Biogeographic origins of southern African Silene (Caryophyllaceae).

Silene (Caryophyllaceae) is distributed predominantly in the northern Hemisphere, where it is most diverse around the Mediterranean Basin. The genus is also well represented in North Africa, extending into tropical, sub-Saharan and southern Africa. Eight native species are recognized in southern Africa, taxonomically placed in two sections: Elisanthe and Silene s.l. Although the taxonomy of the southern African taxa has recently been revised, their phylogenetic relationships and biogeographic history remain unclear. This study aims to infer the phylogenetic position and geographic origins of the southern African taxa. We generated DNA sequences of nuclear and plastid loci from several individuals belonging to all eight species of Silene recognized from southern Africa, and combined our DNA sequences with existing data representing species from major clades (i.e. sections) based on the recently revised Silene infrageneric taxonomy. We used a Bayesian coalescent species tree continuous diffusion approach to co-estimate the species tree and the ancestral areas of representative members of the genus. Our results show that the perennial southern African members of section Elisanthe form a strongly-supported clade with the Eurasian annual S. noctiflora and the Central Asian perennial S. turkestanica. The rest of the perennial species form a strongly-supported clade together with the annual S. aethiopica, which is nested in a larger Mediterranean clade comprising mostly annual species classified in section Silene s.l. Estimates of ancestral areas indicate a late Pleistocene dispersal to southern Africa from central and East Africa for the sub-Saharan members of section Silene s.l. The Elisanthe clade is inferred to have colonized southern Africa through long-distance dispersal from Eurasia during the late Pleistocene. Our findings support the hypothesis of a relatively recent colonization into southern Africa resulting from two independent dispersal events during the Pleistocene.

Congruent population genetic structures and divergence histories in anther-smut fungi and their host plants Silene italica and the Silene nutans species complex.

Study of the congruence of population genetic structure between hosts and pathogens gives important insights into their shared phylogeographical and coevolutionary histories. We studied the population genetic structure of castrating anther-smut fungi (genus Microbotryum) and of their host plants, the Silene nutans species complex, and the morphologically and genetically closely related Silene italica, which can be found in sympatry. Phylogeographical population genetic structure related to persistence in separate glacial refugia has been recently revealed in the S. nutans plant species complex across Western Europe, identifying several distinct lineages. We genotyped 171 associated plant-pathogen pairs of anther-smut fungi and their host plant individuals using microsatellite markers and plant chloroplastic single nucleotide polymorphisms. We found clear differentiation between fungal populations parasitizing S. nutans and S. italica plants. The population genetic structure of fungal strains parasitizing the S. nutans plant species complex mirrored the host plant genetic structure, suggesting that the pathogen was isolated in glacial refugia together with its host and/or that it has specialized on the plant genetic lineages. Using random forest approximate Bayesian computation (ABC-RF), we found that the divergence history of the fungal lineages on S. nutans was congruent with that previously inferred for the host plant and probably occurred with ancient but no recent gene flow. Genome sequences confirmed the genetic structure and the absence of recent gene flow between fungal genetic lineages. Our analyses of individual host-pathogen pairs contribute to a better understanding of co-evolutionary histories between hosts and pathogens in natural ecosystems, in which such studies remain scarce.

Recombination and intraspecific polymorphism for the presence and absence of entire chromosomes in mitochondrial genomes.

Although mitochondrial genomes are typically thought of as single circular molecules, these genomes are fragmented into multiple chromosomes in many eukaryotes, raising intriguing questions about inheritance and (in)stability of mtDNA in such systems. A previous comparison of mitochondrial genomes from two different individuals of the angiosperm species Silene noctiflora found variation in the presence of entire mitochondrial chromosomes. Here, we expand on this work with a geographically diverse sampling of 25 S. noctiflora populations and the closely related species S. turkestanica and S. undulata. Using a combination of deep sequencing and PCR-based screening for the presence of 22 different mitochondrial chromosomes, we found extensive variation in the complement of chromosomes across individuals. Much of this variation could be attributed to recent chromosome loss events, suggesting that the massively expanded and fragmented mitochondrial genomes of S. noctiflora may have entered a phase of genome reduction in which they are losing entire chromosomes at a rapid rate. Sequence analysis of mitochondrial and plastid genomes revealed genealogical differences both between these organelles and within the mitochondrial genome, indicating a history of recombination. Evidence that recombination has generated novel combinations of alleles was more frequent between loci on different mitochondrial chromosomes than it was within chromosomes. Therefore, the fragmentation of mitochondrial genomes and the assortment of chromosomes during mitochondrial inheritance appears to have contributed to a history of sexual-like recombination in the mtDNA of this species.

