ONTOGENETIC AND EVOLUTIONARY IMPLICATIONS OF A NEOTENOUS EXINE IN TAPEINOCHILOS (ZINGIBERALES: COSTACEAE) POLLEN

Tapeinochilos pollen, like that of most angiosperms, is spared by the standard acetolysis treatment because the sporoderm is impregnated with sporopollenin. This genus and its allies in the Costaceae are the only taxa in the eight families of Zingiberales that have acetolysis-resistant pollen. The sporoderm in most of the order is characterized by exine reduced to a wispy coating or layer with delicately anchored spinules and a highly elaborated intine. Ultrastructural studies on the pollen of Tapeinochilos reveal a pattern of wall development that is significantly different from the generalized angiosperm type; namely, there are no columellae, nor is there any significant accretion of sporopollenin following the dissolution of callose and release of microspores. The primexine is composed of rodlets which build up solidly between apertures and become packed into layers to form a thick, stratified exinous covering. No secondary exine develops during the free spore period and the juvenile primexine persists as the protective coat on the mature pollen grain. This pattern of pollen development is viewed as an example of neoteny in which a juvenile or immature character is retained in adulthood.     

ULTRASTRUCTURE AND DEVELOPMENT OF THE NEXINE AND INTINE IN THE POLLEN WALL OF SILENE ALBA (CARYOPHYLLACEAE)

Nexine and intine development in Silene alba (Caryophyllaceae) was investigated by electron microscopy and enzyme cytochemistry. Nexine-2 forms by deposition of sporopollenin along unit membrane lamellae closely associated with the microspore plasma membrane in the late tetrad stage. After the callose wall dissolves, electron density increases along the tangentially oriented fibers of the proximal primexine, forming nexine-1. When the exine is essentially complete, the intine begins to develop. In the nearly mature microspore, acid phosphatase activity appears in the peripheral cytoplasm just prior to its extrusion into the intine of the mature pollen grain.     

POLLEN WALL AND TAPETAL ORBICULAR WALL DEVELOPMENT IN SORGHUM BICOLOR (GRAMINEAE)

Pollen wall development in Sorghum bicolor is morphologically and temporally paralleled by the formation of a prominent orbicular wall on the inner tangential surface of the tapetum. In the late tetrad stage, a thin, nearly uniform primexine forms around each microspore (except at the pore site) beneath the intact callose; concurrently, small spherical bodies (pro-orbicules) appear between the undulate tapetal plasmalemma and the disappearing tapetal primary wall. Within the primexine, differentially staining loci appear, which only develop into young bacula as the callose disappears. Thus, microspore walls are devoid of a visible exine pattern when released from tetrads. Afterwards, sporopollenin accumulates simultaneously on the primexine and bacula, forming the exine, and on the pro-orbicules, forming orbicules. Channels develop in the tectum and nexine, and both layers thicken to complete the microspore exine. Channeled sporopollenin also accumulates on the orbicules. A prominent sporopollenin reticulum interconnects the individual orbicules to produce an orbicular wall; this wall persists even after the tapetal protoplasts degenerate and after anthesis. While the pollen grains become engorged with reserves, a thick intine, containing conspicuous cytoplasmic channels, forms beneath the exine. Fibrous material collects beneath the orbicular wall. The parallel development and morphological similarities between the tapetal and pollen walls are discussed.     

ISOLATION OF EXINES FROM GYMNOSPERM POLLEN

Exines of certain Gymnosperms spontaneously separate from the intine during the process of hydration preceding pollen germination. Exines of pollen of Calocedrus decurrens, Chamaecyparis lawsoniana, Juniperus occidentalis, Sequoia sempervirens, and Pseudotsuga menziesii were isolated from hydrated, autoclaved pollen. Free exines were purified by centrifugation on a discontinuous sucrose gradient of densities 1.14 to 1.27 g/ml. The outer intine dissolved on autoclaving. This method may be applicable to a wide range of genera. Purified exines are of potential use in chemical analyses of sporopollenin and in production of antibodies to exine.     

ULTRASTRUCTURE OF EXINE-LESS POLLEN: HELICONIA (HELICONIACEAE)

Pollen of Heliconia and many of its relatives in the Zingiberales is virtually devoid of a conspicuous, protective exine. The exine is relegated to a few spinules and a thin, electron-dense layer (0.08 μm), whereas the intine can be up to 100 times as thick (8 μm) and structurally complex. Comparative ultrastructural observations on the sporoderm and protoplast are presented for three species and discussion focuses on the possible adaptive significance of the elaborated intine and its potential usefulness in systematic studies.     

