Mesozoic cupules and the origin of the angiosperm second integument.

The second integument of the angiosperm ovule is unique among seed plants, with developmental genetics that are distinct from those of the inner integument1. Understanding how the second integument should be compared to structures in other seed plants is therefore crucial to resolving the long-standing question of the origin of angiosperms2-6. Attention has focused on several extinct plants with recurved cupules that are reminiscent of the anatropous organization of the basic bitegmic ovules of angiosperms1-6, but interpretations have been hampered by inadequate information on the relevant fossils. Here we describe abundant exceptionally well-preserved recurved cupules from a newly discovered silicified peat dating to the Early Cretaceous epoch (around 125.6 million years ago) in Inner Mongolia, China. The new material, combined with re-examination of potentially related fossils, indicates that the recurved cupules of several groups of Mesozoic plants are all fundamentally comparable, and that their structure is consistent with the recurved form and development of the second integument in the bitegmic anatropous ovules of angiosperms. Recognition of these angiosperm relatives (angiophytes) provides a partial answer to the question of angiosperm origins, will help to focus future work on seed plant phylogenetics and has important implications for ideas on the origin of the angiosperm carpel.

Glycome and Proteome Components of Golgi Membranes Are Common between Two Angiosperms with Distinct Cell-Wall Structures.

The plant endoplasmic reticulum-Golgi apparatus is the site of synthesis, assembly, and trafficking of all noncellulosic polysaccharides, proteoglycans, and proteins destined for the cell wall. As grass species make cell walls distinct from those of dicots and noncommelinid monocots, it has been assumed that the differences in cell-wall composition stem from differences in biosynthetic capacities of their respective Golgi. However, immunosorbence-based screens and carbohydrate linkage analysis of polysaccharides in Golgi membranes, enriched by flotation centrifugation from etiolated coleoptiles of maize (Zea mays) and leaves of Arabidopsis (Arabidopsis thaliana), showed that arabinogalactan-proteins and arabinans represent substantial portions of the Golgi-resident polysaccharides not typically found in high abundance in cell walls of either species. Further, hemicelluloses accumulated in Golgi at levels that contrasted with those found in their respective cell walls, with xyloglucans enriched in maize Golgi, and xylans enriched in Arabidopsis. Consistent with this finding, maize Golgi membranes isolated by flotation centrifugation and enriched further by free-flow electrophoresis, yielded >200 proteins known to function in the biosynthesis and metabolism of cell-wall polysaccharides common to all angiosperms, and not just those specific to cell-wall type. We propose that the distinctive compositions of grass primary cell walls compared with other angiosperms result from differential gating or metabolism of secreted polysaccharides post-Golgi by an as-yet unknown mechanism, and not necessarily by differential expression of genes encoding specific synthase complexes.

High porosity with tiny pore constrictions and unbending pathways characterize the 3D structure of intervessel pit membranes in angiosperm xylem.

Pit membranes between xylem vessels play a major role in angiosperm water transport. Yet, their three-dimensional (3D) structure as fibrous porous media remains unknown, largely due to technical challenges and sample preparation artefacts. Here, we applied a modelling approach based on thickness measurements of fresh and fully shrunken pit membranes of seven species. Pore constrictions were also investigated visually by perfusing fresh material with colloidal gold particles of known sizes. Based on a shrinkage model, fresh pit membranes showed tiny pore constrictions of ca. 20 nm, but a very high porosity (i.e. pore volume fraction) of on average 0.81. Perfusion experiments showed similar pore constrictions in fresh samples, well below 50 nm based on transmission electron microscopy. Drying caused a 50% shrinkage of pit membranes, resulting in much smaller pore constrictions. These findings suggest that pit membranes represent a mesoporous medium, with the pore space characterized by multiple constrictions. Constrictions are much smaller than previously assumed, but the pore volume is large and highly interconnected. Pores do not form highly tortuous, bent, or zigzagging pathways. These insights provide a novel view on pit membranes, which is essential to develop a mechanistic, 3D understanding of air-seeding through this porous medium.

Evolution of Darwin's Peloric Gloxinia (Sinningia speciosa) Is Caused by a Null Mutation in a Pleiotropic TCP Gene.