The two "rules of speciation" in species with young sex chromosomes.

The two "rules of speciation," Haldane's rule (HR) and the large-X effect (LXE), are thought to be caused by recessive species incompatibilities exposed in the phenotype due to the hemizygosity of X-linked genes in the heterogametic sex. Thus, the reports of HR and the LXE in species with recently evolved non- or partially degenerate Y-chromosomes, such as Silene latifolia and its relatives, were surprising. Here, I argue that rapid species-specific degeneration of Y-linked genes and associated adjustment of expression of X-linked gametologs (dosage compensation) may lead to rapid evolution of sex-linked species incompatibilities. This process is likely to be too slow in species with old degenerate Y-chromosomes (e.g., in mammals), but Y-degeneration in species with young gene-rich sex chromosomes may be fast enough to play a significant role in speciation. To illustrate this point, I report the analysis of Y-degeneration and the associated evolution of gene expression on the X-chromosome of S. latifolia and Silene dioica, a close relative that shares the same recently evolved sex chromosomes. Despite the recent (≤1MY) divergence of the two species, ~7% of Y-linked genes have undergone degeneration in one but not the other species. This species-specific degeneration appears to drive faster expression divergence of X-linked genes, which may account for HR and the LXE reported for these species. Furthermore, I suggest that "exposure" of autosomal or sex-linked recessive species incompatibilities in the haploid plant gametophyte may mimic the presence of HR in plants. Both haploid expression and species-specific Y-degeneration need to receive more attention if we are to understand the role of these processes in speciation.

The slowdown of Y chromosome expansion in dioecious Silene latifolia due to DNA loss and male-specific silencing of retrotransposons.

BACKGROUND: The rise and fall of the Y chromosome was demonstrated in animals but plants often possess the large evolutionarily young Y chromosome that is thought has expanded recently. Break-even points dividing expansion and shrinkage phase of plant Y chromosome evolution are still to be determined. To assess the size dynamics of the Y chromosome, we studied intraspecific genome size variation and genome composition of male and female individuals in a dioecious plant Silene latifolia, a well-established model for sex-chromosomes evolution. RESULTS: Our genome size data are the first to demonstrate that regardless of intraspecific genome size variation, Y chromosome has retained its size in S. latifolia. Bioinformatics study of genome composition showed that constancy of Y chromosome size was caused by Y chromosome DNA loss and the female-specific proliferation of recently active dominant retrotransposons. We show that several families of retrotransposons have contributed to genome size variation but not to Y chromosome size change. CONCLUSIONS: Our results suggest that the large Y chromosome of S. latifolia has slowed down or stopped its expansion. Female-specific proliferation of retrotransposons, enlarging the genome with exception of the Y chromosome, was probably caused by silencing of highly active retrotransposons in males and represents an adaptive mechanism to suppress degenerative processes in the haploid stage. Sex specific silencing of transposons might be widespread in plants but hidden in traditional hermaphroditic model plants.

Recombination changes at the boundaries of fully and partially sex-linked regions between closely related Silene species pairs.

The establishment of a region of suppressed recombination is a critical change during sex chromosome evolution, leading to such properties as Y (and W) chromosome genetic degeneration, accumulation of repetitive sequences and heteromorphism. Although chromosome inversions can cause large regions to have suppressed recombination, and inversions are sometimes involved in sex chromosome evolution, gradual expansion of the non-recombining region could potentially sometimes occur. We here test whether closer linkage has recently evolved between the sex-determining region and several genes that are partially sex-linked in Silene latifolia, using Silene dioica, a closely related dioecious plants whose XY sex chromosome system is inherited from a common ancestor. The S. latifolia pseudoautosomal region (PAR) includes several genes extremely closely linked to the fully Y-linked region. These genes were added to an ancestral PAR of the sex chromosome pair in two distinct events probably involving translocations of autosomal genome regions causing multiple genes to become partially sex-linked. Close linkage with the PAR boundary must have evolved since these additions, because some genes added in both events now show almost complete sex linkage in S. latifolia. We compared diversity patterns of five such S. latifolia PAR boundary genes with their orthologues in S. dioica, including all three regions of the PAR (one gene that was in the ancestral PAR and two from each of the added regions). The results suggest recent recombination suppression in S. latifolia, since its split from S. dioica.

