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Shigeyuki Kawano - One of the best experts on this subject based on the ideXlab platform.

  • an asexual flower of Silene latifolia and microbotryum lychnidis dioicae promotes sex organ development
    PLOS ONE, 2019
    Co-Authors: Hiroki Kawamoto, Yusuke Kazama, Ayako Koizumi, Tomoko Abe, Kaori Yamanaka, Kotaro Ishii, Shigeyuki Kawano
    Abstract:

    Silene latifolia is a dioecious flowering plant with sex chromosomes in the family Caryophyllaceae. Development of a gynoecium and stamens are suppressed in the male and female flowers of S. latifolia, respectively. Microbotryum lychnidis-dioicae promotes stamen development when it infects the female flower. If suppression of the stamen and gynoecium development is regulated by the same mechanism, suppression of gynoecium and stamen development is released simultaneously with the infection by M. lychnidis-dioicae. To assess this hypothesis, an asexual mutant without a gynoecium or stamen was infected with M. lychnidis-dioicae. A filament of the stamen in the infected asexual mutant was elongated at stages 11 and 12 of flower bud development as well as in the male, but the gynoecium did not form. Instead of the gynoecium, a filamentous structure was suppressed as in the male flower. Developmental suppression of the stamen was released by M. lychnidis-dioicae, but that of gynoecium development was not released. M. lychnidis-dioicae would have a function similar to stamen-promoting factor (SPF), since the elongation of the stamen that is not observed in the healthy asexual mutant was observed after stage 8 of flower bud development. An infection experiment also revealed that a deletion on the Y chromosome of the asexual mutant eliminated genes for maturation of tapetal cells because the tapetal cells did not mature in the asexual mutant infected with M. lychnidis-dioicae.

  • an asexual flower of Silene latifolia and microbotryum lychnidis dioicae promoting its sexual organ development
    bioRxiv, 2019
    Co-Authors: Hiroki Kawamoto, Yusuke Kazama, Ayako Koizumi, Tomoko Abe, Kaori Yamanaka, Kotaro Ishii, Shigeyuki Kawano
    Abstract:

    Abstract Silene latifolia is a dioecious flowering plant with sex chromosomes in the family Caryophyllaceae. Development of a gynoecium and stamens are suppressed in the male and female flowers of S. latifolia, respectively. Microbtryum lychnidis-dioicae promotes stamen development when it infects the female flower. If suppression of the stamen and gynoecium development is regulated by the same mechanism, suppression of gynoecium and stamen development is released simultaneously with the infection by M. lychnidis-dioicae. To assess this hypothesis, an asexual mutant, without gynoecium or stamen, was infected with M. lychnidis-dioicae. A filament of the stamen in the infected asexual mutant was elongated at stages 11 and 12 of the flower bud development as well as the male, but the gynoecium did not form. Instead of the gynoecium, a filamentous structure was suppressed as in the male flower. Developmental suppression of the stamen was released by M. lychnidis-dioicae, but that of gynoecium development was not released. It is thought, therefore, that the suppression of gynoecium development was not released by the infection of M. lychnidis-dioicae. M. lychnidis-dioicae would have a function similar to SPF since the elongation of the stamen that is not observed in the healthy asexual mutant was observed after stage 8 of flower bud development. Such an infection experiment also that the Y chromosome of the asexual mutant has genes related to the differentiation of archesporial cells, but none related to maturation of the tapetal cells.

  • development of the vigs system in the dioecious plant Silene latifolia
    International Journal of Molecular Sciences, 2019
    Co-Authors: Naoko Fujita, Yusuke Kazama, Shigeyuki Kawano, Noriko Yamagishi, Kyoko Watanabe, Saki Ando, Hiroyuki Tsuji, Nobuyuki Yoshikawa, Ken Komatsu
    Abstract:

    (1) Background: Silene latifolia is a dioecious plant, whose sex is determined by XY-type sex chromosomes. Microbotryum lychnidis-dioicae is a smut fungus that infects S. latifolia plants and causes masculinization in female flowers, as if Microbotryum were acting as a sex-determining gene. Recent large-scale sequencing efforts have promised to provide candidate genes that are involved in the sex determination machinery in plants. These candidate genes are to be analyzed for functional characterization. A virus vector can be a tool for functional gene analyses; (2) Methods: To develop a viral vector system in S. latifolia plants, we selected Apple latent spherical virus (ALSV) as an appropriate virus vector that has a wide host range; (3) Results: Following the optimization of the ALSV inoculation method, S. latifolia plants were infected with ALSV at high rates in the upper leaves. In situ hybridization analysis revealed that ALSV can migrate into the flower meristems in S. latifolia plants. Successful VIGS (virus-induced gene silencing) in S. latifolia plants was demonstrated with knockdown of the phytoene desaturase gene. Finally, the developed method was applied to floral organ genes to evaluate its usability in flowers; (4) Conclusion: The developed system enables functional gene analyses in S. latifolia plants, which can unveil gene functions and networks of S. latifolia plants, such as the mechanisms of sex determination and fungal-induced masculinization.

