Lisa Pokorny

Phylotranscriptomic analysis of the origin and early diversification of land plants

Significance Early branching events in the diversification of land plants and closely related algal lineages remain fundamental and unresolved questions in plant evolutionary biology. Accurate reconstructions of these relationships are critical for testing hypotheses of character evolution: for example, the origins of the embryo, vascular tissue, seeds, and flowers. We investigated relationships among streptophyte algae and land plants using the largest set of nuclear genes that has been applied to this problem to date. Hypothesized relationships were rigorously tested through a series of analyses to assess systematic errors in phylogenetic inference caused by sampling artifacts and model misspecification. Results support some generally accepted phylogenetic hypotheses, while rejecting others. This work provides a new framework for studies of land plant evolution. Reconstructing the origin and evolution of land plants and their algal relatives is a fundamental problem in plant phylogenetics, and is essential for understanding how critical adaptations arose, including the embryo, vascular tissue, seeds, and flowers. Despite advances in molecular systematics, some hypotheses of relationships remain weakly resolved. Inferring deep phylogenies with bouts of rapid diversification can be problematic; however, genome-scale data should significantly increase the number of informative characters for analyses. Recent phylogenomic reconstructions focused on the major divergences of plants have resulted in promising but inconsistent results. One limitation is sparse taxon sampling, likely resulting from the difficulty and cost of data generation. To address this limitation, transcriptome data for 92 streptophyte taxa were generated and analyzed along with 11 published plant genome sequences. Phylogenetic reconstructions were conducted using up to 852 nuclear genes and 1,701,170 aligned sites. Sixty-nine analyses were performed to test the robustness of phylogenetic inferences to permutations of the data matrix or to phylogenetic method, including supermatrix, supertree, and coalescent-based approaches, maximum-likelihood and Bayesian methods, partitioned and unpartitioned analyses, and amino acid versus DNA alignments. Among other results, we find robust support for a sister-group relationship between land plants and one group of streptophyte green algae, the Zygnematophyceae. Strong and robust support for a clade comprising liverworts and mosses is inconsistent with a widely accepted view of early land plant evolution, and suggests that phylogenetic hypotheses used to understand the evolution of fundamental plant traits should be reevaluated.

Data access for the 1,000 Plants (1KP) project

The 1,000 plants (1KP) project is an international multi-disciplinary consortium that has generated transcriptome data from over 1,000 plant species, with exemplars for all of the major lineages across the Viridiplantae (green plants) clade. Here, we describe how to access the data used in a phylogenomics analysis of the first 85 species, and how to visualize our gene and species trees. Users can develop computational pipelines to analyse these data, in conjunction with data of their own that they can upload. Computationally estimated protein-protein interactions and biochemical pathways can be visualized at another site. Finally, we comment on our future plans and how they fit within this scalable system for the dissemination, visualization, and analysis of large multi-species data sets.

Algal ancestor of land plants was preadapted for symbiosis

Significance Colonization of land by plants was a critical event for the emergence of extant ecosystems. The innovations that allowed the algal ancestor of land plants to succeed in such a transition remain unknown. Beneficial interaction with symbiotic fungi has been proposed as one of these innovations. Here we show that the genes required for this interaction appeared in a stepwise manner: Some evolved before the colonization of land by plants and others first appeared in land plants. We thus propose that the algal ancestor of land plants was preadapted for interaction with beneficial fungi and employed these gene networks to colonize land successfully. Colonization of land by plants was a major transition on Earth, but the developmental and genetic innovations required for this transition remain unknown. Physiological studies and the fossil record strongly suggest that the ability of the first land plants to form symbiotic associations with beneficial fungi was one of these critical innovations. In angiosperms, genes required for the perception and transduction of diffusible fungal signals for root colonization and for nutrient exchange have been characterized. However, the origin of these genes and their potential correlation with land colonization remain elusive. A comprehensive phylogenetic analysis of 259 transcriptomes and 10 green algal and basal land plant genomes, coupled with the characterization of the evolutionary path leading to the appearance of a key regulator, a calcium- and calmodulin-dependent protein kinase, showed that the symbiotic signaling pathway predated the first land plants. In contrast, downstream genes required for root colonization and their specific expression pattern probably appeared subsequent to the colonization of land. We conclude that the most recent common ancestor of extant land plants and green algae was preadapted for symbiotic associations. Subsequent improvement of this precursor stage in early land plants through rounds of gene duplication led to the acquisition of additional pathways and the ability to form a fully functional arbuscular mycorrhizal symbiosis.

