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In flowering plants, pollinators contribute to gene flow while they also respond to variation in plant traits together determined by genetic, epigenetic and environmental sources of variation. Consequently, a correlation between abundance and diversity of pollinators and the genetic and epigenetic characteristics of plant populations such as diversity or distinctiveness is expected. However, no study has explored these long‐term dimensions of plant–pollinator interactions. Mediterranean narrow endemics often exhibit unexpectedly high levels of population genetic and epigenetic diversity. We hypothesize that pollinators may contribute to explain this pattern. Specifically, given the higher sensitivity of small, isolated population to gene flow, we expect a stronger association of pollinators with population genetic and epigenetic variability in narrow endemics than in widely distributed congeners. We studied five pairs of congeneric plant species, consisting of one narrow endemic with a restricted distribution and one widespread congener, found in the Sierra de Cazorla mountains (SE Spain). We characterized the pollinators in up to three populations per species to estimate their diversity and visitation rates. Additionally, we calculated the genetic and epigenetic diversity and distinctiveness of each population using AFLP markers and methylation‐sensitive AFLP markers (MSAP), respectively. We assessed the relationship between pollinator diversity and visitation rates. The diversity of pollinators did not vary according to the plant´s distribution range, but visitation rate was higher in widespread species. As predicted, only narrow endemics showed a significant association between pollinators and their population genetic and epigenetic characteristics. Specifically, higher pollinator diversity and visitation rates entailed higher population genetic diversity and lower epigenetic distinctiveness. This work shows the importance of investigating the relationship between pollinator diversity and population genetics and epigenetics to better understand the evolution of plant rarity. [ABSTRACT FROM AUTHOR]

Increasing evidence supports a major role for abiotic stress response in the success of plant polyploids, which usually thrive in harsh environments. However, understanding the ecophysiology of polyploids is challenging due to interactions between genome doubling and natural selection. Here, we investigated physiological responses, gene expression, and the epiphenotype of two related Dianthus broteri cytotypes—with different genome duplications (4× and 12×) and evolutionary trajectories—to short extreme temperature events (42/28 °C and 9/5 °C). The 12× cytotype showed higher expression of stress-responsive genes (SWEET1 , PP2C16 , AI5L3 , and ATHB7) and enhanced gas exchange compared with 4×. Under heat stress, both ploidies had greatly impaired physiological performance and altered gene expression, with reduced cytosine methylation. However, the 12× cytotype exhibited remarkable physiological tolerance (maintaining gas exchange and water status via greater photochemical integrity and probably enhanced water storage) while down-regulating PP2C16 expression. Conversely, 4× D. broteri was susceptible to thermal stress despite prioritizing water conservation, showing signs of non-stomatal photosynthetic limitations and irreversible photochemical damage. This cytotype also presented gene-specific expression patterns under heat, up-regulating ATHB7. These findings provide insights into divergent stress response strategies and physiological resistance resulting from polyploidy, highlighting its widespread influence on plant function. [ABSTRACT FROM AUTHOR]

DNA cytosine methylation is an epigenetic mechanism involved in regulation of plant responses to biotic and abiotic stress and its ability to change can vary with the sequence context in which a cytosine appears (CpG, CHG, CHH, where H = Adenine, Thymine, Cytosine). Quantification of DNA methylation in model plant species is frequently addressed by Whole Genome Bisulfite Sequencing (WGBS), which requires a good-quality reference genome. Reduced Representation Bisulfite Sequencing (RRBS) is a cost-effective potential alternative for ecological research with limited genomic resources and large experimental designs. In this study, we provide for the first time a comprehensive comparison between the outputs of RRBS and WGBS to characterize DNA methylation changes in response to a given environmental factor. In particular, we used epiGBS (recently optimized RRBS) and WGBS to assess global and sequence-specific differential methylation after insect and artificial herbivory in clones of Populus nigra cv. 'italica'. We found that, after any of the two herbivory treatments, global methylation percentage increased in CHH, and the shift was detected as statistically significant only by epiGBS. As regards to loci-specific differential methylation induced by herbivory (cytosines in epiGBS and regions in WGBS), both techniques indicated the specificity of the response elicited by insect and artificial herbivory, together with higher frequency of hypo-methylation in CpG and hyper-methylation in CHH. Methylation changes were mainly found in gene bodies and intergenic regions when present at CpG and CHG and in transposable elements and intergenic regions at CHH context. Thus, epiGBS succeeded to characterize global, genome-wide methylation changes in response to herbivory in the Lombardy poplar. Our results support that epiGBS could be particularly useful in large experimental designs aimed to explore epigenetic changes of non-model plant species in response to multiple environmental factors. [ABSTRACT FROM AUTHOR]

