Cytoskeletal Elements, Cell Shaping and Movement in the Angiosperm Pollen Tube

1988 ◽  
Vol 91 (1) ◽  
pp. 49-60 ◽  
Author(s):  
J. HESLOP-HARRISON ◽  
Y. HESLOP-HARRISON ◽  
M. CRESTI ◽  
A. TIEZZI ◽  
A. MOSCATELLI

The ellipsoidal generative cell of the pollen grain of Endymion nonscriptus usually elongates further following germination and entry into the tube, producing attenuated extensions the forward one of which may reach into the vicinity of the vegetative nucleus. This shape change is accompanied by the stretching of the microtubule cytoskeleton of the cell, identified in the present work by immunofluorescence using monoclonal antibodies to tubulin. Complementary observations of living generative cells of Iris pseudacorus showed that they undergo slow undulatory movements accompanied by variation in shape and length during passage through the tube. Such changes must presumably be accompanied by modifications of the microtubule cytoskeleton. Colchicine at 1 mM eliminated microtubules from tubes and most generative cells of E. nonscriptus, but did not radically affect pollen-tube shape or extension growth, nor arrest the movements of the vegetative nucleus and generative cell into and through the tube. Generative cells in colchicinetreated pollen of Galanthus nivalis rounded up and failed to undergo the usual changes in shape during passage through the tube. Secondary consequences were changes in precedence in movement through the tube, and a greater dispersal along its length. On the assumption that no other cytoskeletal elements remain to be discovered, it seems likely that microfilaments rather than microtubules provide the motive force for movement in the tube, although the latter are involved in shaping the generative cell and adapting it to its passage.

1989 ◽  
Vol 94 (2) ◽  
pp. 319-325
Author(s):  
J. HESLOP-HARRISON ◽  
Y. HESLOP-HARRISON

Myosin, detected by immunofluorescence using an antibody to bovine skeletal and smooth muscle myosin, has been localised on individual identifiable organelles from the grasses Alopecurus pratensis and Secale cereale, and on the surfaces of vegetative nuclei and generative cells from pollen and pollen tubes of Hyacinthus orientalis and Helleborus foetidus. Taken in conjunction with recent evidence showing that the growing pollen tube contains an actin cytoskeleton consisting of numerous mainly longitudinally oriented microfilament bundles, and that isolated pollen-tube organelles show ATP-dependent movement along the actin bundles of the giant cells of the characeous algae, this finding suggests that an actomyosin motility system is present in pollen tubes, and indicates that the movements of the different classes of inclusions are driven by interaction of the surface myosin with the actin fibrils at the zones of contact.


1995 ◽  
Vol 108 (7) ◽  
pp. 2549-2563
Author(s):  
D.D. Miller ◽  
S.P. Scordilis ◽  
P.K. Hepler

The presence and localization of actin and myosin have been examined in pollen tubes of Lilium longiflorum and Nicotiana alata. Immunoblot analysis of pollen tube extracts with antibodies to actin, myosins IA and IB, myosin II, and myosin V reveals the presence of these contractile proteins. Immunofluorescence microscopy using various methods to preserve the pollen tubes; chemical fixation, rapid freeze fixation and freeze substitution (RF-FS) followed by rehydration or by embeddment in a methacrylate mixture, was performed to optimize preservation. Immunocytochemistry reaffirmed that actin is localized longitudinally in the active streaming lanes and near the cortical surface of the pollen tube. Myosin I was localized to the plasma membrane, larger organelles, the surface of the generative cell and the vegetative nucleus, whereas, myosin V was found in the vegetative cytoplasm in a punctate fashion representing smaller organelles. Myosin II subfragment 1 and light meromyosin were localized in a punctate fashion on the larger organelles throughout the vegetative cytoplasm. In addition, isolated generative cells and vegetative nuclei labeled only with the myosin I antibody. Competition studies indicated the specificity of the heterologous antibodies utilized in this study suggesting the presence of three classes of myosins in pollen. These results lead to the following hypothesis: Myosin I may move the generative cell and vegetative nucleus unidirectionally through the pollen tube to the tip, while myosin V moves the smaller organelles and myosins I and II move the larger organelles (bidirectionally) that are involved in growth.


1989 ◽  
Vol 93 (2) ◽  
pp. 299-308
Author(s):  
J. HESLOP-HARRISON ◽  
Y. HESLOP-HARRISON

Actin is present in the cytoplasm of the vegetative cell of angiosperm pollens in numerous fusiform, spiculate or toroidal bodies, and also as a sheath enveloping the vegetative nucleus. During activation following hydration, the compact cytoplasmic bodies are translated into skeins of extended fibrils, and circulatory movements begin in the cytoplasm. Throughout this period the vegetative nucleus, with fibrillar actin now associated with the surface, undergoes a continuous change of shape. In the extending tube following germination the actin cytoskeleton consists of numerous mainly longitudinally oriented fibrils. After entry into the tube the vegetative nucleus remains associated with the fibrils, usually extending greatly in length and developing attenuated, often pointed extensions. The observed conformations, which change continuously, suggest that varying local tensions are applied to the vegetative nucleus during passage through the tube. Cytochalasin D breaks up the actin fibril system and brings about a rapid contraction of the nucleus, at the same time eliminating the elastic extensions of the nuclear envelope. Nuclei isolated physically from unfixed tubes also contract in length as the fibrillar components of the cytoskeleton are detached. These findings indicate that the movement of the vegetative nucleus depends on local associations of the nuclear envelope with the actin cytoskeleton of the vegetative cell.


