De novo assembly and comprehensive analysis of the mitochondrial genome of Taxus wallichiana reveals different repeats mediate recombination to generate multiple conformations.

Taxus plants are the exclusive source of paclitaxel, an anticancer drug with significant medicinal and economic value. Interspecies hybridization and gene introgression during evolution have obscured distinctions among Taxus species, complicating their phylogenetic classification. While the chloroplast genome of Taxus wallichiana, a widely distributed species in China, has been sequenced, its mitochondrial genome (mitogenome) remains uncharacterized.We sequenced and assembled the T. wallichiana mitogenome using BGI short reads and Nanopore long reads, facilitating comparisons with other gymnosperm mitogenomes. The T. wallichiana mitogenome spanning 469,949 bp, predominantly forms a circular configuration with a GC content of 50.51%, supplemented by 3 minor configurations mediated by one pair of LRs and two pairs of IntRs. It includes 32 protein-coding genes, 7 tRNA genes, and 3 rRNA genes, several of which exist in multiple copies.We detailed the mitogenome's structure, codon usage, RNA editing, and sequence migration between organelles, constructing a phylogenetic tree to elucidate evolutionary relationships. Unlike typical gymnosperm mitochondria, T. wallichiana shows no evidence of mitochondrial-plastid DNA transfer (MTPT), highlighting its unique genomic architecture. Synteny analysis indicated extensive genomic rearrangements in T. wallichiana, likely driven by recombination among abundant repetitive sequences. This study offers a high-quality T. wallichiana mitogenome, enhancing our understanding of gymnosperm mitochondrial evolution and supporting further cultivation and utilization of Taxus species.

Transcriptome sequencing data provides a solid base to understand the phylogenetic relationships, biogeography and reticulated evolution of the genus Zamia L. (Cycadales: Zamiaceae).

BACKGROUND AND AIMS: Cycad is a key lineage to understand the early evolution of seed plants and their response to past environmental changes. However, tracing the evolutionary trajectory of cycad species is challenging when the robust relationships at inter- or infrageneric level are not well resolved. METHODS: Here, using 2,901 single-copy nuclear genes, we explored the species relationships and gene flow within the second largest genus of cycads, i.e., Zamia, based on phylotranscriptomic analyses of 90% extant Zamia species. Based on a well-resolved phylogenetic framework, we performed gene flow analyses, molecular dating, and biogeographical reconstruction to examine the spatiotemporal evolution of Zamia. We also performed ancestral state reconstruction (ASR) of a total of 62 traits of the genus to comprehensively investigate its morphological evolution. KEY RESULTS: Zamia is comprised of seven major clades corresponding to seven distinct distribution areas in the Americas, with at least three reticulation nodes revealed in this genus. Extant lineages of Zamia initially diversified around 18.4-32.6 (29.14) million years ago (MA) in the Mega-Mexico, and then expanded eastward into the Caribbean and southward into Central and South America. ASR revealed homoplasy in most of the morphological characters. CONCLUSIONS: This study revealed congruent phylogenetic relationships from comparative methods/datasets, with some conflicts being the result of incomplete lineage sorting and ancient/recent hybridization events. The strong association between the clades and the biogeographic areas suggested that ancient dispersal events shaped the modern distribution pattern, and regional climatic factors may have resulted in the following in-situ diversification. Climate cooling starting during the mid Miocene is associated with the global expansion of Zamia to the tropical South America that have dramatically driven lineage diversification in the New World flora, as well as the extinction of cycad species in the nowadays cooler regions of both hemispheres as indicated by the fossil records.

Development of a Target Enrichment Probe Set for Conifer (REMcon).

