Pollen wall development in flowering plants

The outer pollen wall, or exine, is more structurally complex than any other plant cell wall, comprising several distinct layers, each with its own organizational pattern. Since elucidation of the basic events of pollen wall ontogeny using electron microscopy in the 1970s, knowledge of their developmental genetics has increased enormously. However, self-assembly processes that are not under direct genetic control also play an important role in pollen wall patterning. This review integrates ultrastructural and developmental findings with recent models for self-assembly in an attempt to understand the origins of the morphological complexity and diversity that underpin the science of palynology.     

Pollen wall development in Hypecoum imberbe Sm. (Fumariaceae)

We have studied the ontogeny of the pollen wall of Hypecoum imberbe. Our initial observations with conventional transmission electron microscopy (TEM) were inconclusive due to the similarities found in the electrodensity of the foot layer and the endexine, and between this latter layer and the intine. Thus we applied the PTA acetone histochemical test in order to differentiate between the cell wall components. This method proved to be efficient in resolving the differences between the layers and even allowed us to distinguish two strata within the narrow intine layer. A thin foot layer can be distinguished only by the temporary presence of a single white line separating it from the initial deposition of the endexine. The aperture consists of two colpi with no special differentiation. The tapetum is a typical secretory type giving rise to elaioplasts and tapetosomes during development, which persist as individual organelles upon the degradation of the tapetum. As the ultrastructural organisation of the sporoderm seems to offer little protection to the gametophyte, we finally discuss how the structure of the pollen sporoderm might be related to the special morphological characteristics of a flower, its habitat, and the biology of the plant as a whole.     

Pollen wall development in Cryptomeria japonica (Taxodiaceae)

Pollen wall development in Cryptomeria japonica was observed by scanning and transmission electron microscopy. The pollen of C. japonica is characterized by a non-saccate, projecting papilla. The exine of C. japonica consists of the outer granular ectexine and the inner lamellated endexine. At the tetrad stage, the initial granular layer of the pro-ectexine first forms on the microspore plasma membrane. The tripartite lamellae of the pro-endexine form under the pro-ectexine. The prosporopollenin material is deposited on the pro-ectexine and pro-endexine at the free spore stage. The ectexine granule increases its volume and the endexine lamellae thicken. The papilla protrudes during the tetrad stage. The tip of the papilla bends laterally where the exine is thinner. Exine construction in C. japonica is similar to that of Cunninghamia; however, the amount and size of the granular ectexine and lamellated endexine differ. The conspicuous papilla protrudes and bends during the tetrad period.     

Pollen wall development in Malvastrum corchorifolium (Malvaceae)

The Malvaceae family has many species that are the subject of systematic controversy. Its broad spectrum of fruit and pollen morphology has challenged taxonomists because often morphological studies lead to different conclusions. Therefore, this study of selected stages of pollen wall development in Malvastrum corchorifolium is offered as another approach to verify its current taxonomic assignment and assist in the clarification of malvacean ancestry. Aperture number, endexine thickness and the presence of basal cushions are features that concur with its present placement in the Abutileae tribe, Sidinae subtribe and genus Malvastrum. Its advanced spine morphology, long spines with pointed apices and prominent basal cushions, suggest that Malvastrum corchorifolium is highly specialized and evolutionarily advanced.     

Pollen wall development in Sciadopitys verticillata (Sciadopityaceae)

Pollen wall development of Sciadopitys verticillata was observed by transmission electron microscopy. The pollen of S. verticillata is non-saccate and spherical, and the exine consists of the outer thick, sculptured ectexine and the inner lamellated endexine. At the early tetrad stage, the initial ectexine and lamellae of the initial endexine begin to form on the microspore plasma membrane. The ectexine granules gradually swell. Deposition of sporopollenin materials on the ectexine granules then results it their becoming partially connected to each other. Identification of the original small ectexine granules then becomes difficult, and, finally, the ectexine appears as a homogeneous, partially discontinuous layer. The granules of the early ectexine cannot be identified. At maturity, there are four to five endexine lamellae. Recent molecular data have shown that Sciadopitys first branches off from the Cupressaceae plus Taxaceae clade, which is characterized by granular exine. Although the ectexine of Sciadopitys is similar to that of the Cupressaceae during initial development, the morphology of the ectexine is significantly different in the mature pollen. The initial stage of pollen development clearly shows the structural homology of the granular ectexine. Divergence of the exine structure occurs in the later stages.     