  • three dimensional ultrastructural study of the anther of Silene latifolia infected with microbotryum lychnidis dioicae
    PLOS ONE, 2017
    Co-Authors: Hiroki Kawamoto, Aiko Hirata, Shigeyuki Kawano
    Abstract:

    When Microbotryum lychnidis-dioicae infects a male Silene latifolia, M. lychnidis-dioicae smut spores develop in the pollen sac instead of pollen. In contrast, when M. lychnidis-dioicae infects a female S. latifolia, the female flowers become male-like, promoting stamen formation. However, it is unclear when and how M. lychnidis-dioicae invades the anther. It is important to investigate not only whether hyphae exist when the apical meristem tissue differentiates into flowers and anthers, but also whether hyphae exist when stamen filaments form. We used Grocott’s methenamine silver stain and lectin stain, which stain chitin in the fungal cell wall, to search for M. lychnidis-dioicae in flower tissues. A few M. lychnidis-dioicae hyphae were observed intercellularly in the center of the connective of vascular bundles at the early anther developmental stage. Subsequently, large numbers of deeply stained M. lychnidis-dioicae hyphae were observed intercellularly in the cells surrounding the pollen sac, as well as in the center of the pollen sac. Hyphae stained with lectin were observed intercellularly in all of the stamen filaments at flower development stages. Hyphae were observed in the peduncle connecting the flower and stem. It is thought that M. lychnidis-dioicae invaded the anther via the stamen filament over a long period. Additionally, in total, 163 sections of connective were obtained, and the cell structure of each anther was colored and subjected to three-dimensional reconstruction. The M. lychnidis-dioicae hyphae observed in the connective were mainly old hyphae with large vacuoles or dead hyphae (S1 Fig). These hyphae branched out, towards the pollen sac, while growing between the cells. We also observed that the host cells that collapsed near the hyphae had thick cell walls and teliospores. Cell wall collapse and cell degeneration were observed only around hyphae with thick cell walls.

  • cell death and cell cycle arrest of Silene latifolia stamens and pistils after microbotryum lychnidis dioicae infection
    Plant and Cell Physiology, 2016
    Co-Authors: Hiroki Kawamoto, Ayako Koizumi, Kaori Yamanaka, Aiko Hirata, Shigeyuki Kawano
    Abstract:

    Mechanisms of suppression of pistil primordia in male flowers and of stamen primordia in female flowers differ in diclinous plants. In this study, we investigated how cell death and cell cycle arrest are related to flower organ formation in Silene latifolia. Using in situ hybridization and a TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) assay, we detected both cell cycle arrest and cell death in suppressed stamens of female flowers and suppressed pistils of male flowers in S. latifolia. In female flowers infected with Microbotryum lychnidis-dioicae, developmental suppression of stamens is released, and cell cycle arrest and cell death do not occur. Smut spores are formed in S. latifolia anthers infected with M. lychnidis-dioicae, followed by cell death in the endothelium, middle layer, tapetal cells and pollen mother cells. Cell death is difficult to detect using a fluorescein isothiocyanate-labeled TUNEL assay due to strong autofluorescence in the anther. We therefore combined a TUNEL assay in an infrared region with transmission electron microscopy to detect cell death in anthers. We show that following infection by M. lychnidis-dioicae, a TUNEL signal was not detected in the endothelium, middle layer or pollen mother cells, and cell death with outflow of cell contents, including the nucleoplast, was observed in tapetal cells.

Dmitry A Filatov - One of the best experts on this subject based on the ideXlab platform.

  • the location of the pseudoautosomal boundary in Silene latifolia
    Genes, 2020
    Co-Authors: Marc Krasovec, Yu Zhang, Dmitry A Filatov
    Abstract:

    Y-chromosomes contain a non-recombining region (NRY), and in many organisms it was shown that the NRY expanded over time. How and why the NRY expands remains unclear. Young sex chromosomes, where NRY expansion occurred recently or is on-going, offer an opportunity to study the causes of this process. Here, we used the plant Silene latifolia, where sex chromosomes evolved ~11 million years ago, to study the location of the boundary between the NRY and the recombining pseudoautosomal region (PAR). The previous work devoted to the NRY/PAR boundary in S. latifolia was based on a handful of genes with locations approximately known from the genetic map. Here, we report the analysis of 86 pseudoautosomal and sex-linked genes adjacent to the S. latifolia NRY/PAR boundary to establish the location of the boundary more precisely. We take advantage of the dense genetic map and polymorphism data from wild populations to identify 20 partially sex-linked genes located in the “fuzzy boundary”, that rarely recombines in male meiosis. Genes proximal to this fuzzy boundary show no evidence of recombination in males, while the genes distal to this partially-sex-linked region are actively recombining in males. Our results provide a more accurate location for the PAR boundary in S. latifolia, which will help to elucidate the causes of PAR boundary shifts leading to NRY expansion over time.