Horizontal transfer of an adaptive chimeric photoreceptor from bryophytes to ferns

Significance Despite being one of the oldest groups of land plants, the majority of living ferns resulted from a relatively recent diversification following the rise of angiosperms. To exploit fully the new habitats created by angiosperm-dominated ecosystems, ferns had to evolve novel adaptive strategies to cope with the low-light conditions exerted by the angiosperm canopy. Neochrome, an unconventional photoreceptor that allows ferns to “see the light” better, was likely part of the solution. Surprisingly, we discovered that fern neochrome was derived from a bryophyte lineage via horizontal gene transfer (HGT). This finding not only provides the first evidence that a plant-to-plant HGT can have a profound evolutionary impact but also has implications for the evolution of photosensory systems in plants. Ferns are well known for their shade-dwelling habits. Their ability to thrive under low-light conditions has been linked to the evolution of a novel chimeric photoreceptor—neochrome—that fuses red-sensing phytochrome and blue-sensing phototropin modules into a single gene, thereby optimizing phototropic responses. Despite being implicated in facilitating the diversification of modern ferns, the origin of neochrome has remained a mystery. We present evidence for neochrome in hornworts (a bryophyte lineage) and demonstrate that ferns acquired neochrome from hornworts via horizontal gene transfer (HGT). Fern neochromes are nested within hornwort neochromes in our large-scale phylogenetic reconstructions of phototropin and phytochrome gene families. Divergence date estimates further support the HGT hypothesis, with fern and hornwort neochromes diverging 179 Mya, long after the split between the two plant lineages (at least 400 Mya). By analyzing the draft genome of the hornwort Anthoceros punctatus, we also discovered a previously unidentified phototropin gene that likely represents the ancestral lineage of the neochrome phototropin module. Thus, a neochrome originating in hornworts was transferred horizontally to ferns, where it may have played a significant role in the diversification of modern ferns.

The evolutionary history of ferns inferred from 25 low-copy nuclear genes

UNLABELLED • PREMISE OF THE STUDY Understanding fern (monilophyte) phylogeny and its evolutionary timescale is critical for broad investigations of the evolution of land plants, and for providing the point of comparison necessary for studying the evolution of the fern sister group, seed plants. Molecular phylogenetic investigations have revolutionized our understanding of fern phylogeny, however, to date, these studies have relied almost exclusively on plastid data.• METHODS Here we take a curated phylogenomics approach to infer the first broad fern phylogeny from multiple nuclear loci, by combining broad taxon sampling (73 ferns and 12 outgroup species) with focused character sampling (25 loci comprising 35877 bp), along with rigorous alignment, orthology inference and model selection.• KEY RESULTS Our phylogeny corroborates some earlier inferences and provides novel insights; in particular, we find strong support for Equisetales as sister to the rest of ferns, Marattiales as sister to leptosporangiate ferns, and Dennstaedtiaceae as sister to the eupolypods. Our divergence-time analyses reveal that divergences among the extant fern orders all occurred prior to ∼200 MYA. Finally, our species-tree inferences are congruent with analyses of concatenated data, but generally with lower support. Those cases where species-tree support values are higher than expected involve relationships that have been supported by smaller plastid datasets, suggesting that deep coalescence may be reducing support from the concatenated nuclear data.• CONCLUSIONS Our study demonstrates the utility of a curated phylogenomics approach to inferring fern phylogeny, and highlights the need to consider underlying data characteristics, along with data quantity, in phylogenetic studies.

Ancient vicariance and climate-driven extinction explain continental-wide disjunctions in Africa: the case of the Rand Flora genus Canarina (Campanulaceae)

Transoceanic distributions have attracted the interest of scientists for centuries. Less attention has been paid to the evolutionary origins of ‘continent‐wide’ disjunctions, in which related taxa are distributed across isolated regions within the same continent. A prime example is the ‘Rand Flora’ pattern, which shows sister taxa disjunctly distributed in the continental margins of Africa. Here, we explore the evolutionary origins of this pattern using the genus Canarina, with three species: C. canariensis, associated with the Canarian laurisilva, and C. eminii and C. abyssinica, endemic to the Afromontane region in East Africa, as case study. We infer phylogenetic relationships, divergence times and the history of migration events within Canarina using Bayesian inference on a large sample of chloroplast and nuclear sequences. Ecological niche modelling was employed to infer the climatic niche of Canarina through time. Dating was performed with a novel nested approach to solve the problem of using deep time calibration points within a molecular dataset comprising both above‐species and population‐level sampling. Results show C. abyssinica as sister to a clade formed by disjunct C. eminii and C. canariensis. Miocene divergences were inferred among species, whereas infraspecific divergences fell within the Pleistocene–Holocene periods. Although C. eminii and C. canariensis showed a strong genetic geographic structure, among‐population divergences were older in the former than in the latter. Our results suggest that Canarina originated in East Africa and later migrated across North Africa, with vicariance and aridification‐driven extinction explaining the 7000 km/7 million year divergence between the Canarian and East African endemics.