Conifers are an ecologically and economically important seed plant group that can provide significant insights into the evolution of land plants. Molecular phylogenetics has developed as an important approach in evolutionary studies, although there have been relatively few studies of conifers that employ large-scale data sourced from multiple nuclear genes. Target enrichment sequencing (target capture, exon capture, or Hyb-Seq) has developed as a key approach in modern phylogenomic studies. However, until now, there has been no bait set that specifically targets the entire conifer clade. REMcon is a target sequence capture probe set intended for family- and species-level phylogenetic studies of conifers that target c. 100 single-copy nuclear loci. We tested the REMcon probe set using 69 species, including 44 conifer genera across six families and four other gymnosperm taxa, to evaluate the efficiency of target capture to efficiently generate comparable DNA sequence data across conifers. The recovery of target loci was high, with, on average, 94% of the targeted regions recovered across samples with high read coverage. A phylogenetic analysis of these data produced a well-supported topology that is consistent with the current understanding of relationships among conifers. The REMcon bait set will be useful in generating relatively large-scale nuclear data sets consistently for any conifer lineage.

The seed morphospace, a new contribution towards the multidimensional study of angiosperm sexual reproductive biology.

BACKGROUND: The evolutionary success of flowering plants is associated with the vast diversity of their reproductive structures. Despite recent progress in understanding angiosperm-wide trends of floral structure and evolution, a synthetic view of the diversity in seed form and function across angiosperms is lacking. SCOPE: Here we present a roadmap to synthesise the diversity of seed forms in extant angiosperms, relying on the morphospace concept, i.e. a mathematical representation which relates multiple traits and describes the realised morphologies. We provide recommendations on how to broaden the range of measurable traits beyond mass, by using key morphological traits representative of the embryo, endosperm, and seed coat but also fruit attributes (e.g., dehiscence, fleshiness). These key traits were used to construct and analyse a morphospace to detect evolutionary trends and gain insight into how morphological traits relate to seed functions. Finally, we outline challenges and future research directions, combining the morphospace with macroevolutionary comparative methods to underline the drivers that gave rise to the diversity of observed seed forms. CONCLUSIONS: We conclude that this multidimensional approach has the potential, although still untapped, to improve our understanding of covariation among reproductive traits, and further elucidate angiosperm reproductive biology as a whole.

Paternal Inheritance of Mitochondrial DNA May Lead to Dioecy in Conifers.

In angiosperms cytoplasmic DNA is typically passed on maternally through ovules. Genes in the mtDNA may cause male sterility. When male-sterile (female) cytotypes produce more seeds than cosexuals, they pass on more copies of their mtDNA and will co-occur with cosexuals with a neutral cytotype. Cytoplasmic gynodioecy is a well-known phenomenon in angiosperms, both in wild and crop plants. In some conifer families (e.g. Pinaceae) mitochondria are also maternally inherited. However in some other families (e.g. Taxaceae and Cupressaceae) mtDNA is paternally inherited through the pollen. With paternal mtDNA inheritance, male cytotypes that produce more pollen than cosexuals are expected to co-occur with cosexuals. This is uncharted territory. An ESS model shows that the presence of male cytotypes selects for more female allocation in the cosexual, i.e. for sexual specialisation. An allele that switches sex from male to female can then invade. This leads to rapid loss of the neutral cytotype of the cosexual, fixation of the male cytotype and dioecy with 50% males and 50% females. The models suggest that paternal inheritance of mtDNA facilitates the evolution dioecy. Consistent with this hypothesis the Pinaceae are 100% monoecious, while dioecy is common in the Taxaceae family and in the genus Juniperus (Cupressaceae). However, no reliable data are yet available on both mode of inheritance of mtDNA and gender variation of the same species. When cosexuals benefit from reproductive assurance (high selfing rate, low inbreeding depression, low fertilisation) they maintain themselves next to males and females. This predicted pattern with three sex types present in the same population is observed in conifers in nature.

Evolutionary history shapes variation of wood density of tree species across the world.