The long‐known, widely documented inverse relationship between body size and environmental temperature ("temperature‐size rule") has recently led to predictions of body size decline following current climatic warming ("size shrinking effect"). For keystone pollinators such as wild bees, body shrinking in response to warming can have significant effects on pollination processes but there is still little direct evidence of the phenomenon because adequate tests require controlling for confounding factors linked to climate change (e.g., habitat change). This paper assesses the shrinking effect in a community of solitary bees from well‐preserved habitats in the core of a large nature reserve experiencing climatic warming without disturbances or habitat changes. Long‐term variation in mean body mass was evaluated using data from 1704 individual bees (137 species, 27 genera, 6 families) sampled over 1990–2023. Climate warmed at a fast rate during this period, annual mean of daily maximum temperature increasing 0.069°C/year on average during 2000–2020. Changes in bee body mass verified expectations from the size shrinking effect. The mean individual body mass of the community of solitary bees declined significantly, irrespective of whether the analysis referred to the full species sample or only to the subset of species that were sampled in both the old (1990–1997) and recent (2022–2023) periods. On average, body mass declined ~0.7%·year−1, or an estimated average cumulative reduction of ~20 mg per individual bee from 1990 to 2023. Proportional size reduction was greatest among large‐bodied species, ranging from around −0.6%·year−1 for the smallest species to −0.9%·year−1 for the largest ones. Declining rate was steeper for cavity‐nesting than ground‐nesting species. The pollination and mating systems of bee‐pollinated plants in the study region are probably undergoing significant alterations as a consequence of supra‐annual decline in bee body mass. [ABSTRACT FROM AUTHOR]

Understanding the factors that drive community‐wide assembly of plant‐pollinator systems along environmental gradients has considerable evolutionary, ecological, and applied significance. Variation in thermal environments combined with intrinsic differences among pollinators in thermal biology have been proposed as drivers of community‐wide pollinator gradients, but this suggestion remains largely speculative. We test the hypothesis that seasonality in bee pollinator composition in Mediterranean montane habitats of southeastern Spain, which largely reflects the prevalence during the early flowering season of mining bees (Andrena), is a consequence of the latter's thermal biology. Quantitative information on seasonality of Andrena bees in the whole plant community (275 plant species) and their thermal microenvironment was combined with field and laboratory data on key aspects of the thermal biology of 30 species of Andrena (endothermic ability, warming constant, relationships of body temperature with ambient and operative temperatures). Andrena bees were a conspicuous, albeit strongly seasonal component of the pollinator assemblage of the regional plant community, visiting flowers of 153 different plant species (57% of total). The proportion of Andrena relative to all bees reached a maximum among plant species which flowered in late winter and early spring, and declined precipitously from May onward. Andrena were recorded only during the cooler segment of the annual range of air temperatures experienced at flowers by the whole bee assemblage. These patterns can be explained by features of Andrena's thermal biology: null to weak endothermy; ability to forage at much lower body temperature than strongly endothermic bees (difference ~ 10°C); low upper tolerable limit of body temperature, beyond which thermal stress presumably precluded foraging at the warmest period of year; weak thermoregulatory capacity; and high warming constant enhancing ectothermic warming. Our results demonstrate the importance of lineage‐specific pollinator traits as drivers of seasonality in community‐wide pollinator composition; show that exploitation of cooler microclimates by bees does not require strong endothermy; and suggest that intense endothermy and precise thermoregulation probably apply to a minority of bees. Medium‐ and large‐sized bees with low upper thermal limits and weak thermoregulatory ability can actually be more adversely affected by climate warming than large, hot‐blooded, extremely endothermic species. [ABSTRACT FROM AUTHOR]

