Analysis of auxin responses in the fern Ceratopteris richardii identifies the developmental phase as a major determinant for response properties.

The auxin signaling molecule regulates a range of plant growth and developmental processes. The core transcriptional machinery responsible for auxin-mediated responses is conserved across all land plants. Genetic, physiological and molecular exploration in bryophyte and angiosperm model species have shown both qualitative and quantitative differences in auxin responses. Given the highly divergent ontogeny of the dominant gametophyte (bryophytes) and sporophyte (angiosperms) generations, however, it is unclear whether such differences derive from distinct phylogeny or ontogeny. Here, we address this question by comparing a range of physiological, developmental and molecular responses to auxin in both generations of the model fern Ceratopteris richardii. We find that auxin response in Ceratopteris gametophytes closely resembles that of a thalloid bryophyte, whereas the sporophyte mimics auxin response in flowering plants. This resemblance manifests both at the phenotypic and transcriptional levels. Furthermore, we show that disrupting auxin transport can lead to ectopic sporophyte induction on the gametophyte, suggesting a role for auxin in the alternation of generations. Our study thus identifies developmental phase, rather than phylogeny, as a major determinant of auxin response properties in land plants.

Herbarium specimens as tools for exploring the evolution of fatty acid-derived natural products in plants.

Plants synthesize natural products via lineage-specific offshoots of their core metabolic pathways, including fatty acid synthesis. Recent studies have shed light on new fatty acid-derived natural products and their biosynthetic pathways in disparate plant species. Inspired by this progress, we set out to develop tools for exploring the evolution of fatty-acid derived products. We sampled multiple species from all major clades of euphyllophytes, including ferns, gymnosperms, and angiosperms (monocots and eudicots), and we show that the compositional profiles (though not necessarily the total amounts) of fatty-acid derived surface waxes from preserved plant specimens are consistent with those obtained from freshly collected tissue in a semi-quantitative and sometimes quantitative manner. We then sampled herbarium specimens representing 57 monocot species to assess the phylogenetic distribution and evolution, of two fatty acid-derived natural products found in that clade: beta-diketones and alkylresorcinols. These chemical data, combined with analyses of 26 monocot genomes, revealed a co-occurrence (though not necessarily a causal relationship) between whole genome duplication and the evolution of diketone synthases from an ancestral alkylresorcinol synthase-like polyketide synthase. Limitations of using herbarium specimen wax profiles as proxies for those of fresh tissue seem likely to include effects from loss of epicuticular wax crystals, effects from preservation techniques, and variation in wax chemical profiles due to genotype or environment. Nevertheless, this work reinforces the widespread utility of herbarium specimens for studying leaf surface waxes (and possibly other chemical classes) and reveals some of the evolutionary history of fatty acid-derived natural products within monocots.

The role of deep hybridization in fern speciation: Examples from the Thelypteridaceae.

PREMISE: Hybridization is recognized as an important mechanism in fern speciation, with many allopolyploids known among congeners, as well as evidence of ancient genome duplications. Several contemporary instances of deep (intergeneric) hybridization have been noted, invariably resulting in sterile progeny. We chose the christelloid lineage of the family Thelypteridaceae, recognized for its high frequency of both intra- and intergeneric hybrids, to investigate recent hybrid speciation between deeply diverged lineages. We also seek to understand the ecological and evolutionary outcomes of resulting lineages across the landscape. METHODS: By phasing captured reads within a phylogenomic data set of GoFlag 408 nuclear loci using HybPhaser, we investigated candidate hybrids to identify parental lineages. We estimated divergence ages by inferring a dated phylogeny using fossil calibrations with treePL. We investigated ecological niche conservatism between one confirmed intergeneric allotetraploid and its diploid progenitors using the centroid, overlap, unfilling, and expansion (COUE) framework. RESULTS: We provide evidence for at least six instances of intergeneric hybrid speciation within the christelloid clade and estimate up to 45 million years of divergence between progenitors. The niche quantification analysis showed moderate niche overlap between an allopolyploid species and its progenitors, with significant divergence from the niche of one progenitor and conservatism to the other. CONCLUSIONS: The examples provided here highlight the overlooked role that allopolyploidization following intergeneric hybridization may play in fern diversification and range and niche expansions. Applying this approach to other fern taxa may reveal a similar pattern of deep hybridization resulting in highly successful novel lineages.