A new look at the acetolysis method

The acetolysis method intreduced byGunnar Erdtman is still a very welcome and highly successful technique in palynology. However, acetolysis destroys all pollen material with the exception of sporopollenin that forms the outer pollen wall, the exine. Modern palynology in its application to plant systematics and phylogeny must consider all sporoderm characters, not only those of the exine. The neglect of the intine may distort some principal palynological aspects. This is illustrated by cases of total breakdown or gross modification of thin exine structures (e.g. inBeilschmiedia, Strelitzia) and by the clarification of apertures (e.g.,Polyalthia, Fissistigma, Calluna). In our view the investigation of both acetolysed and non-acetolysed pollen is obligatory for a well balanced view of pollen structure and function.     

POLLEN GRAIN DEVELOPMENT AND TAPETAL CHANGES IN STRELITZIA REGINAE (STRELITZIACEAE)

The proexine that forms within the callosic envelope before the end of the microspore tetrad period is thick (about 1 μm) and exceptionally complex. It has components equatable with tectum, columellae, and a nexine that includes lamellar zones. All these components persist in the exine although late in development they become difficult to recognize because this exine is reduced in thickness, apparently by stretching, to a maximum of 0.2 μm. Strelitzia is an example of an exine template, with receptors for sporopollenin, that is not maintained during development. The Strelitzia microspore surface changes from an exine like that on an interaperture sector to the channeled intinelike system common for the apertures of pollen grains. The exine on sterile grains gives what may be a rare view of a stabilized immature exine. The mature exine on viable pollen grains resembles this early exine only in the most impressionistic way. Tapetal cells go through at least one cycle of hyperactivity, dedifferentiation, mitosis, and then again hyperactivity before they finally decline.     

Pollen wall development in Vigna vexillata I. Characterization of wall layers

Light and electron microscope observations characterized the layers that comprise Vigna vexillata L. pollen walls, and identified the timing of their development. Exine sculpturings form an unusually coarse ektexinous reticulum. The structure of the ektexine is granular; this differs from the columellate/tectate type of structure typical of most angiosperm pollen. The ektexine overlies a homogeneous-to-lamellar, electron-dense endexine, which in turn surrounds a thick, microfibrillar intine. Pollen grains are triporate and operculate, with Zwischenkörper and thickened intine underlying the apertures. The ektexine forms during the tetrad period of microspore development, the endexine and Zwischenkörper during the free microspore stage, and the intine during the bicelled (pollen) stage. Coarsely reticulate exine sculpturings and the granular structure of the patterned exine wall of the pollen grains are features that make this species suitable for detailed studies of pollen wall pattern formation.     

Pollen wall ontogeny in <Emphasis Type="Italic">Polemonium caeruleum</Emphasis> (Polemoniaceae) and suggested underlying mechanisms of development

By a detailed ontogenetic study of Polemonium caeruleum pollen, tracing each stage of development at high TEM resolution, we aim to understand the establishment of the pollen wall and to unravel the mechanisms underlying sporoderm development. The main steps of exine ontogeny in Polemonium caeruleum, observed in the microspore periplasmic space, are spherical units, gradually transforming into columns, then to rod-like units (procolumellae), the appearance of the initial tectum, growth of columellae in height and tectum in thickness and initial sporopollenin accumulation on them, the appearance of the endexine lamellae and of dark-contrasted particles on the tectum, the appearance of a sponge-like layer and of the intine in aperture sites, the appearance of the foot layer on the base of the sponge-like layer and of spinules on the tectum, and massive sporopollenin accumulation. This sequence of developmental events fits well to the sequence of self-assembling micellar mesophases. This gives (together with earlier findings and experimental exine simulations) strong evidence that genome and self-assembly probably share control of exine formation. It is highly probable that self-assembly is an intrinsic instrument of evolution.     

Sporopollenin monomer biosynthesis in arabidopsis

Land plants have evolved aliphatic biopolymers that protect their cell surfaces against dehydration, pathogens, and chemical and physical damage. In flowering plants, a critical event during pollen maturation is the formation of the pollen surface structure. The pollen wall consists essentially of the microspore-derived intine and the sporophyte-derived exine. The major component of the exine is termed sporopollenin, a complex biopolymer. The chemical composition of sporopollenin remains poorlycharacterized because it is extremely resistant to chemical and biological degradation procedures. Recent characterization of Arabidopsis thaliana genes and corresponding enzymes involved in exine formation has demonstrated that the sporopollenin polymer consists of phenolic and fatty acid-derived constituents that are covalently coupled by ether and ester linkages. This review illuminates the outlines of a biosynthetic pathway involved in generating monomer constituents of the sporopollenin biopolymer component of the pollen wall.     