Intraindividual epigenetic mosaicism is probably widespread among long‐lived plants, yet its ecological significance as a potential source of variation in fitness‐related traits in plant populations remains virtually unexplored. This paper examines the hypothesis that extant epigenetic variation within plants can have both current and transgenerational fecundity correlates which could eventually translate into fitness variations among different parts of the same individual and their respective offspring.Five modules, each consisting of a terminal branchlet bearing one inflorescence and its subtending leaves, were collected from each of 15 wild‐growing Lavandula latifolia (Lamiaceae) plants. They were characterized epigenotypically by the methylation state of methylation‐sensitive amplified fragment length polymorphism (MS‐AFLP) markers, and phenotypically by fecundity‐related traits (inflorescence length, and size, number and mass of seeds produced). Seeds from the different modules were sown in the greenhouse and resulting 'subprogenies' characterized phenotypically (germination probability and time to emergence, seedling size, susceptibility to fungal disease).All plants sampled were internally heterogeneous with regard to the methylation state of 1%–13% of MS‐AFLP markers. Predictable relationships were found between epigenotypic and phenotypic variation across the extant modules of individual L. latifolia shrubs. Phenotypes of subprogenies from different modules of the same plant grown under homogeneous conditions in the greenhouse were predictably related to the epigenotype of the maternal module which produced the seeds.Synthesis. The variable epigenotypes of different modules in the same plant not only predicted extant phenotypic variation among the modules themselves, but also phenotypic differences among the subprogenies produced by different modules. These relationships linking intraplant epigenotypic mosaicism with both extant and transgenerational heterogeneity in fitness‐related traits support the 'epigenetic mosaicism hypothesis' for plant variation, and also suggest hitherto unexplored ecological consequences of epigenotypically enhanced variation in the context of plant populations and communities. [ABSTRACT FROM AUTHOR]

Summary: Epigenetic mosaicism is a possible source of within‐plant phenotypic heterogeneity, yet its frequency and developmental origin remain unexplored. This study examines whether extant epigenetic heterogeneity within Lavandula latifolia (Lamiaceae) shrubs reflects recent epigenetic modifications experienced independently by different plant parts or, alternatively, it is the cumulative outcome of a steady lifetime process.Leaf samples from different architectural modules (branch tips) were collected from three L. latifolia plants and characterized epigenetically by global DNA cytosine methylation and methylation state of methylation‐sensitive amplified fragment‐length polymorphism (MS‐AFLP) markers. Epigenetic characteristics of modules were then assembled with information on the branching history of plants. Methods borrowed from phylogenetic research were used to assess genealogical signal of extant epigenetic variation and reconstruct within‐plant genealogical trajectory of epigenetic traits.Plants were epigenetically heterogeneous, as shown by differences among modules in global DNA methylation and variation in the methylation states of 6 to 8% of MS‐AFLP markers. All epigenetic features exhibited significant genealogical signal within plants. Events of epigenetic divergence occurred throughout the lifespan of individuals and were subsequently propagated by branch divisions.Internal epigenetic diversification of L. latifolia individuals took place steadily during their development, a process which eventually led to persistent epigenetic mosaicism. See also the Commentary on this article by Wetzel, 231: 1667–1669. [ABSTRACT FROM AUTHOR]