Patterns in Genome-Wide Codon Usage Bias in Representative Species of Lycophytes and Ferns.

The latest research shows that ferns and lycophytes have distinct evolutionary lineages. The codon usage patterns of lycophytes and ferns have not yet been documented. To investigate the gene expression profiles across various plant lineages with respect to codon usage, analyze the disparities and determinants of gene evolution in primitive plant species, and identify appropriate exogenous gene expression platforms, the whole-genome sequences of four distinct species were retrieved from the NCBI database. The findings indicated that Ceratopteris richardii, Adiantum capillus-veneris, and Selaginella moellendorffii exhibited an elevated A/U content in their codon base composition and a tendency to end with A/U. Additionally, S. capillus-veneris had more C/G in its codons and a tendency to end with C/G. The ENC values derived from both ENC-plot and ENC-ratio analyses deviated significantly from the standard curves, suggesting that the codon usage preferences of these four species were primarily influenced by genetic mutations and natural selection, with natural selection exerting a more prominent influence. This finding was further supported by PR2-Plot, neutrality plot analysis, and COA. A combination of RSCU and ENC values was used as a reference criterion to rank the codons and further identify the optimal codons. The study identified 24 high-frequency codons in C. richardii, A. capillus-veneris, and Diphasiastrum complanatum, with no shared optimal codons among the four species. Arabidopsis thaliana and Ginkgo biloba exhibited similar codon preferences to the three species, except for S. moellendorffii. This research offers a theoretical framework at the genomic codon level for investigating the phylogenetic relationships between lycophytes and ferns, shedding light on gene codon optimization and its implications for genetic engineering in breeding.

Morphological and functional evolution of gametophytes in epilithic Hymenasplenium murakami-hatanakae (Aspleniaceae): The fifth family capable of producing the independent gametophytes.

The fern independent gametophytes that can maintain populations by vegetative reproduction without conspecific sporophytes have been considered an unusual phenomenon found in some epiphytic or epilithic species of Hymenophyllaceae, Pteridaceae, Lomariopsidaceae, and Polypodiaceae. By chance, the discovery of mysterious strap-like gametophytes on Izu-Oshima Island, Japan, has led to the hypothesis that Hymenasplenium murakami-hatanakae, a fern species belonging to Aspleniaceae, can also form independent gametophytes. Our investigation revealed gametophyte populations of H. murakami-hatanakae on three islands in the Izu Islands. Based on chloroplast DNA analysis of the gametophyte and sporophyte populations, the gametophytes were found to be maintained by vegetative reproduction without a new supply of spores from sporophytes. A comparison of the surrounding vegetation at the collection sites showed that environmental factors such as light and humidity may influence the maintenance of gametophyte populations. These results clearly show that H. murakami-hatanakae is one of the ferns capable of forming independent gametophytes. This is the first report of independent gametophytes from the suborder Aspleniineae (eupolypod II). The discovery of the independent gametophyte within a phylogenetic lineage previously thought not to form independent gametophytes will provide important insights into the morphological and functional evolution of gametophytes in ferns.

Hydrogen peroxide is required for light-induced stomatal opening across different plant species.