Construction of an SSR and RAD-Marker Based Molecular Linkage Map of Vigna vexillata (L.) A. Rich

Vigna vexillata (L.) A. Rich. (tuber cowpea) is an underutilized crop for consuming its tuber and mature seeds. Wild form of V. vexillata is a pan-tropical perennial herbaceous plant which has been used by local people as a food. Wild V. vexillata has also been considered as useful gene(s) source for V. unguiculata (cowpea), since it was reported to have various resistance gene(s) for insects and diseases of cowpea. To exploit the potential of V. vexillata, an SSR-based linkage map of V. vexillata was developed. A total of 874 SSR markers successfully amplified single DNA fragment in V. vexillata among 1,336 SSR markers developed from Vigna angularis (azuki bean), V. unguiculata and Phaseolus vulgaris (common bean). An F₂ population of 300 plants derived from a cross between salt resistant (V1) and susceptible (V5) accessions was used for mapping. A genetic linkage map was constructed using 82 polymorphic SSR markers loci, which could be assigned to 11 linkage groups spanning 511.5 cM in length with a mean distance of 7.2 cM between adjacent markers. To develop higher density molecular linkage map and to confirm SSR markers position in a linkage map, RAD markers were developed and a combined SSR and RAD markers linkage map of V. vexillata was constructed. A total of 559 (84 SSR and 475 RAD) markers loci could be assigned to 11 linkage groups spanning 973.9 cM in length with a mean distance of 1.8 cM between adjacent markers. Linkage and genetic position of all SSR markers in an SSR linkage map were confirmed. When an SSR genetic linkage map of V. vexillata was compared with those of V. radiata and V. unguiculata, it was suggested that the structure of V. vexillata chromosome was considerably differentiated. This map is the first SSR and RAD marker-based V. vexillata linkage map which can be used for the mapping of useful traits.     

Pod hairs as a factor in Vigna vexillata resistance to the pod-sucking bug, Clavigralla tomentosicollis

Two varieties of Vigna vexillata Benth proved more highly resistant, both under field (free-choice) and screenhouse (no-choice) conditions, to infestation and damage by the pod-sucking bug Clavigralla tomentosicollis (Stål.) (Hemiptera: Coreidae) than cowpea (Vigna unguiculata Walp.) varieties. Trichomes on the pod of V. vexillata partly accounted for the resistance. However, other factors in the pod or seeds are involved in the resistance. Because the pod wall of V. vexillata is not tougher than that of cowpeas, biochemical characteristics of pods and seeds of V. vexillata will be further investigated.

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Résumé Deux variétés de Vigna vexillata se sont avérées plus résistantes aux infestations et dégâts causés par la punaise des cosses Clavigralla tomentosicollis (Stål.) (Hemiptera: Coreidae) que les variétés de Vigna unguiculata, tant en champ (condition de libre choix) qu'en serre (sans choix possible). Les trichomes des cosses de V. vexillata sont en partie responsables de cette résistance. Cependant d'autres facteurs de la cosses ou des fèves sont impliqués dan la résistance. Comme la paroi de la cosse de V. vexillata n'offre pas plus de résistance mécanique que celle de V. unguiculata, les études futures se porteront sur les caractéristiques biochimiques des cosses et des fèves de V. vexillata.

     

Pollen wall development in Eucommia ulmoides (Eucommiaceae)

In Eucommia the foot layer plays a prominent part in microspore wall development. Bacules (>100 nm) are rods originating within the foot layer. Small bacules (diameter ca. 10 nm) form at the same time as the foot layer. The tectum is considered to be made up of these microbacular rods. Spinules ar continuous with the bacules. An endexine is differentiated during a middle to late microspore stage. Except for the pore the furrow includes tectum and foot layer on the endexine. Since the furrow consists of a region of reduced foot layer and the reduction is gradual near the polar ends of furrows, assessment of furrow length depends to some extent upon variations in exine infolding. The pore is well defined, but, because it is crossed by lamellations of the endexine and foot layer and often overlaid by tongues and bridges of foot layer plus tectum (including spinules) it is obscured from view using either light microscopy or scanning electron microscopy.     

Pollen wall development: the associated enzymes and metabolic pathways

Pollen grains are surrounded by a sculpted wall, which protects male gametophytes from various environmental stresses and microbial attacks, and also facilitates pollination. Pollen wall development requires lipid and polysaccharide metabolism, and some key genes and proteins that participate in these processes have recently been identified. Here, we summarise the genes and describe their functions during pollen wall development via several metabolic pathways. A working model involving substances and catalytic enzyme reactions that occur during pollen development is also presented. This model provides information on the complete process of pollen wall development with respect to metabolic pathways.     