The effect of evolutionary history on wood density variation may play an important role in shaping variation in wood density, but this has largely not been tested. Using a comprehensive global dataset including 27,297 measurements of wood density from 2621 tree species worldwide, we test the hypothesis that the legacy of evolutionary history plays an important role in driving the variation of wood density among tree species. We assessed phylogenetic signal in different taxonomic (e.g., angiosperms and gymnosperms) and ecological (e.g., tropical, temperate, and boreal) groups of tree species, explored the biogeographical and phylogenetic patterns of wood density, and quantified the relative importance of current environmental factors (e.g., climatic and soil variables) and evolutionary history (i.e., phylogenetic relatedness among species and lineages) in driving global wood density variation. We found that wood density displayed a significant phylogenetic signal. Wood density differed among different biomes and climatic zones, with higher mean values of wood density in relatively drier regions (highest in subtropical desert). Our study revealed that at a global scale, for angiosperms and gymnosperms combined, phylogeny and species (representing the variance explained by taxonomy and not direct explained by long-term evolution process) explained 84.3% and 7.7% of total wood density variation, respectively, whereas current environment explained 2.7% of total wood density variation when phylogeny and species were taken into account. When angiosperms and gymnosperms were considered separately, the three proportions of explained variation are, respectively, 84.2%, 7.5% and 6.7% for angiosperms, and 45.7%, 21.3% and 18.6% for gymnosperms. Our study shows that evolutionary history outpaced current environmental factors in shaping global variation in wood density.

Enzymatic machinery of wood-inhabiting fungi that degrade temperate tree species.

Deadwood provides habitat for fungi and serves diverse ecological functions in forests. We already have profound knowledge of fungal assembly processes, physiological and enzymatic activities, and resulting physico-chemical changes during deadwood decay. However, in situ detection and identification methods, fungal origins, and a mechanistic understanding of the main lignocellulolytic enzymes are lacking. This study used metaproteomics to detect the main extracellular lignocellulolytic enzymes in 12 tree species in a temperate forest that have decomposed for 8 ½ years. Mainly white-rot (and few brown-rot) Basidiomycota were identified as the main wood decomposers, with Armillaria as the dominant genus; additionally, several soft-rot xylariaceous Ascomycota were identified. The key enzymes involved in lignocellulolysis included manganese peroxidase, peroxide-producing alcohol oxidases, laccase, diverse glycoside hydrolases (cellulase, glucosidase, xylanase), esterases, and lytic polysaccharide monooxygenases. The fungal community and enzyme composition differed among the 12 tree species. Ascomycota species were more prevalent in angiosperm logs than in gymnosperm logs. Regarding lignocellulolysis as a function, the extracellular enzyme toolbox acted simultaneously and was interrelated (e.g. peroxidases and peroxide-producing enzymes were strongly correlated), highly functionally redundant, and present in all logs. In summary, our in situ study provides comprehensive and detailed insight into the enzymatic machinery of wood-inhabiting fungi in temperate tree species. These findings will allow us to relate changes in environmental factors to lignocellulolysis as an ecosystem function in the future.

Mongolitria: A new Early Cretaceous three-valved seed from Northeast Asia.

PREMISE: Fossil seeds recovered from the Early Cretaceous of Mongolia and Inner Mongolia, China, are described and assigned to Mongolitria gen. nov., a new genus of gymnosperm seed. METHODS: Abundant lignitized seeds along with compression specimens isolated from the matrix were studied using a combination of scanning electron microscopy, anatomical sectioning, light microscopy, synchrotron radiation X-ray microtomography, and cuticle preparations. A single permineralized seed was examined by light microscopy of cellulose acetate peels and X-ray microtomography. RESULTS: Two species are recognized, Mongolitria friisae sp. nov. and Mongolitria exesum sp. nov. Both seeds are orthotropous with a short apical micropyle and a small, basal, circular attachment scar. The thick sclerenchymatous integument has a consistently three-parted organization and about 20 conspicuous longitudinal ribs on the surface. Mongolitria exesum differs from M. friisae primarily in its much larger size and thicker seed coat, which also preserves clear evidence of insect damage. CONCLUSIONS: Mongolitria is similar to other fossil seeds that have been assigned to Cycadales, but displays a unique combination of characters not found in any living or extinct cycadaceous plant, leaving its higher-level systematic affinities uncertain. Germination apparently involved splitting of the integument into three valves. Mongolitria was prominent among the plant parts accumulating in peat swamps in eastern Asia during the Early Cretaceous.