  • immediate dosage compensation is triggered by the deletion of y linked genes in Silene latifolia
    Current Biology, 2019
    Co-Authors: Marc Krasovec, Tomoko Abe, Yusuke Kazama, Kotaro Ishii, Dmitry A Filatov
    Abstract:

    Summary The loss of functional genes from non-recombining sex-specific chromosomes [ 1 , 2 ], such as the Y chromosomes in mammals [ 3 ] or W chromosomes in birds [ 4 ], should result in an imbalance of gene products for sex-linked genes [ 5 ]. Different chromosome-wide systems that rebalance gene expression are known to operate in organisms with relatively old sex chromosomes [ 6 ]; e.g., Drosophila overexpress X-linked genes in males [ 7 ], while mammals shut down one of the X chromosomes in females [ 8 ]. It is not known how long it takes for a chromosome-wide dosage compensation system to evolve. To shed light on the early evolution of dosage compensation, we constructed a high-density Y-deletion map and used deletion mutants to manipulate gene dose and analyze gene expression in white campion (Silene latifolia), which evolved dioecy and sex chromosomes only 11 million years ago [ 9 ]. We demonstrate that immediate dosage compensation can be triggered by deletions in a large portion of the p arm of the Y chromosome. Our results indicate that dosage compensation in S. latifolia does not have to evolve gene by gene because a system to upregulate gene expression is already operating on part of the X chromosome, which likely represents an intermediate step in the evolution of a chromosome-wide dosage compensation system in this species.

  • the mutation rate and the age of the sex chromosomes in Silene latifolia
    Current Biology, 2018
    Co-Authors: Marc Krasovec, Michael Chester, Kate Ridout, Dmitry A Filatov
    Abstract:

    Summary Many aspects of sex chromosome evolution are common to both plants and animals [1], but the process of Y chromosome degeneration, where genes on the Y become non-functional over time, may be much slower in plants due to purifying selection against deleterious mutations in the haploid gametophyte [2, 3]. Testing for differences in Y degeneration between the kingdoms has been hindered by the absence of accurate age estimates for plant sex chromosomes. Here, we used genome resequencing to estimate the spontaneous mutation rate and the age of the sex chromosomes in white campion ( Silene latifolia ). Screening of single nucleotide polymorphisms (SNPs) in parents and 10 F 1 progeny identified 39 de novo mutations and yielded a rate of 7.31 × 10 −9 (95% confidence interval: 5.20 × 10 −9 − 8.00 × 10 −9 ) mutations per site per haploid genome per generation. Applying this mutation rate to the synonymous divergence between homologous X- and Y-linked genes (gametologs) gave age estimates of 11.00 and 6.32 million years for the old and young strata, respectively. Based on SNP segregation patterns, we inferred which genes were Y-linked and found that at least 47% are already dysfunctional. Applying our new estimates for the age of the sex chromosomes indicates that the rate of Y degeneration in S. latifolia is nearly 2-fold slower when compared to animal sex chromosomes of a similar age. Our revised estimates support Y degeneration taking place more slowly in plants, a discrepancy that may be explained by differences in the life cycles of animals and plants.

  • DOES LOCAL ADAPTATION CAUSE HIGH POPULATION DIFFERENTIATION OF Silene latifolia Y CHROMOSOMES
    Evolution; international journal of organic evolution, 2011
    Co-Authors: Graham Muir, Roberta Bergero, Deborah Charlesworth, Dmitry A Filatov
    Abstract:

    Natural selection can reduce the effective population size of the nonrecombining Y chromosome, whereas local adaptation of Y-linked genes can increase the population divergence and overall intra-species polymorphism of Y-linked sequences. The plant Silene latifolia evolved a Y chromosome relatively recently, and most known X-linked genes have functional Y homologues, making the species interesting for comparisons of X- and Y-linked diversity and subdivision. Y-linked genes show higher population differentiation, compared to X-linked genes, and this might be maintained by local adaptation in Y-linked genes (or low sequence diversity). Here we attempt to test between these causes by investigating DNA polymorphism and population differentiation using a larger set of Y-linked and X-linked S. latifolia genes (than used previously), and show that net sequence divergence for Y-linked sequences (measured by D(a) , also known as δ) is low, and not consistently higher than X-linked genes. This does not support local adaptation, instead, the higher values of differentiation measures for the Y-linked genes probably result largely from reduced total variation on the Y chromosome, which in turn reflect deterministic processes lowering effective population sizes of evolving Y-chromosomes.