THE EVOLUTION OF POLLEN TETRADS IN ONAGRACEAE

In Onagraceae, pollen is shed in mature tetrads in most Epilobieae, many species of Ludwigia (Jussiaeeae), and two closely related species of the large genus Camissonia (Onagreae). Mature tetrads of Camissonia cardiophylla and representative species of Epilobium and Ludwigia were examined with light, scanning, and transmission electron microscopes. Morphological diagnoses of monad units indicated that individual taxa could be readily distinguished. Statistical analyses of tetrads which remained after acetolysis treatment revealed significant differences in the strength of the binding mechanisms. Mechanisms of tetrad cohesion were found to consist of two principal types. Common to all taxa is cohesion of pollen wall surfaces at the aperture margins; this mechanism is well known in many angiosperm groups. With the exception of Camissonia, the remaining taxa also display binding by means of short exine fragments between adjacent pollen units. These fragments, termed bridges and reported here for the first time, are located in the area extending from the aperture margins to near the center of the proximal exine faces. Thin sections reveal that layers of the bridges are identical with those of the exine. Comparisons were made between bridges and viscin threads, both of which occur on the proximal faces of the grains. Viscin threads are present on all pollen grains in Onagraceae and exhibit distinctive morphologies, and bridges were viewed morphogenetically as related to viscin threads but including an endexine layer and occupying a position near the apertures where cohesion of wall surfaces also occurs. In an evolutionary sense, the formation of mature tetrads almost certainly occurred independently in Camissonia and may have done so in Ludwigia and the Epilobieae.     

SOLUBILITY OF POLLEN EXINES

The sporopollenin of pollen exines of Ambrosia trifida is soluble in fused potassium hydroxide, in strong oxidizing solutions, and in certain organic bases. It is insoluble in other organic and inorganic acids and bases, in lipid solvents, and in detergents. The outer exine layer of gymnosperm and angiosperm pollen dissolves in 2-aminoethanol. The inner exine layer, as well as the exine of pteridophyte spores, is insoluble. The exine dissolution process in 2-aminoethanol involves swelling and disintegration of exine structures, leaving some residual globules. Sporopollenin shares some solubility properties with lignin and cutin but appears to be chemically distinct from these substances.     

Pollen Tetrads of Hedycarya arborea J. R. et G. Forst. (Monimiaceae)

Hedycarya has pollen in permanent tetrads. H. arborea, the New Zealand species, differs from others studied, in having a cap of more or less imperforate tectum at the distal pole of each grain. This polar region is not an aperture and the pollen tube emerges through a papillose part of the external wall of each grain. Transmission electron microscope studies of immature and mature tetrads reveal a most unusual exine structure. "Radial processes" develop by accumulation of sporopollenin around unit membranes of similar dimensions to the plasmalemma, and extend from just beyond the intine to the tectal region. The entire exine is considered ectexinous. During development, members of a tetrad are interconnected by cytoplasmic channels and the synchronous division into generative and vegetative nuclei within each tetrad is attributed to their presence. The channels become closed by the deposition of intine. Comparisons are made with exine structure in some other members of the woody Ranales and with some other plants with tetrad pollen.     

The Apertural Wall in Pollen of Linum grandiflorum

The apertural wall in tricolpate pollen of Linum grandiflorumwas investigated in order to understand its functioning duringdesiccation and rchydration. Whole and sectioned pollen grainswere studied with light or electron microscopy and by cytochemicalmeans. The areas of the apertures were examined in fresh drypollen, in grains moistened on agar gel or removed from compatiblestigmas, and in pollen from mature undehisced anthers The intine was found to consist of an inner ß-glucanlayer and an outer pectic layer. At the apertures the pecticlayer is thickened and overlaid by a ß-glucan layer.The pectinaceous intine stains red with basic fuchsin. The presenceof a third wall layer, the medine, was not confirmed. The aperturalintine thickenings possess considerable imbibitional capacityand at rehydration they appear as swollen lenticular bodies A procedure is described for obtaining intact exine free grains(EFG's) and whole, separated exines of L. grandiflorum. Invariably,the released EFG's consisted of protoplasts encased in the cellulosicintine. In most grains the outer intine remained attached tothe separated exine In L. grandiflorum the outer wall of the aperture expands whilethe protoplast and endintine are still infolded. Apparently,the exintine becomes detached from the endintine during desiccationand re-attaches at rehydration. It is suggested that the transientdetachment controls the influx of water into the vegetativecell Except for morph-specific exine processes no differences instructure of the aperture wall or its functioning at rehydrationwere observed between pin and thrum grains Pollen wallM, apertures, exintine, exine free grains, rehydration, desiccation, Linum grandiflorum     

STUDIES OF SEED FERN POLLEN: DEVELOPMENT OF THE EXINE IN MONOLETES (MEDULLOSALES)

Monoletes pollen extracted from the seed fern synangium Dolerotheca sclerotica Baxter illustrate four stages in the development of the sporoderm. In the first stage the grains are up to 100 μm long and possess an apparent homogeneous exine in which there is little differentiation between the nexine and sexine. Numerous nexine lamellae and the initiation of sexine expansion mark stage 2 in exine ontogeny. Further expansion of the sexine continues in the third stage until the ratio between the nexine and sexine is approximately 1:5. The final stage in maturation of the sporoderm shows an expanded alveolate sexine with some of the sporopollenin units broken and disorganized. It is at this stage of development that nexine lamellae are most prominent. The formation of sporoderm layers in the fossil grains is compared with pollen grain development in living cycads (Cycadophyta) and a model proposed to account for the apparent early formation of nexine lamellae in Monoletes. The evolution of exine components in early pollen types is discussed.     