Phenotypic variation determines the capacity of plants to adapt to changing environments and to colonize new habitats. Deciphering the mechanisms contributing to plant phenotypic variation and their effects on plant ecological interactions and evolutionary dynamics is thus central to all biological disciplines. In the past few decades, research on plant epigenetics is showing that (1) epigenetic variation is related to phenotypic variation and that some epigenetic marks drive major phenotypic changes in plants; (2) plant epigenomes are highly diverse, dynamic, and can respond rapidly to a variety of biotic and abiotic stimuli; (3) epigenetic variation can respond to selection and therefore play a role in adaptive evolution. Yet, current information in terms of species, geographic ranges, and ecological contexts analyzed so far is too limited to allow for generalizations about the relevance of epigenetic regulation in phenotypic innovation and plant adaptation across taxa. In this report, we contextualize the potential role of the epigenome in plant adaptation to the environment and describe the latest research in this field presented during the symposium "Plant epigenetics: phenotypic and functional diversity beyond the DNA sequence" held within the Botany 2020 conference framework in summer 2020. [ABSTRACT FROM AUTHOR]

The ecological dynamics of co‐flowering communities are largely mediated by pollinators. However, current understanding of pollinator‐mediated interactions primarily relies on how co‐flowering plants influence attraction of shared pollinators, and much less is known about plant–plant interactions that occur via heterospecific pollen (HP) transfer. Invaded communities in particular can be highly affected by the transfer of alien pollen, but the strength, drivers and fitness consequences of these interactions at a community scale are not well understood.Here we analyse HP transfer networks in nine coastal communities in the Yucatan Mexico that vary in the relative abundance of invasive flowers to evaluate how HP donation and receipt varies between native and alien plants. We further evaluate whether HP donation and receipt are mediated by floral traits (e.g. display, flower size) or pollinator visitation rate. Finally, we evaluated whether post‐pollination success (proportion of pollen tubes produced) was affected by alien HP receipt and whether the effect varied between native and alien recipients.HP transfer networks exhibit relatively high connectance (c. 15%), suggesting high HP transfer within the studied communities. Significant network nestedness further suggests the existence of species that predominantly act as HP donors or recipients in the community. Species‐level analyses showed that natives receive 80% more HP compared to alien species, and that alien plants donate 40% more HP than natives. HP receipt and donation were mediated by different floral traits and such effects were independent of plant origin (native or alien). The proportion of alien HP received significantly affected conspecific pollen tube success in natives, but not that of alien species.Synthesis. Our results suggest that HP transfer in invaded communities is widespread, and that native and alien species play different roles within HP transfer networks, which are mediated by a different suite of floral traits. Alien species, in particular, play a central role as HP donors and are more tolerant to HP receipt than natives—a finding that points to two overlooked mechanisms facilitating alien plant invasion and success within native co‐flowering communities. [ABSTRACT FROM AUTHOR]

Background Pollen transfer via animals is necessary for reproduction by ~80 % of flowering plants, and most of these plants live in multispecies communities where they can share pollinators. While diffuse plant–pollinator interactions are increasingly recognized as the rule rather than the exception, their fitness consequences cannot be deduced from flower visitation alone, so other proxies, functionally closer to seed production and amenable for use in a broad variety of diverse communities, are necessary. Scope We conceptually summarize how the study of pollen on stigmas of spent flowers can reflect key drivers and functional aspects of the plant–pollinator interaction (e.g. competition, facilitation or commensalism). We critically evaluate how variable visitation rates and other factors (pollinator pool and floral avoidance) can give rise to different relationships between heterospecific pollen and (1) conspecific pollen on the stigma and (2) conspecific tubes/grain in the style, revealing the complexity of potential interpretations. We advise on best practices for using these proxies, noting the assumptions and caveats involved in their use, and explicate what additional data are required to verify interpretation of given patterns. Conclusions We conclude that characterizing pollen on stigmas of spent flowers provides an attainable indirect measure of pollination interactions, but given the complex processes of pollen transfer that generate patterns of conspecific–heterospecific pollen on stigmas these cannot alone determine whether competition or facilitation are the underlying drivers. Thus, functional tests are also needed to validate these hypotheses. [ABSTRACT FROM AUTHOR]