Unlike most crops, which were domesticated through long periods of selection by ancient humans, horticultural plants were primarily domesticated through intentional selection over short time periods. The molecular mechanisms underlying the origin and spread of novel traits in the domestication process have remained largely unexplored in horticultural plants. Gloxinia (Sinningia speciosa), whose attractive peloric flowers influenced the thoughts of Darwin, have been cultivated since the early 19th century, but its origin and genetic basis are currently unknown. By employing multiple experimental approaches including genetic analysis, genotype-phenotype associations, gene expression analysis, and functional interrogations, we showed that a single gene encoding a TCP protein, SsCYC, controls both floral orientation and zygomorphy in gloxinia. We revealed that a causal mutation responsible for the development of peloric gloxinia lies in a 10-bp deletion in the coding sequence of SsCYC. By combining genetic inference and literature searches, we have traced the putative ancestor and reconstructed the domestication path of the peloric gloxinia, in which a 10-bp deletion in SsCYC under selection triggered its evolution from the wild progenitor. The results presented here suggest that a simple genetic change in a pleiotropic gene can promote the elaboration of floral organs under intensive selection pressure.

How deep is the conflict between molecular and fossil evidence on the age of angiosperms?

The timing of the origin of angiosperms is a hotly debated topic in plant evolution. Molecular dating analyses that consistently retrieve pre-Cretaceous ages for crown-group angiosperms have eroded confidence in the fossil record, which indicates a radiation and possibly also origin in the Early Cretaceous. Here, we evaluate paleobotanical evidence on the age of the angiosperms, showing how fossils provide crucial data for clarifying the situation. Pollen floras document a Northern Gondwanan appearance of monosulcate angiosperms in the Valanginian and subsequent poleward spread of monosulcates and tricolpate eudicots, accelerating in the Albian. The sequence of pollen types agrees with molecular phylogenetic inferences on the course of pollen evolution, but it conflicts strongly with Triassic and early Jurassic molecular ages, and the discrepancy is difficult to explain by geographic or taphonomic biases. Critical scrutiny shows that supposed pre-Cretaceous angiosperms either represent other plant groups or lack features that might confidently assign them to the angiosperms. However, the record may allow the Late Jurassic existence of ecologically restricted angiosperms, like those seen in the basal ANITA grade. Finally, we examine recently recognized biases in molecular dating and argue that a thoughtful integration of fossil and molecular evidence could help resolve these conflicts.

Homologs of the STYLISH gene family control nectary development in Aquilegia.

Floral nectaries are an interesting example of a convergent trait in flowering plants, and are associated with the diversification of numerous angiosperm lineages, including the adaptive radiation of the New World Aquilegia species. However, we know very little as to what genes contribute to nectary development and evolution, particularly in noncore eudicot taxa. We analyzed expression patterns and used RNAi-based methods to investigate the functions of homologs from the STYLISH (STY) family in nectar spur development in Aquilegia coerulea. We found that AqSTY1 exhibits concentrated expression in the presumptive nectary of the growing spur tip, and triple gene silencing of the three STY-like genes revealed that they function in style and nectary development. Strong expression of STY homologs was also detected in the nectary-bearing petals of Delphinium and Epimedium. Our results suggest that the novel recruitment of STY homologs to control nectary development is likely to have occurred before the diversification of the Ranunculaceae and Berberidaceae. To date, the STY homologs of the Ranunculales are the only alternative loci for the control of nectary development in flowering plants, providing a critical data point in understanding the evolutionary origin and developmental basis of nectaries.

The role of the integuments in pollen tube guidance in flowering plants.

In angiosperms, pollen tube entry into the ovule generally takes place through the micropyle, but the exact role of the micropyle in pollen tube guidance remains unclear. A limited number of studies have examined eudicots with bitegmic micropyles, but information is lacking in ovules of basal/early-divergent angiosperms with unitegmic micropyles. We have evaluated the role of the micropyle in pollen tube guidance in an early-divergent angiosperm (Annona cherimola) and the evolutionarily derived Arabidopsis thaliana by studying γ-aminobutyric acid (GABA) and arabinogalactan proteins (AGPs) in wild-type plants and integument-defective mutants. A conserved inhibitory role of GABA in pollen tube growth was shown in A. cherimola, in which AGPs surround the egg apparatus. In Arabidopsis, the micropyle formed only by the outer integument in wuschel-7 mutants caused a partial defect in pollen tube guidance. Moreover, pollen tubes were not observed in the micropyle of an inner no outer (ino) mutant in Arabidopsis, but were observed in homologous ino mutants in Annona. The similar distribution of GABA and AGPs observed in the micropyle of Arabidopsis and Annona, together with the anomalies from specific integument mutants, support the role of the inner integument in preventing multiple tube entrance (polytubey) in these two phylogenetically distant genera.