Stomatal movement is vital for plants to exchange gases and adaption to terrestrial habitats, which is regulated by environmental and phytohormonal signals. Here, we demonstrate that hydrogen peroxide (H2O2) is required for light-induced stomatal opening. H2O2 accumulates specifically in guard cells even when plants are under unstressed conditions. Reducing H2O2 content through chemical treatments or genetic manipulations results in impaired stomatal opening in response to light. This phenomenon is observed across different plant species, including lycopodium, fern, and monocotyledonous wheat. Additionally, we show that H2O2 induces the nuclear localization of KIN10 protein, the catalytic subunit of plant energy sensor SnRK1. The nuclear-localized KIN10 interacts with and phosphorylates the bZIP transcription factor bZIP30, leading to the formation of a heterodimer between bZIP30 and BRASSINAZOLE-RESISTANT1 (BZR1), the master regulator of brassinosteroid signaling. This heterodimer complex activates the expression of amylase, which enables guard cell starch degradation and promotes stomatal opening. Overall, these findings suggest that H2O2 plays a critical role in light-induced stomatal opening across different plant species.

Azolla filiculoides extract improved salt tolerance in wheat (Triticum aestivum L.) is associated with prompting osmostasis, antioxidant potential and stress-interrelated genes.

The growth and productivity of crop plants are negatively affected by salinity-induced ionic and oxidative stresses. This study aimed to provide insight into the interaction of NaCl-induced salinity with Azolla aqueous extract (AAE) regarding growth, antioxidant balance, and stress-responsive genes expression in wheat seedlings. In a pot experiment, wheat kernels were primed for 21 h with either deionized water or 0.1% AAE. Water-primed seedlings received either tap water, 250 mM NaCl, AAE spray, or AAE spray + NaCl. The AAE-primed seedlings received either tap water or 250 mM NaCl. Salinity lowered growth rate, chlorophyll level, and protein and amino acids pool. However, carotenoids, stress indicators (EL, MDA, and H2O2), osmomodulators (sugars, and proline), antioxidant enzymes (CAT, POD, APX, and PPO), and the expression of some stress-responsive genes (POD, PPO and PAL, PCS, and TLP) were significantly increased. However, administering AAE contributed to increased growth, balanced leaf pigments and assimilation efficacy, diminished stress indicators, rebalanced osmomodulators and antioxidant enzymes, and down-regulation of stress-induced genes in NaCl-stressed plants, with priming surpassing spray in most cases. In conclusion, AAE can be used as a green approach for sustaining regular growth and metabolism and remodelling the physio-chemical status of wheat seedlings thriving in salt-affected soils.

An adventurous journey toward and away from fern apomixis: Insights from genome size and spore abortion patterns.

PREMISE: Apomixis in ferns is relatively common and obligatory. Sterile hybrids may restore fertility via apomixis at a cost of long-term genetic stagnation. In this study, we outlined apomixis as a possible temporary phase leading to sexuality and analyzed factors relating to transitioning to and away from apomixis, such as unreduced and reduced spore formation in apomict and apo-sex hybrid ferns. METHODS: We analyzed the genome size of 15 fern species or hybrids ("taxa") via flow cytometry. The number of reduced and unreduced gametophytes was established as a proxy for viable spore formation of either type. We also calculated the spore abortion ratio (sign of reduced spores) in several taxa, including the apo-sex hybrid Dryopteris × critica and its 16 apomictically formed offspring. RESULTS: Four of 15 sampled taxa yielded offspring variable in genome size. Specifically, each variable taxon formed one viable reduced plant among 12-451 sampled gametophytes per taxon. Thus, haploid spore formation in the studied apomicts was very rare but possible. Spore abortion analyses indicated gradually decreasing abortion (haploid spore formation) over time. In Dryopteris × critica, abortion decreased from 93.8% to mean 89.5% in one generation. CONCLUSIONS: Our results support apomixis as a transitionary phase toward sexuality. Newly formed apomicts hybridize with sexual relatives and continue to form haploid spores early on. Thus, they may get the genomic content necessary for regular meiosis and restore sexuality. If the missing relative goes extinct, the lineage gets locked into apomixis as may be the case with the Dryopteris affinis complex.

