) crosses among several strains. Our results indicate that at least three different types of sex chromosomes exist: Y, W, and Z, observed in YZ, YW, and ZZ males and in ZW and WW females. Because some combinations of parental sex chromosomes produce unisex offspring and other distorted sex ratios, understanding the sex-determination systems in X. tropicalis is critical for developing this flexible animal model for genetics and ecotoxicology.

Journal ArticleDOI
TL;DR: Gene syntenic and collinear comparisons revealed macrosynteny between sex chromosomes and the corresponding autosomes between these two lineages and provided valuable information for better understanding the genetics and evolution of sex chromosome in dioecious plants.
Abstract: Willows (Salix) and poplars (Populus) are dioecious plants in Salicaceae family Sex chromosome in poplar genome was consistently reported to be associated with chromosome XIX In contrast to poplar, this study revealed that chromosome XV was sex chromosome in willow Previous studies revealed that both ZZ/ZW and XX/XY sex-determining systems could be present in some species of Populus In this study, sex of S suchowensis was found to be determined by the ZW system in which the female was the heterogametic gender Gene syntenic and collinear comparisons revealed macrosynteny between sex chromosomes and the corresponding autosomes between these two lineages By contrast, no syntenic segments were found to be shared between poplar's and willow's sex chromosomes Syntenic analysis also revealed substantial chromosome rearrangements between willow's alternate sex chromatids Since willow and poplar originate from a common ancestor, we proposed that evolution of autosomes into sex chromosomes in these two lineages occurred after their divergence Results of this study indicate that sex chromosomes in Salicaceae are still at the early stage of evolutionary divergence Additionally, this study provided valuable information for better understanding the genetics and evolution of sex chromosome in dioecious plants

Journal ArticleDOI
TL;DR: An unexpected degree of haploidy is revealed in CHO DXB11 and CHO cells in general and the data indicate that chromosome one and four appear to be more stable over the course of the CHO evolution compared to the other chromosomes thus might presenting the most attractive landing platforms for knock-ins of heterologous genes.
Abstract: The DHFR negative CHO DXB11 cell line (also known as DUX-B11 and DUKX) was historically the first CHO cell line to be used for large scale production of heterologous proteins and is still used for production of a number of complex proteins. Here we present the genomic sequence of the CHO DXB11 genome sequenced to a depth of 33x. Overall a significant genomic drift was seen favoring GC → AT point mutations in line with the chemical mutagenesis strategy used for generation of the cell line. The sequencing depth for each gene in the genome revealed distinct peaks at sequencing depths of 0x, 16x, 33x and 49x coverage corresponding to a copy number in the genome of 0, 1, 2 and 3 copies. This indicate that 17% of the genes are haploid revealing a large number of genes which can be knocked out with relative ease. This tendency of haploidy was furthermore shown to be present in eight additional analyzed CHO genomes (15-20% haploidy) but not in the genome of the Chinese hamster. The dhfr gene is confirmed to be haploid in CHO DXB11; transcriptionally active and the remaining allele contains a G410C point mutation causing a Thr137Arg missense mutation. We find ~2.5 million single nucleotide polymorphisms (SNP’s), 44 gene deletions in the CHO DXB11 genome and 9357 SNP's, which interfere with the coding regions of 3458 genes. Copy number variations for nine CHO genomes were mapped to the chromosomes of the Chinese hamster showing unique signatures for each chromosome. The data indicate that chromosome one and four appear to be more stable over the course of the CHO evolution compared to the other chromosomes thus might presenting the most attractive landing platforms for knock-ins of heterologous genes. Our studies reveal an unexpected degree of haploidy in CHO DXB11 and CHO cells in general and highlight the chromosomal changes that have occurred among the CHO cell lines sequenced to date.

