Incorporation of Label into Pollen Tube Walls from Myoinositol-labeled Lilium longiflorum Pistils

Compatible and incompatible pollen tubes growing on detached Lilium longiflorum pistils which had been prelabeled with myoinositol-U-(14)C take up a portion of the label and utilize it for biosynthesis of tube wall substance. The label is transferred from pistil to pollen tubes apparently via the secretion products (exudate) of the pistil. The exudate thus appears to have a major nutritional role in pollen tube growth in vivo.     

FURTHER CYTOLOGICAL STUDIES OF A PERIODIC ACID-SCHIFF'S SUBSTANCE IN THE OVULES OF PASPALUM ORBICULARE AND P. LONGIFOLIUM

A periodic acid-Schiff's substance present in the micropylar end of the ovules of Paspalum orbiculare and P. longifolium was further studied by light and electron microscopy of glutaraldehyde-osmium-fixed and freeze-substituted, osmium-fixed tissues. The PAS substance is water soluble and is found in intercellular spaces between the nucellus and inner integument, the inner and outer integuments, the outer integument and ovary wall, and in the micropyle. Structurally the substance consists of fibrils embedded in a dense, amorphous matrix and may be associated with membranous structures in special layers between the plasmalemma and the cell wall in nucellar and integumentary cells. Part of the water soluble substance is believed to be secreted from the nucellar and integumentary cells. A large amount of this substance may be formed as a result of the dissolution of about one third of the distal micopylar portion of the outer integument prior to anthesis. Many of the electron-dense fibrils seem to be fibrillar intercellular substances and others appear to originate from the cell walls, including the cuticle. Both the matrix and the fibrils may be chemically heterogeneous and together form a mucilagenous substance which may facilitate the final growth of pollen tubes in these two species.     

How to determine the susceptibility of Meligethes aeneus to neonicotinoids?

Three different bioassays were used to determine the susceptibility to the neonicotinoid Biscaya^® of oilseed rape pollen beetles collected from fields in Bavaria. The one in which the test substance was applied to the inner wall of glass tubes is recommended for future studies on pollen beetles because it is not dependent on the availability of plant material and provides precise information on the amount of insecticide required per unit area.     

A model for the mechanism of tip extension in pollen tubes

Three main mechanisms are proposed to account for the tip growth of pollen tubes. (1) The tip region is supported against the internal osmotic pressure of the cell by a fibrillar network, composed mainly of microfilaments, that is stabilized by calcium ions. Tip extension is promoted by a lowering of the local cytoplasmic calcium ion concentration, through uptake by the mitochondria and/or endoplasmic reticulum, which leads to a weakening of the fibrillar network. (2) Vesicles, derived from dictyosomes in the main body of the tube, fuse with the apical plasma membrane, providing new membrane and further carbohydrate for the wall. The rate of fusion is proportional to the rate of diffusion of calcium ion across the plasma membrane at the tip. (3) The callose lining present in the pollen tube wall, except at the tip, renders the wall impermeable and restricts entry of calcium ions to the apical plasma membrane. This restriction limits the rate of vesicle fusion, and tube growth, to the tip.This model is discussed in the light of previous observations on the growth and structure of pollen tubes under normal and experimental conditions.     

GROWTH OF POLLEN TUBES OF OENOTHERA ORGANENSIS THROUGH OTHERWISE INCOMPATIBLE STYLES

Hecht , A. (Washington State U., Pullman.) Growth of pollen tubes of Oenothera organensis through otherwise incompatible styles. Amer. Jour. Bot. 47(1) : 32—36. Illus. 1960.–It has previously been reported that pollen tubes of Oenothera organensis fail to grow from compatible stylar tissue into a graft of stylar tissue containing the same “S” alleles as are present in the pollen tubes. Upon this basis it has been assumed that the incompatibility reaction in this species is equally effective at all levels of the stigma and style. Results contrary to these were obtained by employing a new culturing and grafting technique: styles with attached stigmas were excised from their ovaries and cultured in large Petri dishes. The styles were cut usually 15 mm. below the stigmas, then grafted to similarly cut styles from a plant of another incompatibility class and held together by means of a “splint” made of lactose-gelatin and a square of moistened lens paper. This “splint” was generally effective in keeping the cut ends in contact and in allowing pollen tubes to grow through the scion portion into the stock. In the 95 grafts involving a scion having no “S” alleles in common with those of the pollen and a stock containing the “S” allele of the pollen, tubes grew an average of 22.1 mm. into the stock during a 15-hr. period at 27°C., with some growing as far as 55 mm. under these conditions. These results suggest the possibility of 3 alternative mechanisms: (1) the incompatibility reaction is stronger in the stigma than in the style; (2) in the growth of pollen tubes through compatible tissue some stimulus is received or some substance is formed. which allows continued growth into the otherwise incompatible tissue of the stock; or (3) some of the stylar, inhibitory substance is lost by diffusion into the gelatin mixture at the graft juncture.     