Full-length transcriptome analysis of Ophioglossum vulgatum: effects of experimentally identified chloroplast gene clusters on expression and evolutionary patterns.

Genes with similar or related functions in chloroplasts are often arranged in close proximity, forming clusters on chromosomes. These clusters are transcribed coordinated to facilitate the expression of genes with specific function. Our previous study revealed a significant negative correlation between the chloroplast gene expression level of the rare medicinal fern Ophioglossum vulgatum and its evolutionary rates as well as selection pressure. Therefore, in this study, we employed a combination of SMRT and Illumina sequencing technology to analyze the full-length transcriptome sequencing of O. vulgatum for the first time. In particular, we experimentally identified gene clusters based on transcriptome data and investigated the effects of chloroplast gene clustering on expression and evolutionary patterns. The results revealed that the total sequenced data volume of the full-length transcriptome of O. vulgatum amounted to 71,950,652,163 bp, and 110 chloroplast genes received transcript coverage. Nine different types of gene clusters were experimentally identified in their transcripts. The chloroplast cluster genes may cause a decrease in non-synonymous substitution rate and selection pressure, as well as a reduction in transversion rate, transition rate, and their ratio. While expression levels of chloroplast cluster genes in leaf, sporangium, and stem would be relatively elevated. The Mann-Whitney U test indicated statistically significant in the selection pressure, sporangia and leaves groups (P < 0.05). We have contributed novel full-length transcriptome data resources for ferns, presenting new evidence on the effects of chloroplast gene clustering on expression land evolutionary patterns, and offering new theoretical support for transgenic research through gene clustering.

Apomixis and the paradox of sex in plants.

BACKGROUND: The predominance of sex in eukaryotes, despite the high costs of meiosis and mating, remains an evolutionary enigma. Many theories have been proposed, none of them being conclusive on its own, and they are, in part, not well applicable to land plants. Sexual reproduction is obligate in embryophytes for the great majority of species. SCOPE: This review compares the main forms of sexual and asexual reproduction in ferns and angiosperms, based on the generation cycling of sporophyte and gametophyte (leaving vegetative propagation aside). The benefits of sexual reproduction for maintenance of genomic integrity in comparison to asexuality are discussed in the light of developmental, evolutionary, genetic and phylogenetic studies. CONCLUSIONS: Asexual reproduction represents modifications of the sexual pathway, with various forms of facultative sexuality. For sexual land plants, meiosis provides direct DNA repair mechanisms for oxidative damage in reproductive tissues. The ploidy alternations of meiosis-syngamy cycles and prolonged multicellular stages in the haploid phase in the gametophytes provide a high efficiency of purifying selection against recessive deleterious mutations. Asexual lineages might buffer effects of such mutations via polyploidy and can purge the mutational load via facultative sexuality. The role of organelle-nuclear genome compatibility for maintenance of genome integrity is not well understood. In plants in general, the costs of mating are low because of predominant hermaphroditism. Phylogenetic patterns in the archaeplastid clade suggest that high frequencies of sexuality in land plants are concomitant with a stepwise increase of intrinsic and extrinsic stress factors. Furthermore, expansion of genome size in land plants would increase the potential mutational load. Sexual reproduction appears to be essential for keeping long-term genomic integrity, and only rare combinations of extrinsic and intrinsic factors allow for shifts to asexuality.

Incorporating fossils into the joint inference of phylogeny and biogeography of the tree fern order Cyatheales.