The Development of Plant Genome Sequencing Technology and Its Conservation and Application in Endangered Gymnosperms.

Genome sequencing is widely recognized as a fundamental pillar in genetic research and legal studies of biological phenomena, providing essential insights for genetic investigations and legal analyses of biological events. The field of genome sequencing has experienced significant progress due to rapid improvements in scientific and technological developments. These advancements encompass not only significant improvements in the speed and quality of sequencing but also provide an unparalleled opportunity to explore the subtle complexities of genomes, particularly in the context of rare species. Such a wide range of possibilities has successfully supported the validation of plant gene functions and the refinement of precision breeding methodologies. This expanded scope now includes a comprehensive exploration of the current state and conservation efforts of gymnosperm gene sequencing, offering invaluable insights into their genomic landscapes. This comprehensive review elucidates the trajectory of development and the diverse applications of genome sequencing. It encompasses various domains, including crop breeding, responses to abiotic stress, species evolutionary dynamics, biodiversity, and the unique challenges faced in the conservation and utilization of gymnosperms. It highlights both ongoing challenges and the unveiling of forthcoming developmental trajectories.

Distribution Patterns of Gymnosperm Species along Elevations on the Qinghai-Tibet Plateau: Effects of Climatic Seasonality, Energy-Water, and Physical Tolerance Variables.

Climate change is one of the most prominent factors influencing the spatial distribution of plants in China, including gymnosperms. Climatic factors influence gymnosperm distribution along elevational gradients on the Qinghai-Xizang (Tibet) Plateau (QTP), and understanding how species adapt to these factors is important for identifying the impacts of global climate change. For the first time, we examined the county-level distribution of gymnosperm species on QTP using data from field surveys, published works, monographs, and internet sources. We used simulated distribution data of gymnosperms (N = 79) along the elevational gradients to investigate the overall impact of environmental variables in explaining the richness pattern of gymnosperms. Eighteen environmental variables were classified into three key variable sets (climatic seasonality, energy-water, and physical tolerance). We employed principal component analysis and generalized linear models to assess the impact of climatic variables on the gymnosperm's richness pattern. Gymnosperm species are unevenly distributed across the plateau and decline gradually from the southeast to the northwest. The altitudinal gradients have a unimodal relationship with the richness of gymnosperms, with the maximum species richness at an elevation of 3200 m. The joint effects of physical tolerance and energy-water predictors have explained the highest diversity of gymnosperms at mid-elevation. Because the richness peak correlates significantly with the wettest month's precipitation and moisture index, this confirms the significance of moisture on gymnosperm distributions due to increased precipitation during the wet season. Furthermore, our results provide evidence that climatic seasonality factors are involved in the decline of gymnosperm richness at high elevations. A total of 37% of gymnosperm species on QTP are listed as vulnerable, nearly threatened, or endangered, with elevations ranging from 600 m to 5300 m. As a result, we conclude that gymnosperms are at high risk of extinction because of the current climate fluctuations caused by global climate change. Our research offers fundamental data for the study and protection of gymnosperm species along the steepest elevation gradients.

Quantifying the complexity of plant reproductive structures reveals a history of morphological and functional integration.

Vascular plant reproductive structures have undoubtedly become more complex through time, evolving highly differentiated parts that interact in specialized ways. But quantifying these patterns at broad scales is challenging because lineages produce disparate reproductive structures that are often difficult to compare and homologize. We develop a novel approach for analysing interactions within reproductive structures using networks, treating component parts as nodes and a suite of physical and functional interactions among parts as edges. We apply this approach to the plant fossil record, showing that interactions have generally increased through time and that the concentration of these interactions has shifted towards differentiated surrounding organs, resulting in more compact, functionally integrated structures. These processes are widespread across plant lineages, but their extent and timing vary with reproductive biology; in particular, seed-producing structures show them more strongly than spore or pollen-producing structures. Our results demonstrate that major reproductive innovations like the origin of seeds and angiospermy were associated with increased integration through greater interactions among parts. But they also reveal that for certain groups, particularly Mesozoic gymnosperms, millions of years elapsed between the origin of reproductive innovations and increased interactions among parts within their reproductive structures.