  • Dynamic gene order on the Silene latifolia Y chromosome
    Chromosoma, 2011
    Co-Authors: Elaine C. Howell, Susan J. Armstrong, Dmitry A Filatov
    Abstract:

    Dioecious Silene latifolia evolved heteromorphic sex chromosomes within the last ten million years, making it a species of choice for studies of the early stages of sex chromosome evolution in plants. About a dozen genes have been isolated from its sex chromosomes and basic genetic and deletion maps exist for the X and Y chromosomes. However, discrepancies between Y chromosome maps led to the proposal that individual Y chromosomes may differ in gene order. Here, we use an alternative approach, with fluorescence in situ hybridization (FISH), to locate individual genes on S. latifolia sex chromosomes. We demonstrate that gene order on the Y chromosome differs between plants from two populations. We suggest that dynamic gene order may be a general property of Y chromosomes in species with XY systems, in view of recent work demonstrating that the gene order on the Y chromosomes of humans and chimpanzees are dramatically different.

Deborah Charlesworth - One of the best experts on this subject based on the ideXlab platform.

  • has adaptation occurred in males and females since separate sexes evolved in the plant Silene latifolia
    Proceedings of The Royal Society B: Biological Sciences, 2018
    Co-Authors: Niklaus Zemp, Alex Widmer, Deborah Charlesworth
    Abstract:

    The evolution of separate sexes may involve changed expression of many genes, as each sex adapts to its new state. Evidence is accumulating for sex differences in expression even in organisms that have recently evolved separate sexes from hermaphrodite or monoecious (cosexual) ancestors, such as some dioecious flowering plants. We describe evidence that a dioecious plant species with recently evolved dioecy, Silene latifolia, has undergone adaptive changes that improve functioning in females, in addition to changes that are probably pleiotropic effects of male sterility. The results suggest pervasive adaptations as soon as males and females evolve from their cosexual ancestor.

  • expansion of the pseudo autosomal region and ongoing recombination suppression in the Silene latifolia sex chromosomes
    Genetics, 2013
    Co-Authors: Roberta Bergero, Suo Qiu, Alan Forrest, Helen Borthwick, Deborah Charlesworth
    Abstract:

    There are two very interesting aspects to the evolution of sex chromosomes: what happens after recombination between these chromosome pairs stops and why suppressed recombination evolves. The former question has been intensively studied in a diversity of organisms, but the latter has been studied largely theoretically. To obtain empirical data, we used codominant genic markers in genetic mapping of the dioecious plant Silene latifolia, together with comparative mapping of S. latifolia sex-linked genes in S. vulgaris (a related hermaphrodite species without sex chromosomes). We mapped 29 S. latifolia fully sex-linked genes (including 21 newly discovered from transcriptome sequencing), plus 6 genes in a recombining pseudo-autosomal region (PAR) whose genetic map length is ∼25 cM in both male and female meiosis, suggesting that the PAR may contain many genes. Our comparative mapping shows that most fully sex-linked genes in S. latifolia are located on a single S. vulgaris linkage group and were probably inherited from a single autosome of an ancestor. However, unexpectedly, our maps suggest that the S. latifolia PAR region expanded through translocation events. Some genes in these regions still recombine in S. latifolia, but some genes from both addition events are now fully sex-linked. Recombination suppression is therefore still ongoing in S. latifolia, and multiple recombination suppression events have occurred in a timescale of few million years, much shorter than the timescale of formation of the most recent evolutionary strata of mammal and bird sex chromosomes.

  • slwus1 an x linked gene having no homologous y linked copy in Silene latifolia
    G3: Genes Genomes Genetics, 2012
    Co-Authors: Yusuke Kazama, Roberta Bergero, Deborah Charlesworth, Kiyoshi Nishihara, Tomoko Abe, Makoto T Fujiwara, Shigeyuki Kawano
    Abstract:

    The dioecious plant Silene latifolia has heteromorphic sex chromosomes, and comparison of the positions of sex-linked genes indicates that at least three large inversions have occurred during the evolution of the Y chromosome. In this article, we describe the isolation of a new sex-linked gene from S. latifolia, which provides new information on the evolution of this plant’s young sex chromosomes. By using reverse-transcription polymerase chain reaction degenerate primers based on the Arabidopsis thaliana sequence of WUSCHEL, a flower-development gene, we found two copies in S. latifolia, which we named SlWUS1 and SlWUS2. Southern blot and genetic segregation analysis showed that SlWUS1 is located on the X chromosome and SlWUS2 is autosomal. No Y-linked copy of SlWUS1 was found by either Southern blot analysis under low-stringency conditions or polymerase chain reaction with degenerate primers, so we conclude that SlWUS1 probably has no Y-linked homolog. It is unknown whether the Y chromosome lost the SlWUS1 copy by degeneration of this individual gene or whether deletion of a larger genome region was involved. Several tests lead us to conclude that dosage compensation has not evolved for this sex-linked gene. We mapped the ortholog in the nondioecious relative S. vulgaris (SvWUS1), to compare the location in a species that has no history of having sex chromosomes. SvWUS1 maps to the same linkage group as other fully X-linked genes, indicating that it was not added to the X, but was lost from the Y. Its location differs in the maps from the two species, raising the possibility that the X chromosome, as well as the Y, may have been rearranged.