Present-day geographic and phylogenetic patterns often reflect the geological and climatic history of the planet. Neontological distribution data are often sufficient to unravel a lineage's biogeographic history, yet ancestral range inferences can be at odds with fossil evidence. Here, I use the fossilized birth-death process and the dispersal-extinction cladogenesis model to jointly infer the dated phylogeny and range evolution of the tree fern order Cyatheales. I use data for 101 fossil and 442 extant tree ferns to reconstruct the biogeographic history of the group over the last 220 million years. Fossil-aware reconstructions evince a prolonged occupancy of Laurasia over the Triassic-Cretaceous by Cyathealean tree ferns, which is evident in the fossil record but hidden from analyses relying on neontological data alone. Nonetheless, fossil-aware reconstructions are affected by uncertainty in fossils' phylogenetic placement, taphonomic biases, and specimen sampling and are sensitive to interpretation of paleodistributions and how these are scored. The present results highlight the need and challenges of incorporating fossils into joint inferences of phylogeny and biogeography to improve the reliability of ancestral geographic range estimation.

Unreduced spore formation in a spontaneous chimeric pinnule in an artificially produced haploid Anisocampium niponicum (Athyriaceae, Polypodiales).

Haploid sporophytes of Anisocampium niponicum with 2n = 40, were produced artificially by induced apogamy in vitro. They were subsequently transplanted into pots and two of them have been cultivated for the investigation of sporogenesis and/or production of chimera for more than 20 years. Haploid A. niponicum is sterile, but an abnormal chimeric pinnule that developed spontaneously in a single frond produced sporangia with spores. Each sporangium bore approximately 32 spores that were almost uniform in size. Sowing of these spores resulted in 50 gametophytes. Of 20 gametophytes cultured individually, five produced sporophytes apogamously after eight months. Both the gametophytes and subsequent apogamous sporophytes showed a chromosome number of 2n = 40. Our study demonstrates that a haploid sporophyte offspring can be produced from a haploid mother sporophyte via haploid spores. Since asexual reproduction is a prominent evolutionary process in ferns, the reproduction of a haploid A. niponicum sporophyte by unreduced spore formation might help to elucidate how apogamous ferns occur and evolve.

Characterization and analysis of multi-organ full-length transcriptomes in Sphaeropteris brunoniana and Alsophila latebrosa highlight secondary metabolism and chloroplast RNA editing pattern of tree ferns.

BACKGROUND: Sphaeropteris brunoniana and Alsophila latebrosa are both old relict and rare tree ferns, which have experienced the constant changes of climate and environment. However, little is known about their high-quality genetic information and related research on environmental adaptation mechanisms of them. In this study, combined with PacBio and Illumina platforms, transcriptomic analysis was conducted on the roots, rachis, and pinna of S. brunoniana and A. latebrosa to identify genes and pathways involved in environmental adaptation. Additionally, based on the transcriptomic data of tree ferns, chloroplast genes were mined to analyze their gene expression levels and RNA editing events. RESULTS: In the study, we obtained 11,625, 14,391 and 10,099 unigenes of S. brunoniana root, rachis, and pinna, respectively. Similarly, a total of 13,028, 11,431 and 12,144 unigenes were obtained of A. latebrosa root, rachis, and pinna, respectively. According to the enrichment results of differentially expressed genes, a large number of differentially expressed genes were enriched in photosynthesis and secondary metabolic pathways of S. brunoniana and A. latebrosa. Based on gene annotation results and phenylpropanoid synthesis pathways, two lignin synthesis pathways (H-lignin and G-lignin) were characterized of S. brunoniana. Among secondary metabolic pathways of A. latebrosa, three types of WRKY transcription factors were identified. Additionally, based on transcriptome data obtained in this study, reported transcriptome data, and laboratory available transcriptome data, positive selection sites were identified from 18 chloroplast protein-coding genes of four tree ferns. Among them, RNA editing was found in positive selection sites of four tree ferns. RNA editing affected the protein secondary structure of the rbcL gene. Furthermore, the expression level of chloroplast genes indicated high expression of genes related to the chloroplast photosynthetic system in all four species. CONCLUSIONS: Overall, this work provides a comprehensive transcriptome resource of S. brunoniana and A. latebrosa, laying the foundation for future tree fern research.

Differential expression and co-localization of transcriptional factors during callus transition to differentiation for shoot organogenesis in the water fern Ceratopteris richardii.