Morphological change in the early stages of the mating process of Rhodosporidium toruloides.

The events which occur in the early stages of the mating process of the yeast Rhodosporidium toruloides between strains M-919 (mating type A) and M-1057 (mating type a) were investigated. In preliminary experiments we determined the frequency of mating by two newly designed methods: the liquid culture method and the membrane-filter microculture method. The mating frequencies of strains M-919 and M-1057 were 89% in the liquid culture method and 62% in the membrane-filter microculture method. The early stages in the mating process included the following events: (i) M-919 cells produce constitutively the extracellular inducing substance (A factor), (ii) M-1057 cells receive A factor, and in response to it they form mating tubes and secrete another inducing substance (a factor), (iii) M-919 cells receive a factor, and in response to it they form mating tubes, (iv) mating tubes elongate to the cells or the tubes of mating partner, (v) tips of the growing tubes recognize the opposite mating type cells or their tubes, followed by cell-to-cell fusion.     

The Nutritional Role of Pistil Exudate in Pollen Tube Wall Formation in Lilium longiflorum: I. Utilization of Injected Stigmatic Exudate

A quantity of labeled stigmatic exudate, collected from detached Lilium longiflorum (cv. Ace) pistils labeled with d-glucose-1-¹⁴C, was fractionated on Sephadex G-100 and the polysaccharide component, G-100-I, was injected into the hollow styles of unlabeled detached pistils (cv. Ace) which had been removed on the day after anthesis from the plant. Injected pistils were immediately cross-pollinated with L. longiflorum (cv. No. 44) pollen. Eighty-four hours later, pistils were dissected to recover the pollen tubes, expended exudate, and labeled tissues of the stigma and style. Distribution of label revealed that at least 25% of the carbohydrate substance in excised pollen tubes was derived from G-100-I. The composition of expended exudate adhering to pollen tubes, of pollen tube cytoplasm, and of pollen tube walls suggests that utilization of exudate by growing pollen tubes involves uptake and incorporation into pollen tube cytoplasm of exudate polysaccharide fragments followed by extensive metabolism of at least a portion of the incorporated carbohydrate prior to its utilization for pollen tube wall biosynthesis. Results suggest the presence of at least two polysaccharide components in G-100-I, one which resists major degradation following injection into the style and another which undergoes measurable degradation both before and after entry into the pollen tube.     

Chemically mediated mechanical expansion of the pollen tube cell wall

Morphogenesis of plant cells is tantamount to the shaping of the stiff cell wall that surrounds them. To this end, these cells integrate two concomitant processes: 1), deposition of new material into the existing wall, and 2), mechanical deformation of this material by the turgor pressure. However, due to uncertainty regarding the mechanisms that coordinate these processes, existing models typically adopt a limiting case in which either one or the other dictates morphogenesis. In this report, we formulate a simple mechanism in pollen tubes by which deposition causes turnover of cell wall cross-links, thereby facilitating mechanical deformation. Accordingly, deposition and mechanics are coupled and are both integral aspects of the morphogenetic process. Among the key experimental qualifications of this model are: its ability to precisely reproduce the morphologies of pollen tubes; its prediction of the growth oscillations exhibited by rapidly growing pollen tubes; and its prediction of the observed phase relationships between variables such as wall thickness, cell morphology, and growth rate within oscillatory cells. In short, the model captures the rich phenomenology of pollen tube morphogenesis and has implications for other plant cell types.     