  • DOES LOCAL ADAPTATION CAUSE HIGH POPULATION DIFFERENTIATION OF Silene latifolia Y CHROMOSOMES
    Evolution; international journal of organic evolution, 2011
    Co-Authors: Graham Muir, Roberta Bergero, Deborah Charlesworth, Dmitry A Filatov
    Abstract:

    Natural selection can reduce the effective population size of the nonrecombining Y chromosome, whereas local adaptation of Y-linked genes can increase the population divergence and overall intra-species polymorphism of Y-linked sequences. The plant Silene latifolia evolved a Y chromosome relatively recently, and most known X-linked genes have functional Y homologues, making the species interesting for comparisons of X- and Y-linked diversity and subdivision. Y-linked genes show higher population differentiation, compared to X-linked genes, and this might be maintained by local adaptation in Y-linked genes (or low sequence diversity). Here we attempt to test between these causes by investigating DNA polymorphism and population differentiation using a larger set of Y-linked and X-linked S. latifolia genes (than used previously), and show that net sequence divergence for Y-linked sequences (measured by D(a) , also known as δ) is low, and not consistently higher than X-linked genes. This does not support local adaptation, instead, the higher values of differentiation measures for the Y-linked genes probably result largely from reduced total variation on the Y chromosome, which in turn reflect deterministic processes lowering effective population sizes of evolving Y-chromosomes.

  • patterns of codon usage bias in Silene latifolia
    Molecular Biology and Evolution, 2011
    Co-Authors: Roberta Bergero, Suo Qiu, Kai Zeng, Deborah Charlesworth
    Abstract:

    Patterns of codon usage bias (CUB) convey useful information about the selection on synonymous codons induced by gene expression and contribute to an understanding of substitution patterns observed at synonymous sites. They can also be informative about the distinctive evolutionary properties of sex chromosomes such as genetic degeneration of the Y chromosome, dosage compensation, and hemizygosity of the X chromosome in males, which can affect the selection on codon usage. Here, we study CUB in Silene latifolia, a species of interest for studying the early stages of sex chromosome evolution. We have obtained a large expressed sequence tag data set containing more than 1,608 sequence fragments by 454 sequencing. Using three different methods, we conservatively define 21 preferred codons. Interestingly, the preferred codons in S. latifolia are almost identical to those in Arabidopsis thaliana, despite their long divergence time (we estimate average nonsynonymous site divergence to be 0.216, and synonymous sites are saturated). The agreement suggests that the nature of selection on codon usage has not changed significantly during the long evolutionary time separating the two species. As in many other organisms, the frequency of preferred codons is negatively correlated with protein length. For the 43 genes with both exon and intron sequences, we find a positive correlation between gene expression levels and GC content at third codon positions, but a strong negative correlation between expression and intron GC content, suggesting that the CUB we detect in S. latifolia is more likely to be due to natural selection than to mutational bias. Using polymorphism data, we detect evidence of ongoing natural selection on CUB, but we find little support for effects of biased gene conversion. An analysis of ten sex-linked genes reveals that the X chromosome has experienced significantly more unpreferred to preferred than preferred to unpreferred substitutions, suggesting that it may be evolving higher CUB. In contrast, numbers of substitutions between preferred and unpreferred codons are similar in both directions in the Y-linked genes, contrary to the expectation of genetic degeneration.

Boris Vyskot - One of the best experts on this subject based on the ideXlab platform.

  • agrobacterium rhizogenes mediated transformation of a dioecious plant model Silene latifolia
    New Biotechnology, 2019
    Co-Authors: Vojtech Hudzieczek, Roman Hobza, Tomas Cermak, Radim Cegan, Nela Bacovska, Zuzana Machalkova, Karel Dolezal, Lucie Plihalova, Daniel F Voytas, Boris Vyskot
    Abstract:

    Silene latifolia serves as a model species to study dioecy, the evolution of sex chromosomes, dosage compensation and sex-determination systems in plants. Currently, no protocol for genetic transformation is available for this species, mainly because S. latifolia is considered recalcitrant to in vitro regeneration and infection with Agrobacterium tumefaciens. Using cytokinins and their synthetic derivatives, we markedly improved the efficiency of regeneration. Several agrobacterial strains were tested for their ability to deliver DNA into S. latifolia tissues leading to transient and stable expression of the GUS reporter. The use of Agrobacterium rhizogenes strains resulted in the highest transformation efficiency (up to 4.7% of stable transformants) in hairy root cultures. Phenotypic and genotypic analyses of the T1 generation suggested that the majority of transformation events contain a small number of independent T-DNA insertions and the transgenes are transmitted to the progeny in a Mendelian pattern of inheritance. In short, we report an efficient and reproducible protocol for leaf disc transformation and subsequent plant regeneration in S. latifolia, based on the unique combination of infection with A. rhizogenes and plant regeneration from hairy root cultures using synthetic cytokinins. A protocol for the transient transformation of S.latifolia protoplasts was also developed and applied to demonstrate the possibility of targeted mutagenesis of the sex linked gene SlAP3 by TALENs and CRISPR/Cas9.