Large-Scale Identification and Characterization Analysis of VQ Family Genes in Plants, Especially Gymnosperms.

VQ motif-containing (VQ) proteins are a class of transcription regulatory cofactors widely present in plants, playing crucial roles in growth and development, stress response, and defense. Although there have been some reports on the member identification and functional research of VQ genes in some plants, there is still a lack of large-scale identification and clear graphical presentation of their basic characterization information to help us to better understand this family. Especially in gymnosperms, the VQ family genes and their evolutionary relationships have not yet been reported. In this study, we systematically identified 2469 VQ genes from 56 plant species, including bryophytes, gymnosperms, and angiosperms, and analyzed their molecular and evolutionary features. We found that amino acids are only highly conserved in the VQ domain, while other positions are relatively variable; most VQ genes encode relatively small proteins and do not have introns. The GC content in Poaceae plants is the highest (up to 70%); these VQ proteins can be divided into nine subgroups. In particular, we analyzed the molecular characteristics, chromosome distribution, duplication events, and expression levels of VQ genes in three gymnosperms: Ginkgo biloba, Taxus chinensis, and Pinus tabuliformis. In gymnosperms, VQ genes are classified into 11 groups, with highly similar motifs in each group; most VQ proteins have less than 300 amino acids and are predicted to be located in nucleus. Tandem duplication is an important driving force for the expansion of the VQ gene family, and the evolutionary processes of most VQ genes and duplication events are relatively independent; some candidate VQ genes are preliminarily screened, and they are likely to be involved in plant growth and stress and defense responses. These results provide detailed information and powerful references for further understanding and utilizing the VQ family genes in various plants.

Strong Purifying Selection in Haploid Tissue-Specific Genes of Scots Pine Supports the Masking Theory.

The masking theory states that genes expressed in a haploid stage will be under more efficient selection. In contrast, selection will be less efficient in genes expressed in a diploid stage, where the fitness effects of recessive deleterious or beneficial mutations can be hidden from selection in heterozygous form. This difference can influence several evolutionary processes such as the maintenance of genetic variation, adaptation rate, and genetic load. Masking theory expectations have been confirmed in single-cell haploid and diploid organisms. However, in multicellular organisms, such as plants, the effects of haploid selection are not clear-cut. In plants, the great majority of studies indicating haploid selection have been carried out using male haploid tissues in angiosperms. Hence, evidence in these systems is confounded with the effects of sexual selection and intraspecific competition. Evidence from other plant groups is scarce, and results show no support for the masking theory. Here, we have used a gymnosperm Scots pine megagametophyte, a maternally derived seed haploid tissue, and four diploid tissues to test the strength of purifying selection on a set of genes with tissue-specific expression. By using targeted resequencing data of those genes, we obtained estimates of genetic diversity, the site frequency spectrum of 0-fold and 4-fold sites, and inferred the distribution of fitness effects of new mutations in haploid and diploid tissue-specific genes. Our results show that purifying selection is stronger for tissue-specific genes expressed in the haploid megagametophyte tissue and that this signal of strong selection is not an artifact driven by high expression levels.

Model Species to Investigate the Origin of Flowers.

The angiosperms, or flowering plants, arose at least 135 million years ago (Ma) and rapidly diversified to form over 300,000 species alive today. This group appears, however, to have separated from its closest living relatives, the extant gymnosperms, much earlier: over 300 Ma. Representatives of basally-diverging angiosperm lineages are of key importance to studies aimed at reconstructing the most recent common ancestor of living angiosperms, including its morphological, anatomical, eco-physiological and molecular aspects. Furthermore, evo-devo comparisons of angiosperms with living gymnosperms may help to determine how the many novel aspects of angiosperms, including those of the flower, first came about. This chapter reviews literature on the origin of angiosperms and focusses on basally-diverging angiosperms and gymnosperms that show advantages as potential experimental models, reviewing information and protocols for the use of these species in an evo-devo context. The final section suggests a means by which data from living and fossil groups could be integrated to better elucidate evolutionary events that took place on the long stem-lineage that apparently preceded the radiation of living angiosperms.