BACKGROUND AND AIMS: In flowering plants, regeneration can be achieved by a variety of approaches, and different sets of transcriptional factors are involved in these processes. However, regeneration in taxa other than flowering plants remains a mystery. Ceratopteris richardii is a representative fern capable of both direct and indirect organogenesis, and we aimed to investigate the genetics underlying the transition from callus proliferation to differentiation. METHODS: Morphological and histological analyses were used to determine the type of regeneration involved. RNA sequencing and differential gene expression were used to investigate how the callus switches from proliferation to differentiation. Phylogenetic analysis and RNA in situ hybridization were used to understand whether transcriptional factors are involved in this transition. KEY RESULTS: The callus formed on nascent leaves and subsequently developed the shoot pro-meristem and shoot meristem, thus completing indirect de novo shoot organogenesis in C. richardii. Genes were differentially expressed during the callus transition from proliferation to differentiation, indicating a role for photosynthesis, stimulus response and transmembrane signalling in this transition and the involvement of almost all cell layers that make up the callus. Transcriptional factors were either downregulated or upregulated, which were generally in many-to-many orthology with genes known to be involved in callus development in flowering plants, suggesting that the genetics of fern callus development are both conserved and divergent. Among them, an STM-like, a PLT-like and an ethylene- and salt-inducible ERF gene3-like gene were expressed simultaneously in the vasculature but not in the other parts of the callus, indicating that the vasculature played a role in the callus transition from proliferation to differentiation. CONCLUSIONS: Indirect de novo shoot organogenesis could occur in ferns, and the callus transition from proliferation to differentiation required physiological changes, differential expression of transcriptional factors and involvement of the vasculature.

'Out of Africa' origin of the pantropical staghorn fern genus Platycerium (Polypodiaceae) supported by plastid phylogenomics and biogeographical analysis.

BACKGROUND AND AIMS: The staghorn fern genus Platycerium is one of the most commonly grown ornamental ferns, and it evolved to occupy a typical pantropical intercontinental disjunction. However, species-level relationships in the genus have not been well resolved, and the spatiotemporal evolutionary history of the genus also needs to be explored. METHODS: Plastomes of all the 18 Platycerium species were newly sequenced. Using plastome data, we reconstructed the phylogenetic relationships among Polypodiaceae members with a focus on Platycerium species, and further conducted molecular dating and biogeographical analyses of the genus. KEY RESULTS: The present analyses yielded a robustly supported phylogenetic hypothesis of Platycerium. Molecular dating results showed that Platycerium split from its sister genus Hovenkampia ~35.2 million years ago (Ma) near the Eocene-Oligocene boundary and began to diverge ~26.3 Ma during the late Oligocene, while multiple speciation events within Platycerium occurred during the middle to late Miocene. Biogeographical analysis suggested that Platycerium originated in tropical Africa and then dispersed eastward to southeast Asia-Australasia and westward to neotropical areas. CONCLUSIONS: Our analyses using a plastid phylogenomic approach improved our understanding of the species-level relationships within Platycerium. The global climate changes of both the Late Oligocene Warming and the cooling following the mid-Miocene Climate Optimum may have promoted the speciation of Platycerium, and transoceanic long-distance dispersal is the most plausible explanation for the pantropical distribution of the genus today. Our study investigating the biogeographical history of Platycerium provides a case study not only for the formation of the pantropical intercontinental disjunction of this fern genus but also the 'out of Africa' origin of plant lineages.

The evolutionary innovation of root suberin lamellae contributed to the rise of seed plants.