Genetic diversity defines the evolutionary potential of a species, yet mounting evidence suggests that epigenetic diversity could also contribute to adaptation. Elucidating the complex interplay between genetic and epigenetic variation in wild populations remains a challenge for evolutionary biologists, and the intriguing possibility that epigenetic diversity could compensate for the loss of genetic diversity is one aspect that remains basically unexplored in wild plants. This hypothesis is addressed in this paper by comparing the extent and patterns of genetic and epigenetic diversity of phylogenetically closely related but ecologically disparate species. Seven pairs of congeneric species from Cazorla mountains in south-eastern Spain were studied, each pair consisting of one endemic, restricted-range species associated to stressful environments, and one widespread species occupying more favourable habitats. The prediction was tested that endemic species should have lower genetic diversity due to population fragmentation, and higher epigenetic diversity induced by environmental stress, than their widespread congeners. Genetic (DNA sequence variants) and epigenetic (DNA cytosine methylation variants) diversities and their possible co-variation were assessed in three populations of each focal species using amplified fragment length polymorphism (AFLP) and methylation-sensitive AFLP (MSAP). All species and populations exhibited moderate to high levels of genetic polymorphism irrespective of their ecological characteristics. Epigenetic diversity was greater than genetic diversity in all cases. Only in endemic species were the two variables positively related, but the difference between epigenetic and genetic diversity was greater at populations with low genetic polymorphism. Results revealed that the relationship between genetic and epigenetic diversity can be more complex than envisaged by the simple hypothesis addressed in this study, and highlight the need of additional research on the actual role of epigenetic variation as a source of phenotypic diversity before a realistic understanding of the evolutionary relevance of epigenetic phenomena in plant adaptation can be achieved. [ABSTRACT FROM AUTHOR]

The interactions between pairs of native and alien plants via shared use of pollinators have been widely studied. Community level studies however, are necessary in order to fully understand the factors and mechanisms that facilitate successful plant invasion, but these are still scarce. Specifically, few community level studies have considered how differences in invasion level (alien flower abundance), and degree of floral trait similarity between native and invasive species, mediate effects on native plant-pollinator communities. Here, we evaluated the role of alien species on overall plant-floral visitor network structure, and on species-level network parameters, across nine invaded coastal communities distributed along 205 km in Yucatán, México that vary in alien species richness and flower abundance. We further assessed the potential the role of alien plant species on plant-floral visitor network structure and robustness via computational simulation of native and invasive plant extinction scenarios. We did not find significant differences between native and alien species in their functional floral phenotypes or in their visitation rate and pollinator community composition in these invaded sites. Variation in the proportion of alien plant species and flower abundance across sites did not influence plant-pollinator network structure. Species-level network parameters (i.e., normalized degree and nestedness contribution) did not differ between native and alien species. Furthermore, our simulation analyses revealed that alien species are functionally equivalent to native species and contribute equally to network structure and robustness. Overall, our results suggest that high levels of floral trait similarity and pollinator use overlap may help facilitate the integration of alien species into native plant-pollinator networks. As a result, alien species may also play a similar role than that of natives in the structure and stability of native plant and pollinator communities in the studied coastal sand dune ecosystem. [ABSTRACT FROM AUTHOR]