Positive Selection in Rapidly Evolving Plastid-Nuclear Enzyme Complexes.

Rates of sequence evolution in plastid genomes are generally low, but numerous angiosperm lineages exhibit accelerated evolutionary rates in similar subsets of plastid genes. These genes include clpP1 and accD, which encode components of the caseinolytic protease (CLP) and acetyl-coA carboxylase (ACCase) complexes, respectively. Whether these extreme and repeated accelerations in rates of plastid genome evolution result from adaptive change in proteins (i.e., positive selection) or simply a loss of functional constraint (i.e., relaxed purifying selection) is a source of ongoing controversy. To address this, we have taken advantage of the multiple independent accelerations that have occurred within the genus Silene (Caryophyllaceae) by examining phylogenetic and population genetic variation in the nuclear genes that encode subunits of the CLP and ACCase complexes. We found that, in species with accelerated plastid genome evolution, the nuclear-encoded subunits in the CLP and ACCase complexes are also evolving rapidly, especially those involved in direct physical interactions with plastid-encoded proteins. A massive excess of nonsynonymous substitutions between species relative to levels of intraspecific polymorphism indicated a history of strong positive selection (particularly in CLP genes). Interestingly, however, some species are likely undergoing loss of the native (heteromeric) plastid ACCase and putative functional replacement by a duplicated cytosolic (homomeric) ACCase. Overall, the patterns of molecular evolution in these plastid-nuclear complexes are unusual for anciently conserved enzymes. They instead resemble cases of antagonistic coevolution between pathogens and host immune genes. We discuss a possible role of plastid-nuclear conflict as a novel cause of accelerated evolution.

Species delimitation without prior knowledge: DISSECT reveals extensive cryptic speciation in the Silene aegyptiaca complex (Caryophyllaceae).

Species delimitation is a major focus of biosystematics. In recent years, considerable progress has been achieved with the development of the multispecies coalescent (MSC) model, where species constitute the branches of the species tree or network. However, researchers are faced with the limitation that the MSC method of choice often requires a priori assignment of individuals to species. This not only introduces subjectivitiy into the analyses, but may also lead to meaningless species tree hypotheses, if the allele-to-species assignments are inaccurate. DISSECT is a recently introduced method that does not require a priori allele-to-species assignments, but instead examines the posterior probabilities of groupings (clusterings) of individuals under study. Using the DISSECT approach, we analysed genetic data from 75 individual plants belonging to the Silene aegyptiaca species complex that has previously been divided into 3-5 species. Marginal likelihood estimates from (*)BEAST analyses, run with predefined species classifications, strongly favour those compatible with the DISSECT result over those from morphology- and geography-based taxonomy. We found at least nine species, including several cryptic ones, for which no clear geographical or morphological patterns are correlated. However, the limited data and the possibility of unmodelled processes mean there is still much uncertainty about the true number of MSC species, and for taxonomic purposes, other criteria might be relevant. Nevertheless, we argue that the approach signifies an important step towards objective and testable species delimitations in any organismal group. In particular, it makes it possible to avoid biologically irrelevant species classifications.

RAD mapping reveals an evolving, polymorphic and fuzzy boundary of a plant pseudoautosomal region.

How loss of genetic exchanges (recombination) evolves between sex chromosomes is a long-standing question. Suppressed recombination may evolve when a sexually antagonistic (SA) polymorphism occurs in a partially sex-linked 'pseudoautosomal' region (or 'PAR'), maintaining allele frequency differences between the two sexes, and creating selection for closer linkage with the fully sex-linked region of the Y chromosome in XY systems, or the W in ZW sex chromosome systems. Most evidence consistent with the SA polymorphism hypothesis is currently indirect, and more studies of the genetics and population genetics of PAR genes are clearly needed. The sex chromosomes of the plant Silene latifolia are suitable for such studies, as they evolved recently and the loss of recombination could still be ongoing. Here, we used RAD sequencing to genetically map sequences in this plant, which has a large genome (c. 3 gigabases) and no available whole-genome sequence. We mapped 83 genes on the sex chromosomes, and comparative mapping in the related species S. vulgaris supports previous evidence for additions to an ancestral PAR and identified at least 12 PAR genes. We describe evidence that recombination rates have been reduced in meiosis of both sexes, and differences in recombination between S. latifolia families suggest ongoing recombination suppression. Large allele frequency differences between the sexes were found at several loci closely linked to the PAR boundary, and genes in different regions of the PAR showed striking sequence diversity patterns that help illuminate the evolution of the PAR.