  • dna methylation and genetic degeneration of the y chromosome in the dioecious plant Silene latifolia
    BMC Genomics, 2018
    Co-Authors: Jose Luis Rodriguez Lorenzo, Roman Hobza, Boris Vyskot
    Abstract:

    S. latifolia is a model organism for the study of sex chromosome evolution in plants. Its sex chromosomes include large regions in which recombination became gradually suppressed. The regions tend to expand over time resulting in the formation of evolutionary strata. Non-recombination and later accumulation of repetitive sequences is a putative cause of the size increase in the Y chromosome. Gene decay and accumulation of repetitive DNA are identified as key evolutionary events. Transposons in the X and Y chromosomes are distributed differently and there is a regulation of transposon insertion by DNA methylation of the target sequences, this points to an important role of DNA methylation during sex chromosome evolution in Silene latifolia. The aim of this study was to elucidate whether the reduced expression of the Y allele in S. latifolia is caused by genetic degeneration or if the cause is methylation triggered by transposons and repetitive sequences. Gene expression analysis in S. latifolia males has shown expression bias in both X and Y alleles. To determine whether these differences are caused by genetic degeneration or methylation spread by transposons and repetitive sequences, we selected several sex-linked genes with varying degrees of degeneration and from different evolutionary strata. Immunoprecipitation of methylated DNA (MeDIP) from promoter, exon and intron regions was used and validated through bisulfite sequencing. We found DNA methylation in males, and only in the promoter of genes of stratum I (older). The Y alleles in genes of stratum I were methylation enriched compared to X alleles. There was also abundant and high percentage methylation in the CHH context in most sequences, indicating de novo methylation through the RdDM pathway. We speculate that TE accumulation and not gene decay is the cause of DNA methylation in the S. latifolia Y sex chromosome with influence on the process of heterochromatinization.

  • strong accumulation of chloroplast dna in the y chromosomes of rumex acetosa and Silene latifolia
    Cytogenetic and Genome Research, 2014
    Co-Authors: Pavlina Steflova, Boris Vyskot, Roman Hobza, Eduard Kejnovský
    Abstract:

    Chloroplast DNA (cpDNA) sequences are often found in plant nuclear genomes, but patterns of their chromosomal distribution are not fully understood. The distribution of cpDNA on the sex chromosomes can only be studied in dioecious plant species possessing heteromorphic sex chromosomes. We reconstructed the whole chloroplast genome of Rumex acetosa (sorrel, XY1Y2 system) from next generation sequencing data. We systematically mapped the chromosomal localization of various regions of cpDNA in R. acetosa and in Silene latifolia (white campion, XY system) using fluorescence in situ hybridization. We found that cpDNA was accumulated on the Y chromosomes of both studied species. In R. acetosa, the entire Y chromosome gathered all parts of cpDNA equally. On the contrary, in S. latifolia, the majority of the cpDNA, corresponding to the single copy regions, was localized in the centromere of the Y chromosome, while the inverted repeat region was present also in other loci. We found a stronger accumulation of cpDNA on the more degenerated Y1 and Y2 chromosomes of R. acetosa than in evolutionary younger S. latifolia Y chromosome. Our data stressed the prominent role of the Y chromosome centromere in cpDNA accumulation.

  • genomic diversity in two related plant species with and without sex chromosomes Silene latifolia and s vulgaris
    PLOS ONE, 2012
    Co-Authors: Boris Vyskot, Eduard Kejnovský, Jaroslav Doležel, Zdenek Kubat, Radim Cegan, Nicolas Blavet, Jan Safař, Hana Blavet, Roman Hobza
    Abstract:

    Background Genome size evolution is a complex process influenced by polyploidization, satellite DNA accumulation, and expansion of retroelements. How this process could be affected by different reproductive strategies is still poorly understood. Methodology/Principal Findings We analyzed differences in the number and distribution of major repetitive DNA elements in two closely related species, Silene latifolia and S. vulgaris. Both species are diploid and possess the same chromosome number (2n = 24), but differ in their genome size and mode of reproduction. The dioecious S. latifolia (1C = 2.70 pg DNA) possesses sex chromosomes and its genome is 2.5× larger than that of the gynodioecious S. vulgaris (1C = 1.13 pg DNA), which does not possess sex chromosomes. We discovered that the genome of S. latifolia is larger mainly due to the expansion of Ogre retrotransposons. Surprisingly, the centromeric STAR-C and TR1 tandem repeats were found to be more abundant in S. vulgaris, the species with the smaller genome. We further examined the distribution of major repetitive sequences in related species in the Caryophyllaceae family. The results of FISH (fluorescence in situ hybridization) on mitotic chromosomes with the Retand element indicate that large rearrangements occurred during the evolution of the Caryophyllaceae family. Conclusions/Significance Our data demonstrate that the evolution of genome size in the genus Silene is accompanied by the expansion of different repetitive elements with specific patterns in the dioecious species possessing the sex chromosomes.