Premise: Phenotypic heterogeneity of reiterated, homologous structures produced by individual plants has ecological consequences for plants and their animal consumers. This paper examines experimentally the epigenetic mosaicism hypothesis, which postulates that within‐plant variation in traits of reiterated structures may partly arise from different parts of the same genetic individual differing in patterns or extent of genomic DNA methylation. Methods: Leaves of paired ramets borne by field‐growing Helleborus foetidus plants were infiltrated periodically over the entire flowering period with either a water solution of the demethylating agent zebularine or just water as the control. The effects of the zebularine treatment were assessed by quantifying genome‐wide DNA cytosine methylation in leaves and monitoring inflorescence growth and flower production, number of ovules per flower, pollination success, fruit set, seed set, seed size, and distribution of sap‐feeding insects. Results: Genomic DNA from leaves in zebularine‐treated ramets was significantly less methylated than DNA from leaves in control ones. Inflorescences in treated ramets grew smaller and produced fewer flowers, with fewer ovules and lower follicle and seed set, but did not differ from inflorescences in untreated ramets in pollination success or seed size. The zebularine treatment influenced the within‐plant distribution of sap‐feeding insects. Conclusions: Experimental manipulation of genomic DNA methylation level in leaves of wild‐growing H. foetidus plants induced considerable within‐plant heterogeneity in phenotypic (inflorescences, flowers, fecundity) and ecologically relevant traits (herbivore distribution), which supports the hypothesis that epigenetic mosaicism may partly account for within‐plant variation. [ABSTRACT FROM AUTHOR]

Growing evidence shows that epigenetic mechanisms contribute to complex traits, with implications across many fields of biology. In plant ecology, recent studies have attempted to merge ecological experiments with epigenetic analyses to elucidate the contribution of epigenetics to plant phenotypes, stress responses, adaptation to habitat, and range distributions. While there has been some progress in revealing the role of epigenetics in ecological processes, studies with non-model species have so far been limited to describing broad patterns based on anonymous markers of DNA methylation. In contrast, studies with model species have benefited from powerful genomic resources , which contribute to a more mechanistic understanding but have limited ecological realism. Understanding the significance of epigenetics for plant ecology requires increased transfer of knowledge and methods from model species research to genomes of evolutionarily divergent species, and examination of responses to complex natural environments at a more mechanistic level. This requires transforming genomics tools specifically for studying non-model species, which is challenging given the large and often polyploid genomes of plants. Collaboration among molecular geneticists, ecologists and bioinformaticians promises to enhance our understanding of the mutual links between genome function and ecological processes. [ABSTRACT FROM AUTHOR]

The way in which taxonomic diversity relates to functional diversity is important for understanding the mechanisms that sustain ecosystem function and services. We investigated how an explicit consideration of plant-pollinator interactions influences our view of plant diversity. We studied three plant species-rich communities located in different biodiversity hotspot regions: two soil-specific plant communities with Mediterranean-climate, sandy dolomite outcrops in Andalusia (Spain) and serpentine seeps in California (USA), and a third community in the sub-tropical dry scrublands in Yucatan (Mexico). Sampling at three spatial scales (region, site, plot) and rarefaction analyses were used to characterize and compare spatial and temporal variation of entomophilous plant diversity based on species presence (“static plant diversity”), flower display sizes along the season (“dynamic flower diversity”), and pollinator visitation (“interaction-effective diversity”). The studied communities differed in the static diversity of plants, with sub-tropical dry scrublands being less diverse than the two Mediterranean communities. Reduction of static diversity at local scale was stronger in the richest Mediterranean communities and, thus, static diversity was similar among regions when considering finer (site-level) spatial scales. In addition, the two Mediterranean communities displayed more seasonal variation, thus reducing differences in dynamic diversity among regions, i.e. when considering finer temporal scales. These results suggested that, at finer spatio-temporal scales expected to be relevant for interactions with pollinators, plant communities are not necessarily as diverse as the region where they occur. Accordingly, interaction-effective diversity based on pollinator visitation was in all cases lower than expected relative to the diversity of flowers. Thus, diversity of visited flowers does not perfectly track diversity of flowering species but instead reaches asymptotes at much lower values than expected in the richest communities. Regional species diversity may support the functionality of interactions at broader spatial and temporal scales than they actually occur. [ABSTRACT FROM AUTHOR]