Patterns of molecular evolution in dioecious and non-dioecious Silene.

Dioecy (i.e. having separate sexes) is a rather rare breeding system in flowering plants. Such rareness may result from a high probability of extinction in dioecious species because of less efficient dispersal and the costs of sexual selection, which are expected to harm dioecious species' survival on the long term. These handicaps should decrease the effective population size (Ne) of dioecious species, which in turn should reduce the efficacy of selection. Moreover, sexual selection in dioecious species is expected to specifically affect some genes, which will evolve under positive selection. The relative contribution of these effects is currently unknown and we tried to disentangle them by comparing sequence evolution between dioecious and non-dioecious species in the Silene genus (Caryophyllaceae), where dioecy has evolved at least twice. For the dioecious species in the section Melandrium, where dioecy is the oldest, we found a global reduction of purifying selection, while on some, male-biased genes, positive selection was found. For section Otites, where dioecy evolved more recently, we found no significant differences between dioecious and non-dioecious species. Our results are consistent with the view that dioecy is an evolutionary dead end in flowering plants, although other scenarios for explaining reduced Ne cannot be ruled out. Our results also show that contrasting forces act on the genomes of dioecious plants, and suggest that some time is required before the genome of such plants bears the footprints of dioecy.

SlWUS1; an X-linked gene having no homologous Y-linked copy in Silene latifolia.

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.

Genomic diversity in two related plant species with and without sex chromosomes--Silene latifolia and S. vulgaris.

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.

De novo transcriptome assembly and polymorphism detection in the flowering plant Silene vulgaris (Caryophyllaceae).

Members of the angiosperm genus Silene are widely used in studies of ecology and evolution, but available genomic and population genetic resources within Silene remain limited. Deep transcriptome (i.e. expressed sequence tag or EST) sequencing has proven to be a rapid and cost-effective means to characterize gene content and identify polymorphic markers in non-model organisms. In this study, we report the results of 454 GS-FLX Titanium sequencing of a polyA-selected and normalized cDNA library from Silene vulgaris. The library was generated from a single pool of transcripts, combining RNA from leaf, root and floral tissue from three genetically divergent European subpopulations of S. vulgaris. A single full-plate 454 run produced 959,520 reads totalling 363.6 Mb of sequence data with an average read length of 379.0 bp after quality trimming and removal of custom library adaptors. We assembled 832,251 (86.7%) of these reads into 40,964 contigs, which have a total length of 25.4 Mb and can be organized into 18,178 graph-based clusters or 'isogroups'. Assembled sequences were annotated based on homology to genes in multiple public databases. Analysis of sequence variants identified 13,432 putative single-nucleotide polymorphisms (SNPs) and 1320 simple sequence repeats (SSRs) that are candidates for microsatellite analysis. Estimates of nucleotide diversity from 1577 contigs were used to generate genome-wide distributions that revealed several outliers with high diversity. All of these resources are publicly available through NCBI and/or our website (http://silenegenomics.biology.virginia.edu) and should provide valuable genomic and population genetic tools for the Silene research community.

Geographic and phylogenetic patterns in Silene section Melandrium (Caryophyllaceae) as inferred from chloroplast and nuclear DNA sequences.

The phylogenetic relationships between the five dioecious species in Silene section Melandrium (Caryophyllaceae) and their putative hermaphrodite relatives are investigated based on an extensive geographic and taxonomic sample, using DNA sequence data from the chloroplast genome and the nuclear ribosomal ITS region. The hermaphrodite S. noctiflora (the type species of section Elisanthe) is distantly related to the dioecious species. With the exception of chloroplast sequences in one S. latifolia population from Turkey, the dioecious taxa form a strongly supported monophyletic group (Silene section Melandrium). The phylogenetic structure within section Melandrium differs between chloroplast and nuclear sequences. While there is extensive sharing of chloroplast haplotypes among all the dioecious species (the observed patterns reflect geographic structure), the nuclear ITS phylogeny shows a higher degree of taxonomic structure. Chloroplast-sharing by the section Melandrium species is most plausibly explained by a history of hybridization and extensive backcrossing.