Pollen wall and tapetum development in Plantago major L. (Plantaginaceae): assisting self-assembly

We have attempted to elucidate the underlying mechanisms of sporoderm development and pattern determination in Plantago major through a detailed ontogenetic study, using transmission electron microscopy (TEM) and scanning electron microscopy (SEM). We aim to compare our observations and interpretation with those on other species. Our study of sporoderm development in Plantago from the early tetrad stage to mature pollen grains has shown that pure physical processes, including self-assembly, which are not under direct genetic control, play an important role and represent evidently one of the instruments of evolution. Our observations fit well with the sequence of self-assembling micellar mesophases and show reiteration of some of them, confirming our self-assembly hypothesis. Some attention was also paid to the possible role of rough and smooth endoplasmic reticulum in the cortical cytoplasm of the developing microspores. The tapetum and Ubisch bodies development are also traced. The importance of detailed ontogenetic studies for understanding the establishment of complex pollen walls in any species and for understanding mechanisms underlying sporoderm development was demonstrated. We also present a simulation, obtained in vitro experiments by self-assembly, mimicking pollen grain of Plantago major. It is clear that, in pollen wall development, biological processes and purely physical factors work in tandem.     

Cell wall assembly in fucus zygotes: I. Characterization of the polysaccharide components

Fertilization triggers the assembly of a cell wall around the egg cell of three brown algae, Fucus vesiculosus, F. distichus, and F. inflatus. New polysaccharide polymers are continually being added to the cell wall during the first 24 hours of synchronous embryo development. This wall assembly involves the extracellular deposition of fibrillar material by cytoplasmic vesicles fusing with the plasma membrane. One hour after fertilization a fragmented wall can be isolated free of cytoplasm and contains equal amounts of cellulose and alginic acid with no fucose-containing polymers (fucans) present. Birefringence of the wall caused by oriented cellulose microfibrils is not detected in all zygotes until 4 hours, at which time intact cell walls can be isolated that retain the shape of the zygote. These walls have a relatively low ratio of fucose to xylose and little sulfate when compared to walls from older embryos. When extracts of walls from 4-hour zygotes are subjected to cellulose acetate electrophoresis at pH 7, a single fucan (F₁) can be detected. By 12 hours, purified cell walls are composed of fucans containing a relatively high ratio of fucose to xylose and high levels of sulfate, and contain a second fucan (F₂) which is electrophoretically distinct from F₁. F₂ appears to be deposited in only a localized region of the wall, that which elongates to form the rhizoid cell. Throughout wall assembly, the polyuronide block co-polymer alginic acid did not significantly vary its mannuronic (M) to guluronic (G) acid ratio (0.33-0.55) or its block distribution (MG, 54%; GG, 30%; MM, 16%). From 6 to 24 hours of embryo development, the proportion of the major polysaccharide components found in purified walls is stable. Alginic acid is the major polymer and comprises about 60% of the total wall, while cellulose and the fucans each make-up about 20% of the remainder. During the extracellular assembly of this wall, the intracellular levels of the storage glucan laminaran decreases. A membrane-bound β-1, 3-exoglucanase is found in young zygotes which degrades laminaran to glucose. It is postulated that hydrolysis of laminaran by this glucanase accounts, at least in part, for glucose availability for wall biosynthesis and the increase in respiration triggered by fertilization. The properties and function of alginic acid, the fucans, and cellulose are discussed in relation to changes in wall structure and function during development.     

Characterization of the carbohydrate component of fraction I in the Neurospora crassa cell wall.

The carbohydrate portion of fraction I of the Neurospora crassa cell wall has been analyzed for sugar composition by gas-liquid chromatography and colorimetric methods. The analysis was performed comparatively in a wild-type strain (RL 3-8A) and three morphological mutants: scumbo (FGSC 49), peak-2a (a mutant known to be allelic to biscuit), and ragged (FGSC 296). Fraction I of all strains studied contains glucose, mannose, and galactose as the main sugars. Uronic acids and amino sugars are also present in small amounts. The glycosidic linkages binding the neutral sugars were analyzed by Lindberg's combined gas chromatography-mass spectrometry techniques for identification of the partially methylated alditol acitate sugar derivatives. The main polymeric portion of fraction I seems to be a linear glucan with the glucose residues linked by 1 leads to 3 and 1 leads to 4 bonds. A mannan portion with a branched configuration is also present, with galactose as the sugar residue which serves as branches in the molecule(s). The branched mannan portion appears to increase in amount in correlation with more drastic morphological changes of the mycelia. In this respect, the mutant ragged has the lowest mycelial growth rate and the largest amount of mannan. The importance of the polysaccharide structure of fraction I on the colonial morphology of the mycelia is discussed.     