Journal ArticleDOI
TL;DR: It is demonstrated that holocentrics of the sedge (Cyperaceae) Rhynchospora pubera possess different classes of centromere-specific repeats, and that Holocentromeres of metaphase chromosomes are composed of multiple centromeric units rather than possessing a diffuse organization, thus favoring the polycentric model.
Abstract: Holocentric chromosomes lack a primary constriction, in contrast to monocentrics. They form kinetochores distributed along almost the entire poleward surface of the chromatids, to which spindle fibers attach. No centromere-specific DNA sequence has been found for any holocentric organism studied so far. It was proposed that centromeric repeats, typical for many monocentric species, could not occur in holocentrics, most likely because of differences in the centromere organization. Here we show that the holokinetic centromeres of the Cyperaceae Rhynchospora pubera are highly enriched by a centromeric histone H3 variant-interacting centromere-specific satellite family designated “Tyba” and by centromeric retrotransposons (i.e., CRRh) occurring as genome-wide interspersed arrays. Centromeric arrays vary in length from 3 to 16 kb and are intermingled with gene-coding sequences and transposable elements. We show that holocentromeres of metaphase chromosomes are composed of multiple centromeric units rather than possessing a diffuse organization, thus favoring the polycentric model. A cell-cycle–dependent shuffling of multiple centromeric units results in the formation of functional (poly)centromeres during mitosis. The genome-wide distribution of centromeric repeat arrays interspersing the euchromatin provides a previously unidentified type of centromeric chromatin organization among eukaryotes. Thus, different types of holocentromeres exist in different species, namely with and without centromeric repetitive sequences.

Journal ArticleDOI
TL;DR: Recent progress in understanding point, regional and satellite centromeres, as well as less well-studied centromere types, such as holocentromeres are reviewed.

Journal ArticleDOI
TL;DR: The role of TEs in the initial steps of differentiation and evolution of sex chromosomes is highlighted, and it is clearly demonstrated that the observed TE-rich sex determination locus originated from a TE-poor genomic region, strengthening the link between TE accumulation and emergence of the sex determination Locus.
Abstract: In many organisms, the sex chromosome pair can be recognized due to heteromorphy; the Y and W chromosomes have often lost many genes due to the absence of recombination during meiosis and are frequently heterochromatic. Repetitive sequences are found at a high proportion on such heterochromatic sex chromosomes and the evolution and emergence of sex chromosomes has been connected to the dynamics of repeats and transposable elements. With an amazing plasticity of sex determination mechanisms and numerous instances of independent emergence of novel sex chromosomes, fish represent an excellent lineage to investigate the early stages of sex chromosome differentiation, where sex chromosomes often are homomorphic and not heterochromatic. We have analyzed the composition, distribution, and relative age of TEs from available sex chromosome sequences of seven teleost fish. We observed recent bursts of TEs and simple repeat accumulations around young sex determination loci. More strikingly, we detected transposable element (TE) amplifications not only on the sex determination regions of the Y and W sex chromosomes, but also on the corresponding regions of the X and Z chromosomes. In one species, we also clearly demonstrated that the observed TE-rich sex determination locus originated from a TE-poor genomic region, strengthening the link between TE accumulation and emergence of the sex determination locus. Altogether, our results highlight the role of TEs in the initial steps of differentiation and evolution of sex chromosomes.

Journal ArticleDOI
01 Aug 2015-Genetics
TL;DR: It is shown that the latest improvements of the PacBio sequencing yield assembly of the whole genome of the anther-smut fungus, Microbotryum lychnidis-dioicae, into finished chromosomes or chromosome arms, even for the repeat-rich mating-type chromosomes and centromeres, highlighting the potential of the latest breakthrough PacBio chemistry to resolve complex genome architectures.
Abstract: Sex chromosomes in plants and animals and fungal mating-type chromosomes often show exceptional genome features, with extensive suppression of homologous recombination and cytological differentiation between members of the diploid chromosome pair. Despite strong interest in the genetics of these chromosomes, their large regions of suppressed recombination often are enriched in transposable elements and therefore can be challenging to assemble. Here we show that the latest improvements of the PacBio sequencing yield assembly of the whole genome of the anther-smut fungus, Microbotryum lychnidis-dioicae (the pathogenic fungus causing anther-smut disease of Silene latifolia), into finished chromosomes or chromosome arms, even for the repeat-rich mating-type chromosomes and centromeres. Suppressed recombination of the mating-type chromosomes is revealed to span nearly 90% of their lengths, with extreme levels of rearrangements, transposable element accumulation, and differentiation between the two mating types. We observed no correlation between allelic divergence and physical position in the nonrecombining regions of the mating-type chromosomes. This may result from gene conversion or from rearrangements of ancient evolutionary strata, i.e., successive steps of suppressed recombination. Centromeres were found to be composed mainly of copia-like transposable elements and to possess specific minisatellite repeats identical between the different chromosomes. We also identified subtelomeric motifs. In addition, extensive signs of degeneration were detected in the nonrecombining regions in the form of transposable element accumulation and of hundreds of gene losses on each mating-type chromosome. Furthermore, our study highlights the potential of the latest breakthrough PacBio chemistry to resolve complex genome architectures.