Silene latifolia: the classical model to study heteromorphic sex chromosomes.

This review summarizes older as well as recent data about the model dioecious plant Silene latifolia. This plant has been the subject of more than one hundred years of research efforts and its most conspicuous property is huge and well differentiated heteromorphic sex chromosomes, XX in females and XY in males. Due to this property the S. latifolia sex chromosomes have been often used for cytogenetic studies as well as for flow sorting and laser microdissection. Nowadays S. latifolia is the focus of genomic studies, molecular mapping, phylogenetic and population genetics analyses.

Extensive loss of RNA editing sites in rapidly evolving Silene mitochondrial genomes: selection vs. retroprocessing as the driving force.

Theoretical arguments suggest that mutation rates influence the proliferation and maintenance of RNA editing. We identified RNA editing sites in five species within the angiosperm genus Silene that exhibit highly divergent mitochondrial mutation rates. We found that mutational acceleration has been associated with rapid loss of mitochondrial editing sites. In contrast, we did not find a significant difference in the frequency of editing in chloroplast genes, which lack the mutation rate variation observed in the mitochondrial genome. As found in other angiosperms, the rate of substitution at RNA editing sites in Silene greatly exceeds the rate at synonymous sites, a pattern that has previously been interpreted as evidence for selection against RNA editing. Alternatively, we suggest that editing sites may experience higher rates of C-to-T mutation than other portions of the genome. Such a pattern could be caused by gene conversion with reverse-transcribed mRNA (i.e., retroprocessing). If so, the genomic distribution of RNA editing site losses in Silene suggests that such conversions must be occurring at a local scale such that only one or two editing sites are affected at a time. Because preferential substitution at editing sites appears to occur in angiosperms regardless of the mutation rate, we conclude that mitochondrial rate accelerations within Silene have "fast-forwarded" a preexisting pattern but have not fundamentally changed the evolutionary forces acting on RNA editing sites.

Tolerance of Silene vulgaris to copper: population-related comparison of selected physiological parameters.

Tolerance of three Slovak populations of Silene vulgaris [from the localities "Harmanec" (total soil Cu 34.1 mg kg⁻¹), "Lubietová"; (total soil Cu 831.6 mg kg⁻¹) and "Spania dolina" (total soil Cu 1368.7 mg kg⁻¹) referred as SV1, SV2, and SV3, respectively] and Silene dioica (SD; from the locality "Spania dolina") exposed to 60 µM Cu for 7 days has been studied. Respective controls with 0.21 µM Cu were also cultured. SV3 and SD contained higher total shoot Cu than SV1. Methanol-soluble shoot Cu represented 60, 59, 59, and 55% from total shoot Cu and roots contained 16.7-, 20.4-, 9.9- and 28.9-fold more Cu than shoots in SV1, SV2, SV3, and SD, respectively. Soluble proteins were the least affected in SV3 and SD when Cu-exposed and control plants were compared. Root hydrogen peroxide and superoxide showed similar trend and were correlated with enhancement of ascorbate- and guaiacol-peroxidase activities. Malondialdehyde accumulation increased in SV1 and SV2. Phenylalanine ammonia-lyase activity and total soluble phenols were higher in SV3 Cu-exposed plants compared to SV1 and SV2. Shikimate dehydrogenase activity was enhanced in all Cu-exposed populations. Cinnamyl alcohol dehydrogenase activity and root lignin content were not affected and polyphenol oxidase activity was not detected. Within 14 detected free amino acids, majority of them decreased preferentially in the roots. Only serine increased in shoots and decreased in roots of all populations. Potassium content was not affected in SV3. Na, Ca, and Fe increased in roots of Cu-exposed plants of all populations, while Mg was the least affected and Zn increased in SV2 and SV3 shoots and roots. This study revealed the highest tolerance in population from the locality with the highest soil Cu content. Present findings as complex metabolic responses to Cu stress with special emphasis on phenolic metabolism are discussed.

Silene patula (Siphonomorpha, Caryophyllaceae) in North Africa: a test of colonisation routes using chloroplast markers.