Cretaceous asterid evolution: fruits of Eydeia jerseyensis sp. nov. (Cornales) from the upper Turonian of eastern North America.

BACKGROUND AND AIMS: The asterids (>80 000 extant species) appear in the fossil record with considerable diversity near the Turonian-Coniacian boundary (~90 Ma; Late Cretaceous) and are strongly represented in the earliest diverging lineage, Cornales. These early asterid representatives have so far been reported from western North America and eastern Asia. In this study, we characterize a new cornalean taxon based on charcoalified fruits from the upper Turonian of eastern North America, a separate landmass from western North America at the time, and identify early palaeobiogeographical patterns of Cornales during the Cretaceous. METHODS: Fossils were studied and imaged using scanning electron microscopy and micro-computed tomography (micro-CT) scanning. To assess the systematic affinities of the fossils, phylogenetic analyses were conducted using maximum parsimony. KEY RESULTS: The charcoalified fruits are represented by tri-locular woody endocarps with dorsal apically opening germination valves. Three septa intersect to form a robust central axis. Endocarp ground tissue consists of two zones: an outer endocarp composed of isodiametric sclereids and an inner endocarp containing circum-locular fibres. Central vasculature is absent; however, there are several small vascular bundles scattered within the septa. Phylogenetic analysis places the new taxon within the extinct genus Eydeia. DISCUSSION: Thick-walled endocarps with apically opening germination valves, no central vascular bundle and one seed per locule are indicative of the order Cornales. Comparative analysis suggests that the fossils represent a new species, Eydeia jerseyensis sp. nov. This new taxon is the first evidence of Cornales in eastern North America during the Cretaceous and provides insights into the palaeobiogeography and initial diversification of the order.

Xylem Surfactants Introduce a New Element to the Cohesion-Tension Theory.

Vascular plants transport water under negative pressure without constantly creating gas bubbles that would disable their hydraulic systems. Attempts to replicate this feat in artificial systems almost invariably result in bubble formation, except under highly controlled conditions with pure water and only hydrophilic surfaces present. In theory, conditions in the xylem should favor bubble nucleation even more: there are millions of conduits with at least some hydrophobic surfaces, and xylem sap is saturated or sometimes supersaturated with atmospheric gas and may contain surface-active molecules that can lower surface tension. So how do plants transport water under negative pressure? Here, we show that angiosperm xylem contains abundant hydrophobic surfaces as well as insoluble lipid surfactants, including phospholipids, and proteins, a composition similar to pulmonary surfactants. Lipid surfactants were found in xylem sap and as nanoparticles under transmission electron microscopy in pores of intervessel pit membranes and deposited on vessel wall surfaces. Nanoparticles observed in xylem sap via nanoparticle-tracking analysis included surfactant-coated nanobubbles when examined by freeze-fracture electron microscopy. Based on their fracture behavior, this technique is able to distinguish between dense-core particles, liquid-filled, bilayer-coated vesicles/liposomes, and gas-filled bubbles. Xylem surfactants showed strong surface activity that reduces surface tension to low values when concentrated as they are in pit membrane pores. We hypothesize that xylem surfactants support water transport under negative pressure as explained by the cohesion-tension theory by coating hydrophobic surfaces and nanobubbles, thereby keeping the latter below the critical size at which bubbles would expand to form embolisms.

Tanispermum, a new genus of hemi-orthotropous to hemi-anatropous angiosperm seeds from the Early Cretaceous of eastern North America.

