Evolutionary rate variation at multiple levels of biological organization in plant mitochondrial DNA

We examined patterns of mitochondrial polymorphism and divergence in the angiosperm genus Silene and found substantial variation in evolutionary rates among species and among lineages within species. Moreover, we found corresponding differences in the amount of polymorphism within species. We argue that, along with our earlier findings of rate variation among genes, these patterns of rate heterogeneity at multiple phylogenetic scales are most likely explained by differences in underlying mutation rates. In contrast, no rate variation was detected in nuclear or chloroplast loci. We conclude that mutation rate heterogeneity is a characteristic of plant mitochondrial sequence evolution at multiple biological scales and may be a crucial determinant of how much polymorphism is maintained within species. These dramatic patterns of variation raise intriguing questions about the mechanisms driving and maintaining mutation rate heterogeneity in plant mitochondrial genomes. Additionally, they should alter our interpretation of many common phylogenetic and population genetic analyses.     

Isolation and characterization of highly polymorphic microsatellite loci in the bladder campion,Silene vulgaris (Caryophyllaceae)

This study reports the isolation and characterization of seven highly polymorphic microsatellite loci in Silene vulgaris (Caryophyllaceae). The loci were isolated from two libraries constructed from genomic DNA enriched for CA and GA repeats. These markers yielded nine to 40 alleles per locus (mean 22.1) in a survey of 45 individuals from a single population located in the western Swiss Alps. Average observed heterozygosity ranged from 16.2 to 77.4%. These microsatellite loci should be valuable tools for studying fine‐scale genetic structure.     

Toxicity of arsenate in Silene vulgaris, accumulation and degradation of arsenate-induced phytochelatins

The results presented in this paper describe the short- and long-term toxicity of arsenate in Silene vulgaris . Short-term toxicity, measured as inhibition of root elongation, depended on phosphate nutrition, arsenate being much less toxic at high phosphate supply. At low phosphate levels more arsenic was taken up by the plants. Under chronic exposure, toxicity (measured as inhibition of biomass production) did not increase with time. In addition, the accumulation of phytochelatins (PCs) as a function of toxicity and duration of exposure was studied. Short-term PC accumulation (over a 3 d period) was positively correlated with exposure. Isolation of peptide complexes from prolongedly exposed plants showed that PC₂, PC₃ and PC₄ were present, although the latter not until at least 3 d exposure. Arsenic co-eluted mainly with PC₂ and PC₃. Fractions containing PC₄ were devoid of As, probably due to dissociation of the complexes during extraction or elution. The breakdown of PCs after arresting As exposure was very slow. This could explain the continuous accumulation of PCs throughout longer periods of As exposure.     

Variation in mutation rate and polymorphism among mitochondrial genes of Silene vulgaris

The prevailing wisdom of the plant mitochondrial genome is that it has very low substitution rates, thus it is generally assumed that nucleotide diversity within species will also be low. However, recent evidence suggests plant mitochondrial genes may harbor variable and sometimes high levels of within-species polymorphism, a result attributed to variance in the influence of selection. However, insufficient attention has been paid to the effect of among-gene variation in mutation rate on varying levels of polymorphism across loci. We measured levels of polymorphism in seven mitochondrial gene regions across a geographically wide sample of the plant Silene vulgaris to investigate whether individual mitochondrial genes accumulate polymorphisms equally. We found that genes vary significantly in polymorphism. Tests based on coalescence theory show that the genes vary significantly in their scaled mutation rate, which, in the absence of differences among genes in effective population size, suggests these genes vary in their underlying mutation rate. Further evidence that among-gene variance in polymorphism is due to variation in the underlying mutation rate comes from a significant positive relationship between the number of segregating sites and silent site divergence from an outgroup. Contrary to recent studies, we found unconvincing evidence of recombination in the mitochondrial genome, and generally confirm the standard model of plant mitochondria characterized by low substitution rates and no recombination. We also show no evidence of significant variation in the strength or direction of selection among genes; this result may be expected if there is no recombination. The present study provides some of the most thorough data on plant mitochondrial polymorphism, and provides compelling evidence for mutation rate variation among genes. The study also demonstrates the difficulty in establishing a null model of mitochondrial genome polymorphism, and thus the difficulty, in the absence of a comparative approach, in testing the assumption that low substitution rates in plant mitochondria lead to low polymorphism.     

Heavy metal-induced accumulation of free proline in a metal-tolerant and a nontolerant ecotype of Silene vulgaris

Accumulation of free proline in response to Cu, Cd and Zn was studied in nontolerant and metal-tolerant Silene vulgaris (Moench) Garcke. In the nontolerant ecotype these metals induced a massive accumulation of proline, especially in the leaves. When compared at equimolar concentrations in the nutrient solution, Cu was the most effective inducer, followed by Cd and Zn, respectively. However, when compared at equal toxic strength, as estimated from the degree of root growth inhibition, proline accumulation decreased in the order Cd > Zn > Cu. The threshold exposure levels for proline accumulation coincided with the highest no-effect-concentrations for root growth. In the metal-tolerant ecotype the constitutive proline concentration in the leaves was 5 to 6 times higher than in the nontolerant ecotype. Exposure to Cu and Zn, however, was without any effect on the leaf proline concentration, even at exposure levels that caused a 50% root growth inhibition. Only Cd, when present at concentrations above the highest no-effect-concentration for root growth, induced a further increase of the leaf proline content. Reducing transpiration by placing the plants under a transparent polyethylene cover almost completely inhibited proline accumulation, even at metal accumulation rates in the leaves that caused a 10-fold increase of the proline level in leaves of uncovered plants. The results demonstrate that metal-induced proline accumulation depends on the development of a metal-induced water deficit in the leaves. Differential metal-induced proline accumulation in distinctly metal-tolerant ecotypes is a consequence, rather than a cause of differential metal tolerance.     