2014 ◽  
Vol 68 (2) ◽  
pp. 97-102
Author(s):  
Małgorzata Stępka ◽  
Fabricio Ciampolini ◽  
Mauro Cresti ◽  
Maria Charzyńska

The germinating pollen grain (in vivo on the stigma or in vitro in germination medium) forms a pollen tube which transports the vegetative nucleus and generative cell/two sperm cells participating in the process of double fertilization. The growth of the tube and the transport of organelles and the cells occur due to two major motor systems existing in the pollen tubes of higher plants: the tubuline-dynein/kinesin and the actin-myosin system. In pollen tubes of <em>Ornithogalum virens</em> the actin filaments were labelled with TRITC-phalloidin (2 µg/ml) in the PIPES buffer and the 10% sucrose, without the fixative and DMSO. Omission of the fixative and permeabilizing agent (DMSO) allowed better preservation of the structure, and the "fluorescence" of actin was observed in living pollen tubes. Observations in CLSM (confocal laser scanning microscope) showed that actin is distributed in the vicinity of the cell membrane. This could support the view that actin filaments and the plasmalemma form the pollen tube cortex along which the cytoplasmic movement of organelles, and cell transport occurs.


2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Kazuki Motomura ◽  
Hidenori Takeuchi ◽  
Michitaka Notaguchi ◽  
Haruna Tsuchi ◽  
Atsushi Takeda ◽  
...  

AbstractDuring the double fertilization process, pollen tubes deliver two sperm cells to an ovule containing the female gametes. In the pollen tube, the vegetative nucleus and sperm cells move together to the apical region where the vegetative nucleus is thought to play a crucial role in controlling the direction and growth of the pollen tube. Here, we report the generation of pollen tubes in Arabidopsis thaliana whose vegetative nucleus and sperm cells are isolated and sealed by callose plugs in the basal region due to apical transport defects induced by mutations in the WPP domain-interacting tail-anchored proteins (WITs) and sperm cell-specific expression of a dominant mutant of the CALLOSE SYNTHASE 3 protein. Through pollen-tube guidance assays, we show that the physiologically anuclear mutant pollen tubes maintain the ability to grow and enter ovules. Our findings provide insight into the sperm cell delivery mechanism and illustrate the independence of the tip-localized vegetative nucleus from directional growth control of the pollen tube.


The development of the male gametophyte of Taxus baccata has been studied over a period of 20 weeks, from germination of the microspore in February to spermatogenesis in July. A few days after germination the microspore nucleus divides and a transverse wall forms at the equator cutting off the small generative cell and a large tube cell. The latter immediately begins to expand to form the pollen tube. The first division thus establishes the polarity of the gametophyte and the generative cell is regarded as proximal. The transverse wall is ephemeral, and within six weeks it has disappeared. The nucleus of the generative cell divides while still at the proximal pole. The two daughter nuclei are unequal in size, but they remain associated and together move distally. The larger nucleus eventually becomes the nucleus of the spermatogenous cell, and the smaller the sterile nucleus. The spermatogenous cell acquires a distinctive cytoplasm and becomes surrounded by a wall which arises de novo . The nucleus of the spermatogenous cell enlarges, but always remains towards one side of the cell so that at mitosis the spindle is contained within one hemisphere. After division the wall of the spermatogenous cell is ruptured and the two sperms are released as naked nuclei of equal size. The cytoplasm of the spermatogenous cell degenerates as it enters the tube, but remains recognizable until fertilization.


1989 ◽  
Vol 92 (4) ◽  
pp. 569-574
Author(s):  
X.J. Tang ◽  
P.K. Hepler ◽  
S.P. Scordilis

A myosin heavy chain polypeptide has been identified and localized in Nicotiana pollen tubes using monoclonal anti-myosin antibodies. The epitopes of these antibodies were found to reside on the myosin heavy chain head and rod portion and were, therefore, designated anti-S-1 (myosin S-1) and anti-LMM (light meromyosin). On Western blots of the total soluble pollen tube proteins, both anti-S-1 and anti-LMM label a polypeptide of approximately 175,000 Mr. Immunofluorescence microscopy shows that both antibodies yield numerous fluorescent spots throughout the whole length of the tube, often with an enrichment in the tube tip. These fluorescent spots are thought to represent vesicles and/or organelles in the pollen tubes. In addition to this common pattern, anti-S-1 stains both the generative cell and the vegetative nuclear envelope. The different staining patterns of the nucleus between anti-S-1 and anti-LMM may be caused by some organization and/or anchorage state of the myosin molecules on the nuclear surface that differs from those on the vesicles and/or organelles.


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