PREMISE OF THE STUDY: Exotestal seeds with characters that indicate relationship to extant Austrobaileyales and Nymphaeales are abundant in Early Cretaceous sediments from Portugal and eastern North America, but their variety and unique features provide evidence of extensive extinct diversity among early angiosperms. METHODS: The fossils were extracted from Early Cretaceous sediments from Virginia and Maryland, United States, by sieving in water. After cleaning with HF, HCl and water, they were examined using SEM and SRXTM and compared to seeds of extant and fossil angiosperms. KEY RESULTS: A new genus, Tanispermum gen. nov., with four species (T. hopewellense sp. nov., T. marylandense sp. nov., T. drewriense sp. nov., and T. antiquum sp. nov.) is recognized. Relationship with extant Austrobaileyales and Nymphaeales is indicated by an exotesta of tall palisade sclerenchyma with undulate anticlinal walls forming a jigsaw puzzle-like surface pattern. However, seeds of Tanispermum differ from those of Austrobaileyales and Nymphaeales in their hemi-orthotropous-hemi-anatropous organization. Attempts to place Tanispermum in a phylogenetic context confront a variety of problems, including lack of information on other parts of these extinct plants. CONCLUSIONS: The discovery highlights the extent to which the morphology of extant angiosperms is not representative of the diversity that once existed among early-diverging members of the group. The relictual nature of extant taxa near the base of the angiosperm tree greatly complicates the reconstruction of ancestral character states and emphasizes the need for focused paleobotanical studies to elucidate the extinct diversity that existed early in angiosperm evolution.

Vessel-associated cells in angiosperm xylem: Highly specialized living cells at the symplast-apoplast boundary.

BACKGROUND: Vessel-associated cells (VACs) are highly specialized, living parenchyma cells that are in direct contact with water-conducting, dead vessels. The contact may be sparse or in large tight groups of parenchyma that completely surrounds vessels. VACs differ from vessel distant parenchyma in physiology, anatomy, and function and have half-bordered pits at the vessel-parenchyma juncture. The distinct anatomy of VACs is related to the exchange of substances to and from the water-transport system, with the cells long thought to be involved in water transport in woody angiosperms, but where direct experimental evidence is lacking. SCOPE: This review focuses on our current knowledge of VACs regarding anatomy and function, including hydraulic capacitance, storage of nonstructural carbohydrates, symplastic and apoplastic interactions, defense against pathogens and frost, osmoregulation, and the novel hypothesis of surfactant production. Based on microscopy, we visually represent how VACs vary in dimensions and general appearance between species, with special attention to the protoplast, amorphous layer, and the vessel-parenchyma pit membrane. CONCLUSIONS: An understanding of the relationship between VACs and vessels is crucial to tackling questions related to how water is transported over long distances in xylem, as well as defense against pathogens. New avenues of research show how parenchyma-vessel contact is related to vessel diameter and a new hypothesis may explain how surfactants arising from VAC can allow water to travel under negative pressure. We also reinforce the message of connectivity between VAC and other cells between xylem and phloem.

A new commelinid monocot seed fossil from the early Eocene previously identified as Solanaceae.

PREMISE OF THE STUDY: Fossils provide minimum age estimates for extant lineages. Here we critically evaluate Cantisolanum daturoides Reid & Chandler and two other early putative seed fossils of Solanaceae, an economically important plant family in the Asteridae. METHODS: Three earliest seed fossil taxa of Solanaceae from the London Clay Formation (Cantisolanum daturoides) and the Poole and Branksome Sand Formations (Solanum arnense Chandler and Solanispermum reniforme Chandler) were studied using x-ray microcomputed tomography (MCT) and scanning electron microscopy (SEM). KEY RESULTS: The MCT scans of Cantisolanum daturoides revealed a high level of pyrite preservation at the cellular level. Cantisolanum daturoides can be clearly excluded from Solanaceae and has more affinities to the commelinid monocots based on a straight longitudinal axis, a prominent single layer of relatively thin-walled cells in the testa, and a clearly differentiated micropyle surrounded by radially elongated and inwardly curved testal cells. While the MCT scans show no internal preservation in Solanum arnense and Solanispermum reniforme, SEM images show the presence of several characteristics that allow the placement of these taxa at the stem node of Solanaceae. CONCLUSIONS: Cantisolanum daturoides is likely a member of commelinid monocots and not Solanaceae as previously suggested. The earliest fossil record of Solanaceae is revised to consist of fruit fossil with inflated calyces from the early Eocene of Patagonia (52 Ma) and fossilized seeds from the early to mid-Eocene of Europe (48-46 Ma). The new identity for Cantisolanum daturoides does not alter a late Cretaceous minimum age for commelinids.