Proximity-dependent pollen performance in Silene vulgaris

BACKGROUND AND AIMS: Pollen and seed dispersal in herbaceous insect-pollinated plants are often restricted, inducing strong population structure. To what extent this influences mating within and among patches is poorly understood. This study investigates the influence of population structure on pollen performance using controlled pollinations and genetic markers. METHODS: Population structure was investigated in a patchily distributed population of gynodioecious Silene vulgaris in Switzerland using polymorphic microsatellite markers. Experimental pollinations were performed on 21 hermaphrodite recipients using pollen donors at three spatial scales: (a) self-pollination; (b) within-patch cross-pollinations; and (c) between-patch cross-pollinations. Pollen performance was then compared with respect to crossing distance. KEY RESULTS: The population of S. vulgaris was characterized by a high degree of genetic sub-structure, with neighbouring plants more related to one another than to distant individuals. Inbreeding probably results from both selfing and biparental inbreeding. Pollen performance increased with distance between mates. Between-patch pollen performed significantly better than both self- and within-patch pollen donors. However, no significant difference was detected between self- and within-patch pollen donors. CONCLUSIONS: The results suggest that population structure in animal-pollinated plants is likely to influence mating patterns by favouring cross-pollinations between unrelated plants. However, the extent to which this mechanism could be effective as a pre-zygotic barrier preventing inbred mating depends on the patterns of pollinator foraging and their influence on pollen dispersal.     

Variation in the intensity of inbreeding depression among successive life-cycle stages and generations in gynodioecious Silene vulgaris (Caryophyllaceae)

Inbreeding depression is one of the hypotheses explaining the maintenance of females within gynodioecious plant populations. However, the measurement of fitness components in selfed and outcrossed progeny depends on life-cycle stage and the history of inbreeding. Comparative data indicate that strong inbreeding depression is more likely to occur at later life-cycle stages. We used hermaphrodite individuals of Silene vulgaris originating from three populations located in different valleys in the Swiss Alps to investigate the effect of two generations of self- and cross-fertilization on fitness components among successive stages of the life cycle in a glasshouse experiment. We detected significant inbreeding depression for most life-cycle stages including: the number of viable and aborted seeds per fruit, probability of germination, above ground biomass, probability of flowering, number of flowers per plant, flower size and pollen viability. Overall, the intensity of inbreeding depression increased among successive stages of the life cycle and cumulative inbreeding depression was significantly stronger in the first generation (delta approximately 0.5) compared with the second generation (delta approximately 0.35). We found no evidence for synergistic epistasis in our experiment. Our finding of more intense inbreeding depression during later stages of the life cycle may help to explain the maintenance of females in gynodioecious populations of S. vulgaris because purging of genetic load is less likely to occur.     

IDENTICAL MAJOR GENE LOCI FOR HEAVY METAL TOLERANCES THAT HAVE INDEPENDENTLY EVOLVED IN DIFFERENT LOCAL POPULATIONS AND SUBSPECIES OF SILENE VULGARIS

Heavy metal tolerant Silene vulgaris plants, originating from different metalliferous sites in Germany and one in Ireland, were crossed to each other and to nontolerant plants from a nonmetalliferous site in The Netherlands. Analysis of the crosses suggested that there were two distinct major gene loci for zinc tolerance among a total of five tolerant populations. The tolerance loci for zinc, copper, and cadmium in the Irish plants were shown to be identical with those in the German populations. It is argued that the occurrence of common major genes for tolerance among different geographically isolated populations must have resulted from independent parallel evolution in local nontolerant ancestral populations. Each of the tolerances studied seems to be controlled by only a few specific major genes.     

Gender polymorphism in flowers of Silene vulgaris (Moench) Garcke (Caryophyllaceae)

Silene vulgaris occurs in Israel as a stolonifcrous tetraploid which forms mostly sexually polymorphic populations. Fully hermaphrodite and fully male-sterile plants are linked by a range of intermediate forms which are gynomonoecious and bear hermaphrodite, male-sterile, and partially male-sterile flowers. Male-sterile flowers differ from hermaphrodite ones by having a narrower calyx, a shorter corolla, a thicker style, and a considerably larger stigmatic area with longer papillae which are more markedly bulbous basally. In individual anthers of hermaphrodite flowers, diameters of stained pollen grains showed a continuous variation from 36 to 80 μm with a modal value of 56. About 30% of pollen was found to be non-stainable. Reduction in male fertility occurs at different levels, from small or inviable pollen grains through abortion of single anthers, abortion of entire androecia, and through different degrees of gynomonoecy to totally male-sterile individuals. The proportion of stainable pollen to ovules in hermaphrodite flowers alone was found to be less than 100. From the data presented, less than a third of all ovules produced in the different kinds of flower develop into seeds. This may reflect an inadequacy of pollen, which the local population of S. vulgaris can tolerate because of its stoloniferous habit.