Disentangling knots of rapid evolution: origin and diversification of the moss order Hypnales

Abstract The Hypnales are the largest order of mosses comprising approximately 4200 species. Phylogenetic reconstruction within the group has proven to be difficult due to rapid radiation at an early stage of evolution and, consequently, relationships among clades have remained poorly resolved. We compiled data from four sequence regions, namely, nuclear ITS1–5·8S–ITS2, plastid trnL–F and rps4, and mitochondrial nad5, for 122 hypnalean species and 34 species from closely related groups. Tree topologies from both Bayesian and parsimony analyses resolve the order as monophyletic. Although inferences were made from fast-evolving genes, and despite strong phylogenetic signal in the nuclear ITS1–5·8S–ITS2 data, monophyly, as well as backbone nodes within the Hypnales, remains rather poorly supported except under Bayesian inferences. Ancestral distribution based on Bayesian dispersal-vicariance analysis supports a Gondwanan origin of the Hypnales and subsequent geographical radiation in the area of the former Laurasian supercontinent. Reconstruction of historical biogeography is congruent with mainly tropical and Gondwanan distributions in the sister groups Hypnodendrales, Ptychomniales, and Hookeriales, and with the dating for the oldest pleurocarp and hypnalean fossils. We contrast groupings in the phylogenetic tree with recent classifications and other phylogenetic inferences based on molecular data, and summarise current knowledge on the evolutionary history of, and relationships among, the Hypnales.

Living on the edge: timing of Rand Flora disjunctions congruent with ongoing aridification in Africa

The Rand Flora is a well-known floristic pattern in which unrelated plant lineages show similar disjunct distributions in the continental margins of Africa and adjacent islands—Macaronesia-northwest Africa, Horn of Africa-Southern Arabia, Eastern Africa, and Southern Africa. These lineages are now separated by environmental barriers such as the arid regions of the Sahara and Kalahari Deserts or the tropical lowlands of Central Africa. Alternative explanations for the Rand Flora pattern range from vicariance and climate-driven extinction of a widespread pan-African flora to independent dispersal events and speciation in situ. To provide a temporal framework for this pattern, we used published data from nuclear and chloroplast DNA to estimate the age of disjunction of 17 lineages that span 12 families and nine orders of angiosperms. We further used these estimates to infer diversification rates for Rand Flora disjunct clades in relation to their higher-level encompassing lineages. Our results indicate that most disjunctions fall within the Miocene and Pliocene periods, coinciding with the onset of a major aridification trend, still ongoing, in Africa. Age of disjunctions seemed to be related to the climatic affinities of each Rand Flora lineage, with sub-humid taxa dated earlier (e.g., Sideroxylon) and those with more xeric affinities (e.g., Campylanthus) diverging later. We did not find support for significant decreases in diversification rates in most groups, with the exception of older subtropical lineages (e.g., Sideroxylon, Hypericum, or Canarina), but some lineages (e.g., Cicer, Campylanthus) showed a long temporal gap between stem and crown ages, suggestive of extinction. In all, the Rand Flora pattern seems to fit the definition of biogeographic pseudocongruence, with the pattern arising at different times in response to the increasing aridity of the African continent, with interspersed periods of humidity allowing range expansions.

Phylogenetic analyses of morphological evolution in the gametophyte and sporophyte generations of the moss order Hookeriales (Bryopsida)

Morphological characters from the gametophyte and sporophyte generations have been used in land plants to infer relationships and construct classifications, but sporophytes provide the vast majority of data for the systematics of vascular plants. In bryophytes both generations are well developed and characters from both are commonly used to classify these organisms. However, because morphological traits of gametophytes and sporophytes can have different genetic bases and experience different selective pressures, taxonomic emphasis on one generation or the other may yield incongruent classifications. The moss order Hookeriales has a controversial taxonomic history because previous classifications have focused almost exclusively on either gametophytes or sporophytes. The Hookeriales provide a model for comparing morphological evolution in gametophytes and sporophytes, and its impact on alternative classification systems. In this study we reconstruct relationships among mosses that are or have been included in the Hookeriales based on sequences from five gene regions, and reconstruct morphological evolution of six sporophyte and gametophyte traits that have been used to differentiate families and genera. We found that the Hookeriales, as currently circumscribed, are monophyletic and that both sporophyte and gametophyte characters are labile. We documented parallel changes and reversals in traits from both generations. This study addresses the general issue of morphological reversals to ancestral states, and resolves novel relationships in the Hookeriales.

Geographic barriers and Pleistocene climate change shaped patterns of genetic variation in the Eastern Afromontane biodiversity hotspot

The Eastern African Afromontane forest is getting increased attention in conservation studies because of its high endemicity levels and shrinking geographic distribution. Phylogeographic studies have found evidence of high levels of genetic variation structured across the Great Rift System. Here, we use the epiphytic plant species Canarina eminii to explore causal explanations for this pattern. Phylogeographic analyses were undertaken using plastid regions and AFLP fragments. Population genetic analyses, Statistical Parsimony, and Bayesian methods were used to infer genetic diversity, genealogical relationships, structure, gene flow barriers, and the spatiotemporal evolution of populations. A strong phylogeographic structure was found, with two reciprocally monophyletic lineages on each side of the Great Rift System, high genetic exclusivity, and restricted gene flow among mountain ranges. We explain this pattern by topographic and ecological changes driven by geological rifting in Eastern Africa. Subsequent genetic structure is attributed to Pleistocene climatic changes, in which sky-islands acted as long-term refuges and cradles of genetic diversity. Our study highlights the importance of climate change and geographic barriers associated with the African Rift System in shaping population genetic patterns, as well as the need to preserve the high levels of exclusive and critically endangered biodiversity harboured by current patches of the Afromontane forest.