More than a leak sealant. The mechanical properties of callose in pollen tubes

While callose is a well-known permeability barrier and leak sealant in plant cells, it is largely unknown whether this cell wall polymer can also serve as a load-bearing structure. Since callose occurs in exceptionally large amounts in pollen, we assessed its role for resisting tension and compression stress in this cell. The effect of callose digestion in Solanum chacoense and Lilium orientalis pollen grains demonstrated that, depending on the species, this cell wall polymer represents a major stress-bearing structure at the aperture area of germinating grains. In the pollen tube, it is involved in cell wall resistance to circumferential tension stress, and despite its absence at the growing apex, callose is indirectly involved in the establishment of tension stress resistance in this area. To investigate whether or not callose is able to provide mechanical resistance against compression stress, we subjected pollen tubes to local deformation by microindentation. The data revealed that lowering the amount of callose resulted in reduced cellular stiffness and increased viscoelasticity, thus indicating clearly that callose is able to resist compression stress. Whether this function is relevant for pollen tube mechanics, however, is unclear, as stiffened growth medium caused a decrease in callose deposition. Together, our data provide clear evidence for the capacity of cell wall callose to resist tension and compression stress, thus demonstrating that this amorphous cell wall substance can have a mechanical role in growing plant cells.     

A Preliminary Study on the Structure and Function of the Vascular Transfer Cells in Garlic Scape

The vascular transfer cells in garlic scape havebeen examined with electron microscope. Their structure, distributive feature and adenosine triphosphatase (ATPase) activity are studied. The mature vascular transfer cells exhibit the characteristic cell wall ingrowths. The cell contents include a large nucleus, dense cytoplasm and various normal organelles. It is notable that there are numerous mitochondria with well developed, cristae. Plasmodesmata are extensively present in the wall, and transfer cells are connected to adjacent cells by them. The senescing transfer cells become more vacuolated and have a large central vacuole and dense parietal cytoplasm. Their wall ingrowths seem to degenerate and finally disappear. The transfer cells show a particular pattern of distribution in the vascular bundle of the garlic scape. Some of them are present between the vessels of xylem and the sieve tubes of phloem. However, more abundant cell wall ingrowths occur on those walls which abut on, or are close to the vessel of xylem. The other transfer cells are located between the sieve tubes and parenehyma cells. The phloem transfer cell which is adjacent to sieve tube has developed from companion cell. All the transfer cells are mainly concerned with the loading and unloading of sieve tubes. And they may play an important role in facilitating intensive material transfer between two independent systems (i.e. the vessels and sieve tubes, the symplast and apoplast). The results of the cytochemical localization of ATPase using a lead precipitation technique exhibit strong enzyme activity on the plasmalemma of the transfer cells. It is suggested that the transfer cells are especially active in solute movement through them to which cellular energy metabolism coupled.     

2, 6-dichlorobenzonitrile Causes Multiple Effects on Pollen Tube Growth beyond Altering Cellulose Synthesis in Pinus bungeana Zucc

Cellulose is an important component of cell wall, yet its location and function in pollen tubes remain speculative. In this paper, we studied the role of cellulose synthesis in pollen tube elongation in Pinus bungeana Zucc. by using the specific inhibitor, 2, 6-dichlorobenzonitrile (DCB). In the presence of DCB, the growth rate and morphology of pollen tubes were distinctly changed. The organization of cytoskeleton and vesicle trafficking were also disturbed. Ultrastructure of pollen tubes treated with DCB was characterized by the loose tube wall and damaged organelles. DCB treatment induced distinct changes in tube wall components. Fluorescence labeling results showed that callose, and acidic pectin accumulated in the tip regions, whereas there was less cellulose when treated with DCB. These results were confirmed by FTIR microspectroscopic analysis. In summary, our findings showed that inhibition of cellulose synthesis by DCB affected the organization of cytoskeleton and vesicle trafficking in pollen tubes, and induced changes in the tube wall chemical composition in a dose-dependent manner. These results confirm that cellulose is involved in the establishment of growth direction of pollen tubes, and plays important role in the cell wall construction during pollen tube development despite its lower quantity.     