Polyploidization is a significant evolutionary force in plants which involves major genomic and genetic changes, frequently regulated by epigenetic factors. We explored whether natural polyploidization in Dianthus broteri complex resulted in substantial changes in global DNA cytosine methylation associated to ploidy., Global cytosine methylation was estimated by high-performance liquid chromatography ( HPLC) in 12 monocytotypic populations with different ploidies (2×, 4×, 6×, 12×) broadly distributed within D. broteri distribution range. The effects of ploidy level and local variation on methylation were assessed by generalized linear mixed models (GLMMs)., Dianthus broteri exhibited a higher methylation percent (˜33%) than expected by its monoploid genome size and a large variation among study populations (range: 29.3-35.3%). Global methylation tended to increase with ploidy but did not significantly differ across levels due to increased variation within the highest-order polyploidy categories. Methylation varied more among hexaploid and dodecaploid populations, despite such cytotypes showing more restricted geographic location and increased genetic relatedness than diploids and tetraploids., In this study, we demonstrate the usefulness of an HPLC method in providing precise and genome reference-free global measure of DNA cytosine methylation, suitable to advance current knowledge of the roles of this epigenetic mechanism in polyploidization processes. [ABSTRACT FROM AUTHOR]

Methylation of DNA cytosines affects whether transposons are silenced and genes are expressed, and is a major epigenetic mechanism whereby plants respond to environmental change. Analyses of methylation-sensitive amplification polymorphism (MS-AFLP or MSAP) have been often used to assess methyl-cytosine changes in response to stress treatments and, more recently, in ecological studies of wild plant populations. MSAP technique does not require a sequenced reference genome and provides many anonymous loci randomly distributed over the genome for which the methylation status can be ascertained. Scoring of MSAP data, however, is not straightforward, and efforts are still required to standardize this step to make use of the potential to distinguish between methylation at different nucleotide contexts. Furthermore, it is not known how accurately MSAP infers genome-wide cytosine methylation levels in plants. Here, we analyse the relationship between MSAP results and the percentage of global cytosine methylation in genomic DNA obtained by HPLC analysis. A screening of literature revealed that methylation of cytosines at cleavage sites assayed by MSAP was greater than genome-wide estimates obtained by HPLC, and percentages of methylation at different nucleotide contexts varied within and across species. Concurrent HPLC and MSAP analyses of DNA from 200 individuals of the perennial herb Helleborus foetidus confirmed that methyl-cytosine was more frequent in CCGG contexts than in the genome as a whole. In this species, global methylation was unrelated to methylation at the inner CG site. We suggest that global HPLC and context-specific MSAP methylation estimates provide complementary information whose combination can improve our current understanding of methylation-based epigenetic processes in nonmodel plants. [ABSTRACT FROM AUTHOR]

DNA cytosine methylation is a widespread epigenetic mechanism in eukaryotes, and plant genomes commonly are densely methylated. Genomic methylation can be associated with functional consequences such as mutational events, genomic instability or altered gene expression, but little is known on interspecific variation in global cytosine methylation in plants. In this paper, we compare global cytosine methylation estimates obtained by HPLC and use a phylogenetically-informed analytical approach to test for significance of evolutionary signatures of this trait across 54 angiosperm species in 25 families. We evaluate whether interspecific variation in global cytosine methylation is statistically related to phylogenetic distance and also whether it is evolutionarily correlated with genome size (C-value). Global cytosine methylation varied widely between species, ranging between 5.3% (Arabidopsis) and 39.2% (Narcissus). Differences between species were related to their evolutionary trajectories, as denoted by the strong phylogenetic signal underlying interspecific variation. Global cytosine methylation and genome size were evolutionarily correlated, as revealed by the significant relationship between the corresponding phylogenetically independent contrasts. On average, a ten-fold increase in genome size entailed an increase of about 10% in global cytosine methylation. Results show that global cytosine methylation is an evolving trait in angiosperms whose evolutionary trajectory is significantly linked to changes in genome size, and suggest that the evolutionary implications of epigenetic mechanisms are likely to vary between plant lineages. [ABSTRACT FROM AUTHOR]