Journal ArticleDOI
TL;DR: The Sry transgene in FCG mice is present in multiple copies at one locus on chromosome 3, which does not interrupt known genes, suggesting that differences between XX and XY mice with the same type of gonad are not caused by difference in prenatal androgen levels.
Abstract: The “four core genotypes” (FCG) mouse model has emerged as a major model testing if sex differences in phenotypes are caused by sex chromosome complement (XX vs. XY) or gonadal hormones or both. The model involves deletion of the testis-determining gene Sry from the Y chromosome and insertion of an Sry transgene onto an autosome. It produces XX and XY mice with testes, and XX and XY mice with ovaries, so that XX and XY mice with the same type of gonad can be compared to assess phenotypic effects of sex chromosome complement in cells and tissues. We used PCR to amplify the Sry transgene and adjacent genomic sequences, to resolve the location of the Sry transgene to chromosome 3 and confirmed this location by fluorescence in situ hybridization (FISH) of the Sry construct to metaphase chromosomes. Using quantitative PCR, we estimate that 12–14 copies of the transgene were inserted. The anogenital distance (AGD) of FCG pups at 27–29 days after birth was not different in XX vs. XY males, or XX vs. XY females, suggesting that differences between XX and XY mice with the same type of gonad are not caused by difference in prenatal androgen levels. The Sry transgene in FCG mice is present in multiple copies at one locus on chromosome 3, which does not interrupt known genes. XX and XY mice with the same type of gonad do not show evidence of different androgen levels prenatally.

Journal ArticleDOI
TL;DR: It is shown that SMCHD1 forms an active GHKL-ATPase homodimer, contrasting with canonical SMC complexes, which exist as tripartite ring structures, and that a parallel pathway accounts for chromatin loading at a minority of sites, notably the inactive X chromosome.
Abstract: The chromosomal protein SMCHD1 plays an important role in epigenetic silencing at diverse loci, including the inactive X chromosome, imprinted genes, and the facioscapulohumeral muscular dystrophy locus. Although homology with canonical SMC family proteins suggests a role in chromosome organization, the mechanisms underlying SMCHD1 function and target site selection remain poorly understood. Here we show that SMCHD1 forms an active GHKL-ATPase homodimer, contrasting with canonical SMC complexes, which exist as tripartite ring structures. Electron microscopy analysis demonstrates that SMCHD1 homodimers structurally resemble prokaryotic condensins. We further show that the principal mechanism for chromatin loading of SMCHD1 involves an LRIF1-mediated interaction with HP1γ at trimethylated histone H3 lysine 9 (H3K9me3)-modified chromatin sites on the chromosome arms. A parallel pathway accounts for chromatin loading at a minority of sites, notably the inactive X chromosome. Together, our results provide key insights into SMCHD1 function and target site selection.

Journal ArticleDOI
TL;DR: It is shown that concurrent delivery of gRNAs designed to target two different sites in a human chromosome introduce DNA double-strand breaks in the chromosome and give rise to targeted deletions of the intervening genomic segment.
Abstract: : The CRISPR/Cas9 system has emerged as an intriguing new technology for genome engineering. It utilizes the bacterial endonuclease Cas9 which, when delivered to eukaryotic cells in conjunction with a user-specified small guide RNA (gRNA), cleaves the chromosomal DNA at the target site. Here we show that concurrent delivery of gRNAs designed to target two different sites in a human chromosome introduce DNA double-strand breaks in the chromosome and give rise to targeted deletions of the intervening genomic segment. Predetermined genomic DNA segments ranging from several-hundred base pairs to 1 Mbp can be precisely deleted at frequencies of 1% to 10%, with no apparent correlation between the size of the deleted fragment and the deletion frequency. The high efficiency of this technique holds promise for large genomic deletions that could be useful in generation of cell and animal models with engineered chromosomes. Keywords : CRISPR/Cas9; large genomic deletion; HPRT