  • next generation sequencing based analysis of repetitive dna in the model dioceous plant Silene latifolia
    PLOS ONE, 2011
    Co-Authors: Jiři Macas, Eduard Kejnovský, Andrea Koblizkova, Pavel Neumann, Petr Novak, Boris Vyskot
    Abstract:

    Background Silene latifolia is a dioceous plant with well distinguished X and Y chromosomes that is used as a model to study sex determination and sex chromosome evolution in plants. However, efficient utilization of this species has been hampered by the lack of large-scale sequencing resources and detailed analysis of its genome composition, especially with respect to repetitive DNA, which makes up the majority of the genome. Methodology/Principal Findings We performed low-pass 454 sequencing followed by similarity-based clustering of 454 reads in order to identify and characterize sequences of all major groups of S. latifolia repeats. Illumina sequencing data from male and female genomes were also generated and employed to quantify the genomic proportions of individual repeat families. The majority of identified repeats belonged to LTR-retrotransposons, constituting about 50% of genomic DNA, with Ty3/gypsy elements being more frequent than Ty1/copia. While there were differences between the male and female genome in the abundance of several repeat families, their overall repeat composition was highly similar. Specific localization patterns on sex chromosomes were found for several satellite repeats using in situ hybridization with probes based on k-mer frequency analysis of Illumina sequencing data. Conclusions/Significance This study provides comprehensive information about the sequence composition and abundance of repeats representing over 60% of the S. latifolia genome. The results revealed generally low divergence in repeat composition between the sex chromosomes, which is consistent with their relatively recent origin. In addition, the study generated various data resources that are available for future exploration of the S. latifolia genome.

Roman Hobza - One of the best experts on this subject based on the ideXlab platform.

  • the formation of sex chromosomes in Silene latifolia and s dioica was accompanied by multiple chromosomal rearrangements
    Frontiers in Plant Science, 2020
    Co-Authors: Václav Bačovský, Radim Cegan, Denisa Simonikova, Eva Hřibova, Roman Hobza
    Abstract:

    The genus Silene includes a plethora of dioecious and gynodioecious species. Two species, Silene latifolia (white campion) and Silene dioica (red campion), are dioecious plants, having heteromorphic sex chromosomes with an XX/XY sex determination system. The X and Y chromosomes differ mainly in size, DNA content and posttranslational histone modifications. Although it is generally assumed that the sex chromosomes evolved from a single pair of autosomes, it is difficult to distinguish the ancestral pair of chromosomes in related gynodioecious and hermaphroditic plants. We designed an oligo painting probe enriched for X-linked scaffolds from currently available genomic data and used this probe on metaphase chromosomes of S. latifolia (2n = 24, XY), S. dioica (2n = 24, XY), and two gynodioecious species, S. vulgaris (2n = 24) and S. maritima (2n = 24). The X chromosome-specific oligo probe produces a signal specifically on the X and Y chromosomes in S. latifolia and S. dioica, mainly in the subtelomeric regions. Surprisingly, in S. vulgaris and S. maritima, the probe hybridized to three pairs of autosomes labeling their p-arms. This distribution suggests that sex chromosome evolution was accompanied by extensive chromosomal rearrangements in studied dioecious plants.

  • The distribution of epigenetic histone marks differs between the X and Y chromosomes in Silene latifolia
    Planta, 2019
    Co-Authors: Václav Bačovský, Andreas Houben, Katrin Kumke, Roman Hobza
    Abstract:

    Main conclusionContrasting patterns of histone modifications between the X and Y chromosome in Silene latifolia show euchromatic histone mark depletion on the Y chromosome and indicate hyperactivation of one X chromosome in females.AbstractSilene latifolia (white campion) is a dioecious plant with heteromorphic sex chromosomes (24, XX in females and 24, XY in males), and a genetically degenerated Y chromosome that is 1.4 times larger than the X chromosome. Although the two sex chromosomes differ in their DNA content, information about epigenetic histone marks and evidence of their function are scarce. We performed immunolabeling experiments using antibodies specific for active and suppressive histone modifications as well as pericentromere-specific histone modifications. We show that the Y chromosome is partially depleted of histone modifications important for transcriptionally active chromatin, and carries these marks only in the pseudo-autosomal region, but that it is not enriched for suppressive and pericentromere histone marks. We also show that two of the active marks are specifically enriched in one of the X chromosomes in females and in the X chromosome in males. Our data support recent findings that genetic imprinting mediates dosage compensation of sex chromosomes in S. latifolia .