From the sap's perspective: The nature of vessel surfaces in angiosperm xylem.

PREMISE OF THE STUDY: Xylem sap in angiosperms moves under negative pressure in conduits and cell wall pores that are nanometers to micrometers in diameter, so sap is always very close to surfaces. Surfaces matter for water transport because hydrophobic ones favor nucleation of bubbles, and surface chemistry can have strong effects on flow. Vessel walls contain cellulose, hemicellulose, lignin, pectins, proteins, and possibly lipids, but what is the nature of the inner, lumen-facing surface that is in contact with sap? METHODS: Vessel lumen surfaces of five angiosperms from different lineages were examined via transmission electron microscopy and confocal and fluorescence microscopy, using fluorophores and autofluorescence to detect cell wall components. Elemental composition was studied by energy-dispersive X-ray spectroscopy, and treatments with phospholipase C (PLC) were used to test for phospholipids. KEY RESULTS: Vessel surfaces consisted mainly of lignin, with strong cellulose signals confined to pit membranes. Proteins were found mainly in inter-vessel pits and pectins only on outer rims of pit membranes and in vessel-parenchyma pits. Continuous layers of lipids were detected on most vessel surfaces and on most pit membranes and were shown by PLC treatment to consist at least partly of phospholipids. CONCLUSIONS: Vessel surfaces appear to be wettable because lignin is not strongly hydrophobic and a coating with amphiphilic lipids would render any surface hydrophilic. New questions arise about these lipids and their possible origins from living xylem cells, especially about their effects on surface tension, surface bubble nucleation, and pit membrane function.

Evolution of root apical meristem structures in vascular plants: plasmodesmatal networks.

PREMISE OF THE STUDY: The apical meristem generates indeterminate apical growth of the stem and root of vascular plants. Our previous examination showed that shoot apical meristems (SAMs) can be classified into two types based on plasmodesmatal networks (PNs), which are important elements in symplasmic signaling pathways within the apical meristem. Here, we examined the PNs of root apical meristems (RAMs) in comparison with those of SAMs. METHODS: Root apical meristems of 18 families and 22 species of lycophytes and euphyllophytes were analyzed. Plasmodesmata (PD) in cell walls in median longitudinal sections of RAMs were enumerated using transmission electron micrographs, and the PD density per 1 µm2 of each cell wall was calculated. KEY RESULTS: Root apical meristems with prominent apical cells of monilophytes (euphyllophytes) and Selaginellaceae (lycophytes) had high PD densities, while RAMs with plural initial cells of gymnosperms and angiosperms (euphyllophytes), and of Lycopodiaceae and Isoetaceae (lycophytes) had low PD densities. This correlation between structures of apical meristems and PD densities is identical to that in SAMs already described. CONCLUSIONS: Irrespective of their diversified structures, the RAMs of vascular plants can be classified into two types with respect to PNs: the fern (monilophyte) type, which has a lineage-specific PN with only primary PD, and the seed-plant type, which has an interspecific PN with secondary PD in addition to primary PD. PNs may have played a key role in the evolution of apical meristems in vascular plants.

Floral meristem size and organ number correlation in Eucryphia (Cunoniaceae).

We present a comparative flower ontogenetic study in five species of the genus Eucryphia with the aim of testing whether differences in the organ number observed can be explained by changes in the meristematic size of floral meristem and floral organs. Species native to Oceania, viz. E. milliganii, E. lucida and E. moorei, have the smallest gynoecia with ca. 6 carpels, while the Chilean E. glutinosa and E. cordifolia present more than ten carpels. E. milliganii has the smallest flower with the lowest stamen number (ca. 50), while the other species produce around 200 stamens and more. Standardized measurements of meristematic sectors were taken in 49 developing flowers that were classified into three well-defined ontogenetic stages. Sizes of meristems varied significantly among species within each developmental stage as revealed by ANOVA analyses. Significant regressions between organ number and corresponding meristem size were consistent with the premise that a larger meristem size prior to organ initiation could be determining for a higher organ number. Flower organogenesis in Eucryphia also involves relevant meristem expansion while the organs are initiated, which results in a particular androecium patterning with a chaotic stamen arrangement. Meristem expansion also appears to be slower but more extensive in species with larger initial meristematic size, suggesting that flower phenotype can be determined in ontogeny by this heterochronic interplay of space and time.