Journal ArticleDOI
TL;DR: Findings indicate that Y-linked gene loss emerges as an additional driver of gene transposition from the sex chromosomes, a phenomenon thought to be driven primarily by meiotic sex chromosome inactivation, and is widespread among mammalian species.
Abstract: Although the mammalian X and Y chromosomes evolved from a single pair of autosomes, they are highly differentiated: the Y chromosome is dramatically smaller than the X and has lost most of its genes. The surviving genes are a specialized set with extraordinary evolutionary longevity. Most mammalian lineages have experienced delayed, or relatively recent, loss of at least one conserved Y-linked gene. An extreme example of this phenomenon is in the Japanese spiny rat, where the Y chromosome has disappeared altogether. In this species, many Y-linked genes were rescued by transposition to new genomic locations, but until our work presented here, this has been considered an isolated case. We describe eight cases of genes that have relocated to autosomes in mammalian lineages where the corresponding Y-linked gene has been lost. These gene transpositions originated from either the X or Y chromosomes, and are observed in diverse mammalian lineages: occurring at least once in marsupials, apes, and cattle, and at least twice in rodents and marmoset. For two genes - EIF1AX/Y and RPS4X/Y - transposition to autosomes occurred independently in three distinct lineages. Rescue of Y-linked gene loss through transposition to autosomes has previously been reported for a single isolated rodent species. However, our findings indicate that this compensatory mechanism is widespread among mammalian species. Thus, Y-linked gene loss emerges as an additional driver of gene transposition from the sex chromosomes, a phenomenon thought to be driven primarily by meiotic sex chromosome inactivation.

Journal ArticleDOI
TL;DR: Evidence is provided that, in Silene latifolia, this largely involved losses of Y-linked genes, and not suppressed expression ofY-linked alleles, or gene additions to the X chromosome, which suggests that chromosome-wide dosage compensation does not occur in this plant.

Journal ArticleDOI
TL;DR: The findings described here provide the first evidence for a highly dynamic centromere structure within a group of closely related species, challenging previous concepts of centromeret evolution.
Abstract: In most eukaryotes, centromere is determined by the presence of the centromere-specific histone variant CenH3. Two types of chromosome morphology are generally recognized with respect to centromere organization. Monocentric chromosomes possess a single CenH3-containing domain in primary constriction, whereas holocentric chromosomes lack the primary constriction and display dispersed distribution of CenH3. Recently, metapolycentric chromosomes have been reported in Pisum sativum, representing an intermediate type of centromere organization characterized by multiple CenH3-containing domains distributed across large parts of chromosomes that still form a single constriction. In this work, we show that this type of centromere is also found in other Pisum and closely related Lathyrus species, whereas Vicia and Lens genera, which belong to the same legume tribe Fabeae, possess only monocentric chromosomes. We observed extensive variability in the size of primary constriction and the arrangement of CenH3 domains both between and within individual Pisum and Lathyrus species, with no obvious correlation to genome or chromosome size. Search for CenH3 gene sequences revealed two paralogous variants, CenH3-1 and CenH3-2, which originated from a duplication event in the common ancestor of Fabeae species. The CenH3-1 gene was subsequently lost or silenced in the lineage leading to Vicia and Lens, whereas both genes are retained in Pisum and Lathyrus. Both of these genes appear to have evolved under purifying selection and produce functional CenH3 proteins which are fully colocalized. The findings described here provide the first evidence for a highly dynamic centromere structure within a group of closely related species, challenging previous concepts of centromere evolution.

Journal ArticleDOI
TL;DR: Oocyte loss cannot be explained simply by asynapsis checkpoint models, but is related to the gene content of asynapsed chromosomes, despite being enriched for γH2AFX and other checkpoint proteins.
Abstract: Chromosome abnormalities are common in the human population, causing germ cell loss at meiotic prophase I and infertility. The mechanisms driving this loss are unknown, but persistent meiotic DNA damage and asynapsis may be triggers. Here we investigate the contribution of these lesions to oocyte elimination in mice with chromosome abnormalities, e.g. Turner syndrome (XO) and translocations. We show that asynapsed chromosomes trigger oocyte elimination at diplonema, which is linked to the presence of phosphorylated H2AFX (γH2AFX). We find that DNA double-strand break (DSB) foci disappear on asynapsed chromosomes during pachynema, excluding persistent DNA damage as a likely cause, and demonstrating the existence in mammalian oocytes of a repair pathway for asynapsis-associated DNA DSBs. Importantly, deletion or point mutation of H2afx restores oocyte numbers in XO females to wild type (XX) levels. Unexpectedly, we find that asynapsed supernumerary chromosomes do not elicit prophase I loss, despite being enriched for γH2AFX and other checkpoint proteins. These results suggest that oocyte loss cannot be explained simply by asynapsis checkpoint models, but is related to the gene content of asynapsed chromosomes. A similar mechanistic basis for oocyte loss may operate in humans with chromosome abnormalities.