POLLEN MORPHOLOGY IN THE GENUS MIMULUS (SCROPHULARIACEAE) AND ITS TAXONOMIC SIGNIFICANCE

The pollen morphology of 117 species and varieties of Mimulus was examined by light and scanning electron microscopy. Five major and 8 more tentative, minor types were found based on the differential correlation of aperture type, exine morphology, pollen grain diameter and other characters: type 1—synaperturate, usually ±spiraperturate, exine perforate to microreticulate with supratectal processes; type II—trocolporate, exine microreticulate (IIa and IIb, supratectal processes absent; IIa, mean polar axis 16–19 μm; IIb, mean polar axis 25–35 μrn; IIc, supratectal processes present); type III—tricolpate, colpus membrane ±psilate. exine with supratectal processes (IIIa, exine microreticulate and 1.4–2.0 μm thick, polar axis ≥ 30 μm; 111b, exine densely perforate and 2.2–2.8 μm thick, polar axis ≤ 23 μm); type IV—tricolpate, colpus covered with spinulose granules (operculate), exine microreticulate with supratectal processes; type V—5–7 stephanocolpate (Va and Vb, colpus margins ±straight and nongranular; Va, exine microreticulate with supratectal spinules; Vb, exine perforate with supratectal spinules or spinulose verrucae; Vc, colpus margins ragged and granular, exine microreticulate with supratectal processes). The pollen data correlate well with geographical and macromorphological data and, where the latter are ambiguous, often provide important clues toward the resolution of conflicting interpretations of infrageneric classification and generic delimitation.     

云南松花粉形态研究

在云南松(Pinus yunnanensis Fr.)小孢子发生发育过程中,花粉母细胞、四分孢子及花粉粒均见有粘连现象。花粉气囊的形态、大小变化复杂多样。除一般具两个正常气囊的花粉粒外,还观察到气囊不发育、具一个气囊、二个异形气囊、三个气囊和四个气囊的花粉粒。成熟花粉壁从外至内可分为外壁外层、外壁内层、内壁外层和内壁内层,它们的构成成分及形态均有明显差别。贮存后花粉的内壁结构发生了明显变化。     

Exploding pollen in Montrichardia arborescens (Araceae)

Pollen grains of Montrichardia are inaperturate with psilate ornamentation. The pollen wall is formed by a thin ectexine and an extraordinarily thick intine. In living as well as in dead pollen grains contact with water leads to a rapid swelling of the intine followed by an explosive opening of the exine. Within a few seconds a thick tube is formed, which is not the pollen tube. The pollen protoplast is situated at the tip of the tube. These intine tubes are interpreted as pollen connecting tools to keep pollen grains together and adhere them to the cuticle of the hairless pollinators.     

Release of proteins from the pollen wall of Linum grandiflorum

Summary The emission of proteins from the pollen wall of Linum grandiflorum stained with Coomassie blue was followed directly in moistened grains as well as in pollen prints. Within the first minute of the grain being moistened exine-borne proteins emerged from both inter-apertural and apertural sites; subsequently, proteins of a different nature were discharged from the apertures only. In a fraction of the grains the release of intine proteins was not preceded by that of exine proteins. Pin and thrum pollen did not differ in terms of mode or site of this protein emission. The presence and emergence of exine proteins from the apertures is explained by the process of infolding of the colpal wall at desiccation and its expansion at rehydration, which causes an initial trapping and subsequent re-exposure of surface materials. This explanation may also account for the occurrence of poral sporophytic proteins in the pollens of many dictoyledons.     

Some characteristics of pollen wall cytochemistry and ultrastructure in Japanese larch (Larix leptolepis Gord.)

Summary The mature pollen of Larix leptolepis Gord. (Conifer) contains five different cell types, and the plasma membrane of the vegetative cell is continuous and organized. The pollen wall is composed of two morphologically and cytochemically distinct domains: the exine and the intine. In the multilayered exine, the ektexine appears granular and the endexine, lamellar. The intine is thick and bilayered with a microfibrillar structure occupying its inner portion. Cytochemical reactions of the exine and the intine are similar to those found in angiosperms. Pollen wall involvement in the male female recognition system is discussed with respecl to the angiosperms.