John R. Rowley

The fundamental structure of the pollen exine

The “fundamental” structure, the groundwork of the exine, is a three-dimensional network recoverable from exines of pteridophyte spores and the pollen of gymnosperms and angiosperms following many different degrading methods. It can also be derived from fossil exines and untreated exines during early stages of microspore development as well. The dimensions of these networks are commonly about 70 nm measured from the centre of one lumen to the centre of the next with individual lumina being about 40 nm in diameter. My approach here is to consider the interaction between this three-dimensional network and both radially arranged microchannels and laterally arranged white-line-centred lamellations and what it can suggest to us about substructural arrangement within exines. — Through oxidation microchannels can be hollowed out to diameters of 40–70 nm indicating that three-dimensional networks are superpositioned around microchannels. — White-line-centred lamellations have two features of exceptional interest with respect to how they pass through the three-dimensional network. They are both wider than the meshes of the network and apparently come and go during development. I consider the endoaperture of Epilobium to be a useful model in this interpretation. Exine unit structures are attached to either side of white-line-centred lamellations in these endoapertures; using this system as a model I suggest that white-line lamellations can be junction planes between units structures. The important feature of my interpretation is that all substructures of exine units take part in the white line structure including those appearing after partial degradation of the exine as a 3-D network. In this model white-line lamellations can be transposed into subunits of rod-shaped unit structures and the reverse. Sketches of this reversible system were prepared for the Thanikaimoni memorial volume of the Journal of Palynology.

Substructure in exines of Artemisia vulgaris (Asteraceae)

Abstract Unit and subunit structure is exposed in exines of Artemisia vulgaris after partial oxidation methods which include 2-aminoethanol followed by exine expansion and exposure to potassium permanganate. Tectal bacules traced ontogenetically from a plasma membrane—glycocalyx unit-complex (“tuft”) consist of a super-coiled helical subunit (binder) wound around several axially straight helical subunits (core subunits). Fracturing occurs between tuft units when partial oxidation includes contraction of exines in HCL or ethanol prior to potassium permanganate. Tuft units are radial in the non-tectal bacules and throughout the nexine. Partial oxidation was controlled to give an end-product having a reduced although sufficient presence of sporopollenin to conserve the diagnostically taxon-distinct exine. The sporopolleninous remnant resisted acetolysis although without support (SiO) exines disintegrated into helical subunit-sized structures, transparent to electrons and unreactive to stains (including osmium tetroxide). Components of the partially oxidized exine having a binder and core unit structure, stain positively for acid polyanions (polysaccharides?) and protein. Osmophilia (unsaturated lipids + ?), extinguished during potassium permanganate treatment, had been located largely in what is interpreted to be the core region of tuft units.

A model of exine substructure based on dissection of pollen and spore exines

Abstract The exine according to our model, consists of filamentous subunits (∼ 15–40nm in diam.), each containing an axially orientated tubule (∼ 10–15nm in diam.) with numerous lateral branches. Relatively few lateral branches are exposed at the surface of exinous subunits so that most of the polysaccharide‐, protein‐, and lipid‐containing tubular complex embedded within the sporopolleninous subunit is protected from degradation and unavailable to stains or ions that could cause molecules of the complex to swell or contract. In the intact exine, the few branches of the tubular complex (glycocalyx units) that are exposed at surfaces of subunits may affect exine size and staining, but their effect is not discernible from exogenous and nonstructural endogenous substances in the microcapillary space between exinous subunits. The existence of glycocalyx units is evident following treatment that etches the sporopollenin from exinous subunits (and eliminates exogenous and endogenous substances), without inactiv...

Are the endexines of pteridophytes, gymnosperms and angiosperms structurally equivalent?