Seed plants overtook ferns to become the dominant plant group during the late Carboniferous, a period in which the climate became colder and dryer1,2. However, the specific innovations driving the success of seed plants are not clear. Here we report that the appearance of suberin lamellae (SL) contributed to the rise of seed plants. We show that the Casparian strip and SL vascular barriers evolved at different times, with the former originating in the most recent common ancestor (MRCA) of vascular plants and the latter in the MRCA of seed plants. Our results further suggest that most of the genes required for suberin formation arose through gene duplication in the MRCA of seed plants. We show that the appearance of the SL in the MRCA of seed plants enhanced drought tolerance through preventing water loss from the stele. We hypothesize that SL provide a decisive selective advantage over ferns in arid environments, resulting in the decline of ferns and the rise of gymnosperms. This study provides insights into the evolutionary success of seed plants and has implications for engineering drought-tolerant crops or fern varieties.

Fern proteins show promise against crop pests.

These and other ancient plants may provide alternatives to chemical insecticides.

Leaf structure and water relations of an allotetraploid Mediterranean fern and its diploid parents.

Allopolyploidy is a common speciation mechanism in plants; however, its physiological and ecological consequences in niche partitioning have been scarcely studied. In this sense, leaf traits are good proxies to study the adaptive capacity of allopolyploids and diploid parents to their respective environmental conditions. In the present work, leaf water relations (assessed through pressure-volume curves) and structural and anatomical traits of the allotetraploid fern Oeosporangium tinaei and its diploid parents, Oeosporangium hispanicum and Oeosporangium pteridioides, were studied under controlled conditions in response to a water stress (WS) cycle. O. hispanicum showed the lowest osmotic potential at turgor loss point (πtlp ) and leaf capacitance, together with higher leaf mass per area (LMA), leaf thickness (LT), leaf density (LD), and leaf dry matter content (LDMC), whereas O. pteridioides presented the opposite set of traits (high πtlp and capacitance, and low LMA, LT, LD, and LDMC). O. tinaei showed an intermediate position for most of the studied traits. The responsiveness (osmotic and elastic adjustments) to WS was low, although most of the traits explained the segregation of the three species across a range of drought tolerance according to the rank: O. hispanicum > O. tinaei > O. pteridioides. These trait differences may underlie the niche segregation among coexisting populations of the three species in the Mediterranean basin.

Expression and Functional Studies of Leaf, Floral, and Fruit Developmental Genes in Non-model Species.

Researchers working on evolutionary developmental plant biology are inclined to choose non-model taxa to address how specific features have been acquired during ontogeny and fixed during phylogeny. In this chapter we describe methods to extract RNA, to assemble de-novo transcriptomes, to isolate orthologous genes within gene families, and to evaluate expression and function of target genes. We have successfully optimized these protocols for non-model plant species including ferns, gymnosperms, and a large assortment of angiosperms. In the latter, we have ranged a large number of families including Aristolochiaceae, Apodanthaceae, Chloranthaceae, Orchidaceae, Papaveraceae, Rubiaceae, Solanaceae, and Tropaeolaceae.

A Ceratopteris EXCESS MICROSPOROCYTES1 suppresses reproductive transition in the fern vegetative leaves.

Land plant sexual reproduction involves the transition of cells from somatic to reproductive identity during post-embryonic development. In Arabidopsis, the leucine-rich repeat receptor-like kinase EXCESS MICROSPOROCYTES1 (EXS/EMS1) restricts the number of sporogenous cells during the transition from diploid tissue to haploid spore production by promoting the formation of the tapetum cell layer within the anther. Although all land plants studied contain EMS1 genes, its function is unknown beyond a few angiosperms. In the model fern Ceratopteris (Ceratopteris richardii), we discovered an EMS1 homolog (CrEMS1) that functions to suppress formation of reproductive structures on vegetative leaves of the fern sporophyte, a role not found in angiosperms. Suppression of CrEMS1 by RNAi did not affect sporogenesis on reproductive leaves but did affect antheridium production of the fern gametophyte. Expression patterns of CrEMS1 across developmental stages suggest threshold levels of CrEMS1 control the specification of reproductive organs during both generations of the fern. Additional EMS1 homologs present in the fern genome suggest a dynamic role of EMS1 receptors in the evolution of reproductive development in vascular plants.