An integrative view on the systematic position of the cupressophyte Cephalotaxus.

We made an in-depth review of historical studies of the cupressophyte conifer genus Cephalotaxus Siebold & Zucc. with an emphasis on its systematic position. We suggest that the systematic position of the genus is better understood using an integrative approach, so the evolution of phenetic characters is discussed within the context of recent phylogenomics. We propose that the genus should be classified as a separate family Cephalotaxaceae belonging to the clade consisting of Cupressaceae, Cephalotaxaceae, and Taxaceae; the family Cephalotaxaceae is sister to the Taxaceae but not nested within the Taxaceae and is characterized by a unique set of characters including morphology, anatomy, embryology, and chemistry. The family Cephalotaxaceae shows transitional characters between the Cupressaceae and the Taxaceae; the family possesses female cones with a primary cone axis bearing 5-8 pairs of decussate bracts, which is similar to the typical female cones of the Cupressaceae, on the one hand, and may have given rise to the reduced female cone of the Taxaceae with one terminal ovule partially or completely enclosed in a fleshy aril. In parallel, the compound male cone of the Cephalotaxaceae evolved into the seemingly "simple" male cones of the Taxaceae by means of reduction, elimination, and fusion.

Somatic Embryogenesis in Conifers: One Clade to Rule Them All?

Somatic embryogenesis (SE) in conifers is usually characterized as a multi-step process starting with the development of proembryogenic cell masses and followed by histodifferentiation, somatic embryo development, maturation, desiccation, and plant regeneration. Our current understanding of conifers' SE is mainly derived from studies using Pinaceae species as a model. However, the evolutionary relationships between conifers are not clear. Some hypotheses consider conifers as a paraphyletic group and Gnetales as a closely related clade. In this review, we used an integrated approach in order to cover the advances in knowledge on SE in conifers and Gnetales, discussing the state-of-the-art and shedding light on similarities and current bottlenecks. With this approach, we expect to be able to better understand the integration of these clades within current studies on SE. Finally, the points discussed raise an intriguing question: are non-Pinaceae conifers less prone to expressing embryogenic competence and generating somatic embryos as compared to Pinaceae species? The development of fundamental studies focused on this morphogenetic route in the coming years could be the key to finding a higher number of points in common between these species, allowing the success of the SE of one species to positively affect the success of another.

Taxonomic status and nomenclature of Cephalotaxuslanceolata (Cephalotaxaceae).

The nomenclature of Cephalotaxuslanceolata is controversial. After a thorough literature investigation, the nomenclatural problems have been resolved. This name was published in W. C. Cheng et al. (1975) and, although ascribed to "K. M. Feng", there is no suggestion that the descriptive material in the protologue was provided by K. M. Feng. Under Art. 46.5 of the ICN, this name should be attributed to K. M. Feng ex W. C. Cheng et al. but not to K. M. Feng alone. It has been claimed that the name is an illegitimate later homonym of one published by Beissner in 1901, but Beissner never accepted this name in any of his publications and so, under Art. 36.1, he did not validly publish an earlier homonym. Cephalotaxuslanceolatus was first validly published in W. C. Cheng et al. (1975).

Insect pollination in deep time.

Inferring insect pollination from compression fossils and amber inclusions is difficult because of a lack of consensus on defining an insect pollinator and the challenge of recognizing this ecological relationship in deep time. We propose a conceptual definition for such insects and an operational classification into pollinator or presumed pollinator. Using this approach, we identified 15 insect families that include fossil pollinators and show that pollination relationships have existed since at least the Upper Jurassic (~163 Ma). Insects prior to this can only be classified as presumed pollinators. This gives a more nuanced insight into the origin and evolution of an ecological relationship that is vital to the establishment, composition and conservation of modern terrestrial ecosystems.