Understanding the role of floral development in the evolution of angiosperm flowers: clarifications from a historical and physico-dynamic perspective.

Flower morphology results from the interaction of an established genetic program, the influence of external forces induced by pollination systems, and physical forces acting before, during and after initiation. Floral ontogeny, as the process of development from a meristem to a fully developed flower, can be approached either from a historical perspective, as a "recapitulation of the phylogeny" mainly explained as a process of genetic mutations through time, or from a physico-dynamic perspective, where time, spatial pressures, and growth processes are determining factors in creating the floral morphospace. The first (historical) perspective clarifies how flower morphology is the result of development over time, where evolutionary changes are only possible using building blocks that are available at a certain stage in the developmental history. Flowers are regulated by genetically determined constraints and development clarifies specific transitions between different floral morphs. These constraints are the result of inherent mutations or are induced by the interaction of flowers with pollinators. The second (physico-dynamic) perspective explains how changes in the physical environment of apical meristems create shifts in ontogeny and this is reflected in the morphospace of flowers. Changes in morphology are mainly induced by shifts in space, caused by the time of initiation (heterochrony), pressure of organs, and alterations of the size of the floral meristem, and these operate independently or in parallel with genetic factors. A number of examples demonstrate this interaction and its importance in the establishment of different floral forms. Both perspectives are complementary and should be considered in the understanding of factors regulating floral development. It is suggested that floral evolution is the result of alternating bursts of physical constraints and genetic stabilization processes following each other in succession. Future research needs to combine these different perspectives in understanding the evolution of floral systems and their diversification.

Bordered pits in xylem of vesselless angiosperms and their possible misinterpretation as perforation plates.

Vesselless wood represents a rare phenomenon within the angiosperms, characterizing Amborellaceae, Trochodendraceae and Winteraceae. Anatomical observations of bordered pits and their pit membranes based on light, scanning and transmission electron microscopy (SEM and TEM) are required to understand functional questions surrounding vesselless angiosperms and the potential occurrence of cryptic vessels. Interconduit pit membranes in 11 vesselless species showed a similar ultrastructure as mesophytic vessel-bearing angiosperms, with a mean thickness of 245 nm (± 53, SD; n = six species). Shrunken, damaged and aspirated pit membranes, which were 52% thinner than pit membranes in fresh samples (n = four species), occurred in all dried-and-rehydrated samples, and in fresh latewood of Tetracentron sinense and Trochodendron aralioides. SEM demonstrated that shrunken pit membranes showed artificially enlarged, > 100 nm wide pores. Moreover, perfusion experiments with stem segments of Drimys winteri showed that 20 and 50 nm colloidal gold particles only passed through 2 cm long dried-and-rehydrated segments, but not through similar sized fresh ones. These results indicate that pit membrane shrinkage is irreversible and associated with a considerable increase in pore size. Moreover, our findings suggest that pit membrane damage, which may occur in planta, could explain earlier records of vessels in vesselless angiosperms.

Stomatal cell wall composition: distinctive structural patterns associated with different phylogenetic groups.

Background and Aims: Stomatal morphology and function have remained largely conserved throughout ∼400 million years of plant evolution. However, plant cell wall composition has evolved and changed. Here stomatal cell wall composition was investigated in different vascular plant groups in attempt to understand their possible effect on stomatal function. Methods: A renewed look at stomatal cell walls was attempted utilizing digitalized polar microscopy, confocal microscopy, histology and a numerical finite-elements simulation. The six species of vascular plants chosen for this study cover a broad structural, ecophysiological and evolutionary spectrum: ferns ( Asplenium nidus and Platycerium bifurcatum ) and angiosperms ( Arabidopsis thaliana and Commelina erecta ) with kidney-shaped stomata, and grasses (angiosperms, family Poaceae) with dumbbell-shaped stomata ( Sorghum bicolor and Triticum aestivum ). Key Results: Three distinct patterns of cellulose crystallinity in stomatal cell walls were observed: Type I (kidney-shaped stomata, ferns), Type II (kidney-shaped stomata, angiosperms) and Type III (dumbbell-shaped stomata, grasses). The different stomatal cell wall attributes investigated (cellulose crystallinity, pectins, lignin, phenolics) exhibited taxon-specific patterns, with reciprocal substitution of structural elements in the end-walls of kidney-shaped stomata. According to a numerical bio-mechanical model, the end walls of kidney-shaped stomata develop the highest stresses during opening. Conclusions: The data presented demonstrate for the first time the existence of distinct spatial patterns of varying cellulose crystallinity in guard cell walls. It is also highly intriguing that in angiosperms crystalline cellulose appears to have replaced lignin that occurs in the stomatal end-walls of ferns serving a similar wall strengthening function. Such taxon-specific spatial patterns of cell wall components could imply different biomechanical functions, which in turn could be a consequence of differences in environmental selection along the course of plant evolution.