Abstract Substructures have the same size and configuration, like wire-wound springs, in all parts of the exine of Artemisia (tectum, bacules, foot layer and the endexine), and similar substructures are also present in all parts of the exospore of Lycopodium . Substructures like these are illustrated here for pollen exines of Poa, Betula (endexine), and Fagus as well. The generalized radial orientations of these substructures are at right angles to the “lamellations” common in endexines and in exospores. In the ectexine of angiosperm pollen the substructures are organized into unit structures (tufts) that are 70–200 nm or more in diameter. The tufts are structurally like plasmodesmata of tapetal cells and like viscin threads. In pollen of angiosperms and gymnosperms there is at some stages of development a white line at the junction between units of the ectexine and the endexine. The exine substructures cross the ectexine-endexine junction, and it can be assumed that exine units are continuous across these zones. I would say that endexine operates like the exospore of pteridophytes. According to my interpretation, the endexine opens up and closes off large irregular channels functioning as a pumping system. I consider the arrangement, size and appearance of endexine and exospore substructures to be similar in pteridophytes ( Lycopodium ), gymnosperms and angiosperms. Even if they are shown definitively to be structurally the same the distinctive term “exospore” ought to be retained because of the development and exclusive nature of the exospore.

Fibrils, microtubules and lamellae in pollen grains

Abstract Microtubules 24 mμ in diameter were seen in the cytoplasm of Populus tremula microspores near the time of microspore mitosis and intine formation. The microtubules ran between the plasma membrane and the nuclear envelope, where they appeared to attach to annuli of nuclear pores. Except for the nexine 2 and large spines, the exine of Nuphar luteum contained tubular fibrils oriented generally perpendicular to the cell surface in late microspore stages. On end the fibrils were pentagonal, 7–9 mμ in diameter, with a dense wall about 2 mμ in thickness, and a low dense central area 2.5–4 mμ in diameter. The dense wall of the fibrils was composed of about five subunits, each on the order of 2 mμ in diameter. In surface view the fibrils appeared as striated or twisted rods. In the small spines there is a direct relationship between the area of the spinules in section and the number of fibrils. The diameter of fibrils is minimal in spinules and increases toward the tectum and nexine 1 from 7–9 mμ to 10–18 mμ. The added material appeared granular (ca.2 mμ in diameter) when fibrils were seen in transverse section. The pentagonal shape was retained in larger fibrils which suggests that the increase in size is brought about by addition of material to specific sites on the fibrils, perhaps like certain helical viruses, as for example tobacco mosaic virus. The affinity of the fibrils for electron stains is like that of protein and nucleoprotein rather than sporopollenin, at least in its homogeneous form. In Anthurium some of the exine forms on lamellae of unit membrane dimensions. Sporopollenin accumulates on such membranes (often asymmetrically) to form the annular lamellae of the germinal apertures. The annular lamellae become 0.4 μ or more in thickness but retain a line of low density characteristic of the unit membrane (ca.4 mμ in diameter) until near maturity.

Developmental origins of structural diversity in pollen walls of Compositae

Compositae exhibit some of the most complex and diverse pollen grains in flowering plants. This paper reviews the evolutionary and developmental origins of this diversity in pollen structure using recent models based on the behaviour of colloids and formation of micelles in the differentiating microspore glycocalyx and primexine. The developmental model is consistent with observations of structures recovered by pollen wall dissolution. Pollen wall diversity in Compositae is inferred to result from small changes in the glycocalyx, for example ionic concentration, which trigger the self-assembly of highly diverse structures. Whilst the fine details of exine substructure are, therefore, not under direct genetic control, it is likely that genes establish differences in the glycocalyx which define the conditions for self-assembly. Because the processes described here for Compositae can account for some of the most complex exine structures known, it is likely that they also operate in pollen walls with much simpler organisation.

Formation of Pollen Exine Bacules and Microchannels on a Glycocalyx

Abstract In Epilobium and Chamaenerion a glycocalyx, containing mucopolysaccharides, is present between the plasma membrane and outer surface of the tectum during the microspore period. The glycocalyx is built up by rod-shaped units. The orientation of these rods is roughly perpendicular to the cell surface although their traces are vermiculate. The early accumulation of exine material occurs between the rod-shaped regions containing mucopolysaccharide material. Eventual encirclement of the rods with exine material results in the formation of cylindrical channels, of extremely uniform size, common to most Onagraceous pollen exines. Although use of the terms primexine matrix or template exine is avoided in this report, they are unquestionably useful and can be applied in the absence of the histochemical data necessary for defining a glycocalyx. They delimit an ontogenetic period and a space, however, whereas a glycocalyx might be expected to be present throughout development and across the entire sporoderm...