Cell surface arabinogalactan-proteins and their relation to cell proliferation and viability

Summary We have used high-pressure freezing followed by freeze substitution (HPF/FS) to preserve in vivo grown lily pollen tubes isolated from the style. The results indicated that HPF/FS (i) allows excellent preservation of the pollen tubes, (ii) maintains in situ the stylar matrix secreted by the transmitting tract cells, and (iii) preserves the interactions that exist between pollen tubes. Particular attention has been given to the structure of the pollen tube cell wall and the zone of adhesion. The cell wall is composed of an outer fibrillar layer and an inner layer of material similar in texture and nature to the stylar matrix and that is not callose. The stylar matrix labels strongly for arabinogalactan proteins (AGPs) recognized by monoclonal antibody JIM13. The zone of adhesion between pollen tubes contains distinct matrix components that are not recognized by JIM13, and apparent cross-links between the two cell walls. This study indicates that HPF/FS can be used successfully to preserve in vivo grown pollen tubes for ultrastructural investigations as well as characterization of the interactions between pollen tubes and the stylar matrix.Abbreviations AGPs arabinogalactan proteins - FS freeze substitution - HPF high-pressure freezing     

Microbial Oxidation of Cephalosporins to Cephalosporin Sulfoxides

The highest inhibition rate of conidial germination of Pyricularia oryzae was shown by extracts of rice plant leaves inoculated by a pathogen after treatment with probenazole, a rice blast controlling agent. Four anti-conidial germination substances were isolated from these extracts. Substances A, C and D inhibited the germination of the conidia at concentrations between 100 and 200 mcg/ml, and substance B caused morphological changes in the germination tubes of the conidia with a little inhibition of germination. These substances were differentiated from momilactone A, B and the degraded or metabolized products of probenazole. Besides anti-conidial germination activity, they showed antimicrobial activities against several kinds of phytopathogenic bacteria of fungi on agar plates by diffusion method.     

General tube law for collapsible thin and thick-wall tubes

Modeling the complex deformations of cylindrical tubes under external pressure is of interest in engineering and physiological applications. The highly non-linear post-buckling behavior of cross-section of the tube during collapse attracted researchers for years. Major efforts were concentrated on studying the behavior of thin-wall tubes. Unfortunately, the knowledge on post-buckling of thick-wall tubes is still incomplete, although many experimental and several theoretical studies have been performed. In this study we systematically studied the effect of the wall thickness on post-buckling behavior of the tube. For this purpose, we utilized a computational model for evaluation of the real geometry of the deformed cross-sectional area due to negative transmural (internal minus external) pressure. We also developed an experimental method to validate the computational results. Based on the computed cross-sections of tubes with different wall thicknesses, we developed a general tube law that accounts for thin or thick wall tubes and fits the numerical data of computed cross-sectional areas versus transmural pressures.     

Molecular modeling and simulation of Mycobacterium tuberculosis cell wall permeability

The low permeability of the mycobacterial cell wall is thought to contribute to the intrinsic drug resistance of mycobacteria. In this study, the permeability of the Mycobacterium tuberculosis cell wall is studied by computer simulation. Thirteen known drugs with diverse chemical structures were modeled as solutes undergoing transport across a model for the M. tuberculosis cell wall. The properties of the solute-membrane complexes were investigated by means of molecular dynamics simulation, especially the diffusion coefficients of the solute molecules inside the cell wall. The molecular shape of the solute was found to be an important factor for permeation through the M. tuberculosis cell wall. Predominant lateral diffusion within, as opposed to transverse diffusion across, the membrane/cell wall system was observed for some solutes. The extent of lateral diffusion relative to transverse diffusion of a solute within a biological cell membrane may be an important finding with respect to absorption distribution, metabolism, elimination, and toxicity properties of drug candidates. Molecular similarity measures among the solutes were computed, and the results suggest that compounds having high molecular similarity will display similar transport behavior in a common membrane/cell wall environment. In addition, the diffusion coefficients of the solute molecules across the M. tuberculosis cell wall model were compared to those across the monolayers of dipalmitoylphosphatidylethanolamine and dimyristoylphosphatidylcholine, are two common phospholipids in bacterial and animal membranes. The differences among these three groups of diffusion coefficients were observed and analyzed.     

Ultrastructure of Wall Swellings of Germinating Sporangia of Phytophthora palmivora (Butl.) Butl.

Germ tubes of directly germinating sporangia of P. palmivoraincubated in yeast extract solution at 30 ?C usually developedinto prominent swellings from which hyphae later emerged. Thegerm tubes arose as an extension of a new germination wall formedinternal to the sporangial wall prior to germination. The germtube swellings contained typical hyphal organelles. The germtube swelling possessed a thicker wall than both hyphae growingout of it and germ tubes that did not form swellings.     