  • agrobacterium rhizogenes mediated transformation of a dioecious plant model Silene latifolia
    New Biotechnology, 2019
    Co-Authors: Vojtech Hudzieczek, Roman Hobza, Tomas Cermak, Radim Cegan, Nela Bacovska, Zuzana Machalkova, Karel Dolezal, Lucie Plihalova, Daniel F Voytas, Boris Vyskot
    Abstract:

    Silene latifolia serves as a model species to study dioecy, the evolution of sex chromosomes, dosage compensation and sex-determination systems in plants. Currently, no protocol for genetic transformation is available for this species, mainly because S. latifolia is considered recalcitrant to in vitro regeneration and infection with Agrobacterium tumefaciens. Using cytokinins and their synthetic derivatives, we markedly improved the efficiency of regeneration. Several agrobacterial strains were tested for their ability to deliver DNA into S. latifolia tissues leading to transient and stable expression of the GUS reporter. The use of Agrobacterium rhizogenes strains resulted in the highest transformation efficiency (up to 4.7% of stable transformants) in hairy root cultures. Phenotypic and genotypic analyses of the T1 generation suggested that the majority of transformation events contain a small number of independent T-DNA insertions and the transgenes are transmitted to the progeny in a Mendelian pattern of inheritance. In short, we report an efficient and reproducible protocol for leaf disc transformation and subsequent plant regeneration in S. latifolia, based on the unique combination of infection with A. rhizogenes and plant regeneration from hairy root cultures using synthetic cytokinins. A protocol for the transient transformation of S.latifolia protoplasts was also developed and applied to demonstrate the possibility of targeted mutagenesis of the sex linked gene SlAP3 by TALENs and CRISPR/Cas9.

  • dna methylation and genetic degeneration of the y chromosome in the dioecious plant Silene latifolia
    BMC Genomics, 2018
    Co-Authors: Jose Luis Rodriguez Lorenzo, Roman Hobza, Boris Vyskot
    Abstract:

    S. latifolia is a model organism for the study of sex chromosome evolution in plants. Its sex chromosomes include large regions in which recombination became gradually suppressed. The regions tend to expand over time resulting in the formation of evolutionary strata. Non-recombination and later accumulation of repetitive sequences is a putative cause of the size increase in the Y chromosome. Gene decay and accumulation of repetitive DNA are identified as key evolutionary events. Transposons in the X and Y chromosomes are distributed differently and there is a regulation of transposon insertion by DNA methylation of the target sequences, this points to an important role of DNA methylation during sex chromosome evolution in Silene latifolia. The aim of this study was to elucidate whether the reduced expression of the Y allele in S. latifolia is caused by genetic degeneration or if the cause is methylation triggered by transposons and repetitive sequences. Gene expression analysis in S. latifolia males has shown expression bias in both X and Y alleles. To determine whether these differences are caused by genetic degeneration or methylation spread by transposons and repetitive sequences, we selected several sex-linked genes with varying degrees of degeneration and from different evolutionary strata. Immunoprecipitation of methylated DNA (MeDIP) from promoter, exon and intron regions was used and validated through bisulfite sequencing. We found DNA methylation in males, and only in the promoter of genes of stratum I (older). The Y alleles in genes of stratum I were methylation enriched compared to X alleles. There was also abundant and high percentage methylation in the CHH context in most sequences, indicating de novo methylation through the RdDM pathway. We speculate that TE accumulation and not gene decay is the cause of DNA methylation in the S. latifolia Y sex chromosome with influence on the process of heterochromatinization.

  • strong accumulation of chloroplast dna in the y chromosomes of rumex acetosa and Silene latifolia
    Cytogenetic and Genome Research, 2014
    Co-Authors: Pavlina Steflova, Boris Vyskot, Roman Hobza, Eduard Kejnovský
    Abstract:

    Chloroplast DNA (cpDNA) sequences are often found in plant nuclear genomes, but patterns of their chromosomal distribution are not fully understood. The distribution of cpDNA on the sex chromosomes can only be studied in dioecious plant species possessing heteromorphic sex chromosomes. We reconstructed the whole chloroplast genome of Rumex acetosa (sorrel, XY1Y2 system) from next generation sequencing data. We systematically mapped the chromosomal localization of various regions of cpDNA in R. acetosa and in Silene latifolia (white campion, XY system) using fluorescence in situ hybridization. We found that cpDNA was accumulated on the Y chromosomes of both studied species. In R. acetosa, the entire Y chromosome gathered all parts of cpDNA equally. On the contrary, in S. latifolia, the majority of the cpDNA, corresponding to the single copy regions, was localized in the centromere of the Y chromosome, while the inverted repeat region was present also in other loci. We found a stronger accumulation of cpDNA on the more degenerated Y1 and Y2 chromosomes of R. acetosa than in evolutionary younger S. latifolia Y chromosome. Our data stressed the prominent role of the Y chromosome centromere in cpDNA accumulation.