Floral morphology and anatomy of Ophiocaryon, a paedomorphic genus of Sabiaceae.

Background and Aims: Ophiocaryon is a lesser known genus in Sabiaceae. This study examines flowers of six Ophiocaryon species in comparison with Meliosmaalba, to identify taxonomically informative characters for understanding relationships within the family Sabiaceae, to imply previously unknown pollination mechanisms of Ophiocaryon, and to contribute to the placement of Sabiaceae within the early-diverging eudicots. Methods: Floral morphology and anatomy of six Ophiocaryon species and M. alba were studied and described using scanning electron microscopy, clearing techniques and resin sectioning. Key Results: Novel characters of Ophiocaryon were identified, e.g. conical cells on petals, different kinds of orbicules in anthers, stomata on nectary appendage tips and ovary, two distinct surface patterns on stamens and ovary, tanniferous cell layers in the ovary wall, and acorn-shaped unitegmic ovules with very short integuments. Comparison of floral characters between Ophiocaryon and Meliosma found that the calyx, corolla, androecium and gynoecium of Ophiocaryon resemble an undeveloped state of the latter taxon, reflecting a paedomorphic regression of the flower of Ophiocaryon. The flower morphology and anatomy of Ophiocaryon was compared with its putative sister species M. alba, but no clear shared derived characters could be detected. Moreover, the findings of scent, presence of conical cells on petals and a nectary suggest flowers are pollinated by small insects with a secondary pollen presentation on the cupula of fertile stamens. Conclusions: We found that Ophiocaryon may be derived from ancestors that were similar to extant Meliosma in their flower structure and pollination mechanism. However, the lack of shared derived characters between Ophiocaryon and its phylogenetic sister group M. alba is puzzling and requires further investigations on the diversity of the latter species.

The morphophysiological dormancy in Amborella trichopoda seeds is a pleisiomorphic trait in angiosperms.

Background and Aims: Recent parsimony-based reconstructions suggest that seeds of early angiosperms had either morphophysiological or physiological dormancy, with the former considered as more probable. The aim of this study was to determine the class of seed dormancy present in Amborella trichopoda , the sole living representative of the most basal angiosperm lineage Amborellales, with a view to resolving fully the class of dormancy present at the base of the angiosperm clade. Methods: Drupes of A. trichopoda without fleshy parts were germinated and dissected to observe their structure and embryo growth. Pre-treatments including acid scarification, gibberellin treatment and seed excision were tested to determine their influence on dormancy breakage and germination. Character-state mapping by maximum parsimony, incorporating data from the present work and published sources, was then used to determine the likely class of dormancy present in early angiosperms. Key Results: Germination in A. trichopoda requires a warm stratification period of at least approx. 90 d, which is followed by endosperm swelling, causing the water-permeable pericarp-mesocarp envelope to split open. The embryo then grows rapidly within the seed, to radicle emergence some 17 d later and cotyledon emergence after an additional 24 d. Gibberellin treatment, acid scarification and excision of seeds from the surrounding drupe tissues all promoted germination by shortening the initial phase of dormancy, prior to embryo growth. Conclusions: Seeds of A. trichopoda have non-deep simple morphophysiological dormancy, in which mechanical resistance of the pericarp-mesocarp envelope plays a key role in the initial physiological phase. Maximum parsimony analyses, including data obtained in the present work, indicate that morphophysiological dormancy is likely to be a pleisiomorphic trait in flowering plants. The significance of this conclusion for studies of early angiosperm evolution is discussed.