Based on morphological characters, the North African Silene patula has been divided into two subspecies, ssp. patula found North of Kabylies and Atlas Mountains, and ssp. amurensis found south of these regions. In order to test the hypothesis that S. patula could have derived from S. italica through the Sicilian Channel during the Messinian, we sequenced three chloroplast loci, trnH-psbA, trnS-trnG and rpl12-rps20. Fifteen haplotypes were found on 211 herbarium specimens, associated with a huge differentiation within species. The hypothesis that S. patula had independently evolved as two different subspecies North and South of the mountains is refuted and a morphological adaptation to different pollinators is suggested to explain the differences between the two taxa. The Kabylies-Numidie-Kroumirie gathers a large proportion of haplotypes, which points to this region as a probable refugium or place of origin from which spatial expansions have subsequently occurred towards Morocco and the Aurès Mountains.

Conflicting phylogenetic signals in the SlX1/Y1 gene in Silene.

BACKGROUND: Increasing evidence from DNA sequence data has revealed that phylogenies based on different genes may drastically differ from each other. This may be due to either inter- or intralineage processes, or to methodological or stochastic errors. Here we investigate a spectacular case where two parts of the same gene (SlX1/Y1) show conflicting phylogenies within Silene (Caryophyllaceae). SlX1 and SlY1 are sex-linked genes on the sex chromosomes of dioecious members of Silene sect. Elisanthe. RESULTS: We sequenced the homologues of the SlX1/Y1 genes in several Sileneae species. We demonstrate that different parts of the SlX1/Y1 region give different phylogenetic signals. The major discrepancy is that Silene vulgaris and S. sect. Conoimorpha (S. conica and relatives) exchange positions. To determine whether gene duplication followed by recombination (an intralineage process) may explain the phylogenetic conflict in the Silene SlX1/Y1 gene, we use a novel probabilistic, multiple primer-pair PCR approach. We did not find any evidence supporting gene duplication/loss as explanation to the phylogenetic conflict. CONCLUSION: The phylogenetic conflict in the Silene SlX1/Y1 gene cannot be explained by paralogy or artefacts, such as in vitro recombination during PCR. The support for the conflict is strong enough to exclude methodological or stochastic errors as likely sources. Instead, the phylogenetic incongruence may have been caused by recombination of two divergent alleles following ancient interspecific hybridization or incomplete lineage sorting. These events probably took place several million years ago. This example clearly demonstrates that different parts of the genome may have different evolutionary histories and stresses the importance of using multiple genes in reconstruction of taxonomic relationships.

Survey of repetitive sequences in Silene latifolia with respect to their distribution on sex chromosomes.

We carried out a global survey of all major types of transposable elements in Silene latifolia, a model species with sex chromosomes that are in the early stages of their evolution. A shotgun genomic library was screened with genomic DNA to isolate and characterize the most abundant elements. We found that the most common types of elements were the subtelomeric tandem repeat X-43.1 and Gypsy retrotransposons, followed by Copia retrotransposons and LINE non-LTR elements. SINE elements and DNA transposons were less abundant. We also amplified transposable elements with degenerate primers and used them to screen the library. The localization of elements by FISH revealed that most of the Copia elements were accumulated on the Y chromosome. Surprisingly, one type of Gypsy element, which was similar to Ogre elements known from legumes, was almost absent on the Y chromosome but otherwise uniformly distributed on all chromosomes. Other types of elements were ubiquitous on all chromosomes. Moreover, we isolated and characterized two new tandem repeats. One of them, STAR-C, was localized at the centromeres of all chromosomes except the Y chromosome, where it was present on the p-arm. Its variant, STAR-Y, carrying a small deletion, was specifically localized on the q-arm of the Y chromosome. The second tandem repeat, TR1, co-localized with the 45S rDNA cluster in the subtelomeres of five pairs of autosomes. FISH analysis of other Silene species revealed that some elements (e.g., Ogre-like elements) are confined to the section Elisanthe while others (e.g. Copia or Athila-like elements) are present also in more distant species. Similarly, the centromeric satellite STAR-C was conserved in the genus Silene whereas the subtelomeric satellite X-43.1 was specific for Elisanthe section. Altogether, our data provide an overview of the repetitive sequences in Silene latifolia and revealed that genomic distribution and evolutionary dynamics differ among various repetitive elements. The unique pattern of repeat distribution is found on the Y chromosome, where some elements are accumulated while other elements are conspicuously absent, which probably reflects different forces shaping the Y chromosome.