Sporoderm ontogeny in Cabomba aquatica (Cabombaceae)

Abstract The study on Cabomba aquatica sporoderm ontogeny was undertaken to reveal the principles of development, their connection with the exine substructure and the resistance of the sporopollenin part of the exine to degradation. Procolumellae appear as the first elements of the exine at early tetrad stage, located in the plasma membrane glycocalyx. These procolumellae correspond to elementary exine units (tufts), by Rowley. Very soon after this, procolumellae are seen consisting of several tufts and turn into columellae. Then the grouping of the columellae occurs: sporopollenin accumulates at the proximal parts of the columellae, uniting several of them. Simultaneously a thin and interrupted tectum is introduced, and the columellae grow in height. The process of grouping of individual columellae persists at the stage of the disintegrating tetrad, so that the diameter of a new-formed complex columellae has increased 3 times. The complex columellae consist of clumps of columellae. The initial striae appear in the tectum at an early free microspore stage. Simultaneously, the foot layer as a very thin layer appears. In the course of further development some striae increase in volume (macro-striae), whereas the groups of the striae between them preserve their initial size (micro-striae). Sporopollenin Acceptor Particles (SAPs) appear on the surface of the striae. The diameter of the complex columellae is 6 times more than of procolumellae, the height of the exine increases 7–8 times. The individual columellae, which are the constructive elements of each complex columella, support adjacent supratectal striae and determine the formation of them. In each depression between the striae a row of radially oriented channels penetrates the tectum. These macro-channels are rather wide – unlike micro-channels which penetrate the columellae and the foot layer in different directions. At the stage of microspore cell vacuolisation, when the intine is introduced, the volume of the microspores increases and the exine is stretched out, the complex columellae are pulled apart and can be fragmented to pieces. The oxidative treatment with potassium permanganate brings about the etching of the superficial sporopollenin layer on the surface of the columellae and reveals a spirally arranged cluster of SAPs, the latter becoming a basis for sporopollenin accumulation in the form of a super-spiral around the complex columellae. These data give evidence of SAPs to be responsible for the growth of the elements of the exine by precisely located accumulation of receptor-dependant sporopollenin. The substructure of the exine – tuft units – becomes also evident as a result of oxidative treatment. These are revealed mainly in the tectum as the basic units of micro- and macro-stria. The binder parts of the tuft units resist degradation, but core zones do not. This gives evidence of the secondary nature of the sporopollenin deposited inside the tufts, whereas the binder zone of them seems to be the location of the primary accumulated, receptor-dependant sporopollenin.

Microchannels in the pollen grain exine

Abstract Direct evidence for the location of microchannels within larger diameter structures in modern pollen grain exines is provided by selective oxidation of the exine bordering the microchannels. These large structures form a three‐dimensional network that can be seen throughout exines in all transmission electron microscope studies involving experimental degradation of the exine in extant taxa and also in some fossil exines. From our results we conclude that components forming this reticulated pattern must be aligned around channel‐containing structures and that the superpositioning of the network of loops can be expected to be mostly radial in orientation, like the microchannels. Our approach was to oxidize the exines of Betula verrucosa and Chenopodium album after first causing the exines either to expand or contract. In expanded exines microchannels were etched out, creating cylindrical cavities 40–50 nm in diameter and the entire exine became highly acceptable to staining. Compaction of exines re...

Pollen wall of Ephedra foliata

Abstract The pollen wall of Ephedra foliata was examined by LM, SEM and TEM. The pollen wall consists of three layers: ectexine, foot layer and endexine. The ectexine has a solid tectum underlain by infratectal processes. The infratectal components appear granular in dried grains but there are columellae‐like rods in fresh fixed grains that were not subject to dehydration and the elevation of ridges. The Ephedra foot layer is part of the ectexine and has the same contrast. There is a white line lamellation between the foot layer and outer component of the endexine that is probably a junction plane between units of the foot layer and the endexine. The endexine consists of many cylindrical sheets that extend parallel with the long axis of the pollen grain. These sheets appear interconnected and may be branched or recurved in some sites. In pollen that had been acetolyzed the endexine was stained lighter than the ectexine, the reverse of most fresh grains. This is due to the endexine being porous and the mat...