Persistent Symmetry Frustration in Pollen Tubes

Pollen tubes are extremely rapidly growing plant cells whose morphogenesis is determined by spatial gradients in the biochemical composition of the cell wall. We investigate the hypothesis (MP) that the distribution of the local mechanical properties of the wall, corresponding to the change of the radial symmetry along the axial direction, may lead to growth oscillations in pollen tubes. We claim that the experimentally observed oscillations originate from the symmetry change at the transition zone, where both intervening symmetries (cylindrical and spherical) meet. The characteristic oscillations between resonating symmetries at a given (constant) turgor pressure and a gradient of wall material constants may be identified with the observed growth-cycles in pollen tubes.     

Pollen tubes of flavonol-deficient Petunia show striking alterations in wall structure leading to tube disruption

Despite the vital role that flavonols play in fertilization and pollen tube growth of a number of species such as petunia and maize, their function is still unclear. Pollen tubes of the flavonol-deficient transformant T17.02 of Petunia hybrida L. are able to germinate and start growing in vitro, but eventually disrupt at the tip approximately 2 h after germination. In order to establish the possible role of flavonols in this process, wild-type and flavonol-deficient pollen tubes were subjected to cytological and ultrastructural analyses and screened for differences. The results showed that before disruption of the flavonol-deficient pollen tubes, the structure of the primary wall at the tip dramatically changed from layered to granular. Secretory vesicles at the tip still fused with the wall but lost their capacity to melt into the wall and to form layers. Instead they remained as dark, electron-dense granular structures surrounded by an electron-translucent matrix. Apparently the matrix is not able to sustain the wall's coherence and as a consequence the tube disrupts. No other remarkable cytological or ultrastructural differences between the transformant and the wild-type pollen tubes could be found before tip disruption. Even a morphometric analysis of abundance and distribution of endoplasmic reticulum, dictyosomes and mitochondria did not reveal any significant difference. However, for the first time, obvious morphological differences were observed in the wall of the flavonol-deficient pollen tubes. We conclude that flavonols act on precursors of the pollen tube wall of petunia and interfere with a cross-linking system in the wall, possibly via extensins. Received: 23 February 1998 / Accepted: 13 August 1998     

Ultrastructure of the stylar canal cells ofCitrus limon (Rutaceae)

The ultrastructure of the canal cells and the canal filling substance ofCitrus limon have been studied. At maturity the canal cells are very rich in cytoplasm. Their inner tangential walls lining the canal are much thickened and formed by two layers: the outer corresponds to the original wall, the inner is formed by subsequent deposition of abundant materials of different origin. This thickening occurs at the same time as the filling of the stylar canal. Both events are paralleled by considerable dictyosomic activity, the formation of a large amount of rough endoplasmic reticulum, and the incorporation of small cytoplasmic masses into the cell wall, due to plasmalemma evaginations. — The material in the stylar canal has a heterogeneous ultrastructure aspect and consists of polysaccharides, proteins and lipids; it presumably provides nutrients for the growing pollen tubes.Research performed under CNR program Biology of Reproduction.     

Ultrastructure of feeding tubes formed in giant-cells induced in plants by the root-knot nematodeMeloidogyne incognita

Summary The plant pathogenic nematodeMeloidogyne incognita forms conspicuous tubular structures referred to as feeding tubes in special food cells, called giant-cells, induced and maintained in susceptible host roots by feeding nematodes. Feeding tubes are formed by nematode secretions injected into giant-cells via a stylet and apparently function to facilitate withdrawal of soluble assimilates by the parasite. In giant-cells in roots of the four host species examined in this study, feeding tube morphology was identical. Tubes were straight to slightly curved structures just less than 1 μm wide and up to slightly more than 70 μm long. At the ultrastructural level, each tube consisted of a 190–290 nm thick, electron-dense, crystalline wall surrounding an electron-transparent lumen with a diameter of 340–510 nm. The distal end of the tube was sealed with wall material. Older tubes were found free in the host cytoplasm while the proximal ends of young tubes were attached to the host cell wall via short wall ingrowths through which the nematode's stylet was inserted. An elaborate membrane system was associated with the feeding tubes and was most extensive around newly formed tubes. Contiguous to the feeding tube wall, this membrane system consisted of strands of smooth endoplasmic reticulum while rough endoplasmic reticulum predominated toward the outer margin of the membrane system. Vacuoles and mitochondria were excluded from a zone of cytoplasm surrounding feeding tubes. This zone of exclusion, as well as the membrane system noted above, tended to be less pronounced or absent around older tubes no longer being used by the nematode.