Pollen development in orchids

Summary Cytokinesis in microsporocytes of moth orchids is unusual in that it occurs simultaneously after meiosis, the cytoplasm does not infurrow in the division planes, and cell plates are deposited in association with centrifugal expansion of phragmoplasts. Microtubules radiating from the nuclear envelopes appear to be of fundamental importance in establishment of division planes. Primary interzonal spindles develop between sister nuclei and interaction of radial microtubules triggers development of secondary interzonal spindles between non-sister nuclei. From three to six or more phragmoplasts, depending upon the arrangement of nuclei in the coenocyte, develop from these postmeiotic arrays. The phragmoplasts consist of co-aligned microtubules and F-actin organized into bundles that are broad proximal to the mid-plane and taper distally. Ultrastructure of the phragmoplast/cell plate reveals that abundant ER is associated with vesicle aggregation and coalescence. Cell plates are deposited in association with phragmoplasts as they expand centrifugally to join the parental wall and/or fuse with one another in the interior of the cell.Abbreviations CLSM confocal laser scanning microscope/microscopy - FITC flnorescein isothiocyanate - PPB preprophase band of microtubules - TEM transmission electron microscope/microscopy     

Rice caryopsis development I: Dynamic changes in different cell layers

Rice caryopsis as one of the most important food sources for humans has a complex structure that is composed of maternal tissues including the pericarp and testa and filial tissues including the endosperm and embryo. Although rice caryopsis studies have been conducted previously, a systematic characterization throughout the entire developmental process is still lacking. In this study, detailed morphological examinations of caryopses were made during the entire 30‐day developmental process. We observed some rapid changes in cell differentiation events and cataloged how cellular degeneration processes occurred in maternal tissues. The differentiations of tube cells and cross cells were achieved by 9 days after pollination (DAP). In the testa, the outer integument was degenerated by 3 DAP, while the outer layer of the inner integument degenerated by 7 DAP. In the nucellus, all tissues with the exception of the nucellar projection and the nucellar epidermis degenerated in the first 5 DAP. By 21 DAP, all maternal tissues, including vascular bundles, the nucellar projection and the nucellar epidermal cells were degenerated. In summary, this study provides a complete atlas of the dynamic changes in cell differentiation and degeneration for individual maternal cell layers of rice caryopsis.     

Pollen ultrastructure in anther cultures of Datura innoxia. II. The generative-cell wall.

In young pollen grains of Datura innoxia, a wall of the usual hemispherical type separates the 2 gametophytic cells initially and, in the electron microscope, appears as an electron-translucent matrix which is contiguous with the intine. Before detachment of the generative cell from the intine, the matrix decreases in thickness and in places is dispersed altogether leaving the plasmalemmae on either side of it in close apposition. A particularly prominent zone, triangular in profile, is left where the wall joins with the intine. After detachment of the cell, remnants of the matrix can be seen distributed irregularly around the cell and it is supposed that these are partly derived from material in the triangular zone as the cell is drawn away from the intine. The wall residues persist throughout the maturation phase of the pollen and are considered to be either callose resulting from incomplete digestion of the initial wall, or some other polysaccharide material which is unevenly laid down along the wall and concentrated at the junction with the intine. In pollen induced into embryogenesis by anther culture, wall material is also distributed irregularly around the detached cell in a series of discrete zones, but these are more extensive than in vivo, closer together and in many instances highly dilated. The wall profiles thus have a beaded appearance, the 'beads' being connected together by short links of the 2 apposed plasmalemmae. The contents of the swollen zones have a similar electron density to that of the matrix in vivo but also show traces of a fibrillar component. It is postulated that this unusual swelling is a prelude to dispersal of the wall by disruption of the plasmalemmal links and to the establishment of cytoplasmic continuity between the 2 cells. The significance of such binucleate pollen grains in the formation of non-haploid embryos is discussed.     

Cell wall storage polysaccharides in cotyledons ofLupinus angustifolius L. I. Deposition during seed development

Summary Cotyledons of developing seeds ofLupinus angustifolius cv. Unicrop were examined by light and electron microscopy from 14 days after anthesis until maturity. Cell wall storage polysaccharides are deposited as secondary wall in the mesophyll cells from about 30 days after anthesis. Wall thickness increases from 0.2 to 20 m except in the pit areas around the plasmodesmata. Concentric layers within the secondary wall were revealed following staining with periodic acid-Schiff's reagent and with calcofluor white. Layers are seen as alternating bands of closely- and loosely-packed fibrils in the electron microscope. During maturation, these layers become compressed and radial striations appeared. During wall thickening, dictyosome vesicles contain fibrils of carbohydrate material which is apparently discharged into the periplasm. Evidence strongly suggests that the Golgi apparatus is active in wall deposition. Protein and lipid reserves fill the mesophyll cells at maturity. Starch which was abundant during development is present only in trace amounts in the cotyledons of mature seeds.This work was carried out at the former ARC Unit of Developmental Botany, Cambridge.