Dissertation
28 Dec 2015
TL;DR: In this paper, the effect of spontaneous tetraploidization on colorectal cancer genomes has been investigated using a siRNA screen of genes commonly mutated in genome-doubled tumours of multiple cancer types.
Abstract: Tetraploidy has long been proposed as an intermediate cellular stage en route to the aneuploidy and chromosomal instability that is observed in many cancer types. Although tetraploidy has been shown to be an unstable cellular state, an in depth analysis of the effect of a spontaneous tetraploidisation event on the cancer genome has not been carried out. Using an isogenic system of naturally occurring tetraploid cells derived from a chromosomally stable colorectal cancer cell line, the effect of tetraploidisation on genome stability over time was assessed. Tetraploid cells were shown to have increased structural and numerical instability on a per cell but not per chromosome basis. Over time the tetraploid genome became increasingly genomically unstable, which is likely due to the increased ability of tetraploid clones to propagate segregation errors. The genomic landscape of tetraploid clones began to recapitulate the genomic architecture of chromosomally unstable colorectal cancer, and allowed for the selection of clinically high risk chromosomal losses over time. Genome doubling was further shown to be a predictive marker of poor prognosis in colorectal cancer. Exhaustive analysis of DNA and mRNA failed to reveal any common changes in tetraploid clones that are likely to explain their aneuploidy tolerance phenotype. Instead a focussed siRNA screen of genes commonly mutated in genome-doubled tumours of multiple cancer types was carried out. Given the high-risk clinical phenotypes associated with tetraploidy and chromosomal instability, it remains a priority to identify the mechanisms allowing tumour cells to undergo tetraploidisation and to sustain chromosome segregation errors.

Journal ArticleDOI
TL;DR: In several angiosperms affected by WGD, it is shown that chromosome number reduction (CNR) is best explained by intra- and/or inter-chromosomal crossovers to form new chromosomes that utilize the existing telomeres of 'invaded' and centromeres of '.
Abstract: Whole-genome duplication (WGD) is central to the evolution of many eukaryotic genomes, in particular rendering angiosperm (flowering plant) genomes much less stable than those of animals. Following repeated duplication/triplication(s), angiosperm chromosome numbers have usually been restored to a narrow range, as one element in a 'diploidization' process that re-establishes diploid heredity. In several angiosperms affected by WGD, we show that chromosome number reduction (CNR) is best explained by intra- and/or inter-chromosomal crossovers to form new chromosomes that utilize the existing telomeres of 'invaded' and centromeres of 'invading' chromosomes, the alternative centromeres and telomeres being lost. Comparison with the banana (Musa acuminata) genome supports a 'fusion model' for the evolution of rice (Oryza sativa) chromosomes 2 and 3, implying that the grass common ancestor had seven chromosomes rather than the five implied by a 'fission model.' The 'invading' and 'invaded' chromosomes are frequently homoeologs, originating from duplication of a common ancestral chromosome and with greater-than-average DNA-level correspondence to one another. Telomere-centric CNR following recursive WGD in plants is also important in mammals and yeast, and may be a general mechanism of restoring small linear chromosome numbers in higher eukaryotes.

Journal ArticleDOI
TL;DR: This review highlights the involvement of selected Y chromosome genes in cancer development in men, and suggests that the Y-linked genes are involved in male health and diseases in more frequently than expected.
Abstract: Male and female differ genetically by their respective sex chromosome composition, that is, XY as male and XX as female. Although both X and Y chromosomes evolved from the same ancestor pair of autosomes, the Y chromosome harbors male-specific genes, which play pivotal roles in male sex determination, germ cell differentiation, and masculinization of various tissues. Deletions or translocation of the sex-determining gene, SRY, from the Y chromosome causes disorders of sex development (previously termed as an intersex condition) with dysgenic gonads. Failure of gonadal development results not only in infertility, but also in increased risks of germ cell tumor (GCT), such as gonadoblastoma and various types of testicular GCT. Recent studies demonstrate that either loss of Y chromosome or ectopic expression of Y chromosome genes is closely associated with various male-biased diseases, including selected somatic cancers. These observations suggest that the Y-linked genes are involved in male health and diseases in more frequently than expected. Although only a small number of protein-coding genes are present in the male-specific region of Y chromosome, the impacts of Y chromosome genes on human diseases are still largely unknown, due to lack of in vivo models and differences between the Y chromosomes of human and rodents. In this review, we highlight the involvement of selected Y chromosome genes in cancer development in men.