scholarly journals Microtubules play a role in trafficking prevacuolar compartments to vacuoles in tobacco pollen tubes

Open Biology ◽  
2018 ◽  
Vol 8 (10) ◽  
pp. 180078 ◽  
Author(s):  
Elisabetta Onelli ◽  
Monica Scali ◽  
Marco Caccianiga ◽  
Nadia Stroppa ◽  
Piero Morandini ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Pollen Tube ◽  
Biochemical Analysis ◽  
Pollen Tubes ◽  
Degradation Pathways ◽  
Transient Transformation ◽  
Secretory Pathways ◽  
Tobacco Pollen ◽  
Fine Regulation ◽  
Pollen Tube Elongation

Fine regulation of exocytosis and endocytosis plays a basic role in pollen tube growth. Excess plasma membrane secreted during pollen tube elongation is known to be retrieved by endocytosis and partially reused in secretory pathways through the Golgi apparatus. Dissection of endocytosis has enabled distinct degradation pathways to be identified in tobacco pollen tubes and has shown that microtubules influence the transport of plasma membrane internalized in the tip region to vacuoles. Here, we used different drugs affecting the polymerization state of microtubules together with SYP21, a marker of prevacuolar compartments, to characterize trafficking of prevacuolar compartments in Nicotiana tabacum pollen tubes. Ultrastructural and biochemical analysis showed that microtubules bind SYP21-positive microsomes. Transient transformation of pollen tubes with LAT52-YFP-SYP21 revealed that microtubules play a key role in the delivery of prevacuolar compartments to tubular vacuoles.

scholarly journals Coordinated Localization and Antagonistic Function of NtPLC3 and PI4P 5-Kinases in the Subapical Plasma Membrane of Tobacco Pollen Tubes

Plants ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 452 ◽  
Author(s):  
Irene Stenzel ◽  
Till Ischebeck ◽  
Linh Hai Vu-Becker ◽  
Mara Riechmann ◽  
Praveen Krishnamoorthy ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Pollen Tube ◽  
Tip Growth ◽  
Pollen Tubes ◽  
Dominant Negative ◽  
Membrane Domains ◽  
Tobacco Pollen ◽  
Polar Tip Growth ◽  
Spinning Disc

Polar tip growth of pollen tubes is regulated by the membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2), which localizes in a well-defined region of the subapical plasma membrane. How the PtdIns(4,5)P2 region is maintained is currently unclear. In principle, the formation of PtdIns(4,5)P2 by PI4P 5-kinases can be counteracted by phospholipase C (PLC), which hydrolyzes PtdIns(4,5)P2. Here, we show that fluorescence-tagged tobacco NtPLC3 displays a subapical plasma membrane distribution which frames that of fluorescence-tagged PI4P 5-kinases, suggesting that NtPLC3 may modulate PtdIns(4,5)P2-mediated processes in pollen tubes. The expression of a dominant negative NtPLC3 variant resulted in pollen tube tip swelling, consistent with a delimiting effect on PtdIns(4,5)P2 production. When pollen tube morphologies were assessed as a quantitative read-out for PtdIns(4,5)P2 function, NtPLC3 reverted the effects of a coexpressed PI4P 5-kinase, demonstrating that NtPLC3-mediated breakdown of PtdIns(4,5)P2 antagonizes the effects of PtdIns(4,5)P2 overproduction in vivo. When analyzed by spinning disc microscopy, fluorescence-tagged NtPLC3 displayed discontinuous membrane distribution omitting punctate areas of the membrane, suggesting that NtPLC3 is involved in the spatial restriction of plasma membrane domains also at the nanodomain scale. Together, the data indicate that NtPLC3 may contribute to the spatial restriction of PtdIns(4,5)P2 in the subapical plasma membrane of pollen tubes.

Dynamics of the apical vesicle accumulation and the rate of growth are related in individual pollen tubes

2001 ◽  
Vol 114 (14) ◽  
pp. 2685-2695 ◽  
Author(s):  
R. M. Parton ◽  
S. Fischer-Parton ◽  
M. K. Watahiki ◽  
A. J. Trewavas
Keyword(s):  
Growth Rate ◽  
Plasma Membrane ◽  
Pollen Tube ◽  
Staining Pattern ◽  
Periodic Oscillation ◽  
Quantitative Relationship ◽  
Pollen Tubes ◽  
Apical Region ◽  
Secretory Vesicles ◽  
Animal Cells

Regulated secretory vesicle delivery, vesicle fusion and rapid membrane recycling are all contentious issues with respect to tip growth in plant, fungal and animal cells. To examine the organisation and dynamics of membrane movements at the growing pollen tube apex and address the question of their relationship to growth, we have used the membrane stain FM4-64 both as a structural marker and as a quantitative assay. Labelling of living Lilium Longiflorum pollen tubes by FM4-64 resulted in a distinct staining pattern in the tube apex, which corresponds spatially to the previously identified cone-shaped `apical clear zone' containing secretory vesicles. Dye uptake could be inhibited by sodium azide and followed a strict temporal sequence from the plasma membrane to a population of small (1-2 μm diameter) discrete internal structures, with subsequent appearance of dye in the apical region and ultimately in vacuolar membranes. Washout of the dye rapidly removed the plasma membrane staining, which was followed by a gradual decline in the apical fluorescence over more than an hour. Injected aqueous FM4-64 solution showed a relatively even distribution within the pollen tube. Association of FM4-64 with apical secretory vesicles was supported by the effects of the inhibitors Brefeldin-A and Cytochalasin-D, which are known to affect the localisation and number of such vesicles, on the FM4-64 staining pattern. Examination of the dynamics of FM4-64 labelling in the pollen tube tip by time-lapse observation, supported by fluorescence-recovery-after-photobleaching (FRAP) analysis, suggested the possibility of distinct pathways of bulk membrane movement both towards and, significantly, away from the apex. Quantitative analysis of FM4-64 distribution in the apex revealed that fluctuations in fluorescence 5 to 10 μm subapically, and to a lesser extent the apical 3 μm, could be related to the periodic oscillation in pollen tube growth rate. This data reveals a quantitative relationship between FM4-64 staining and growth rate within an individual tube.

Confocal image analysis of spatial variations in immunocytochemically identified calmodulin during pollen hydration, germination and pollen tube tip growth in Nicotiana tabacum L.

Zygote ◽  
1994 ◽  
Vol 2 (1) ◽  
pp. 63-68 ◽  
Author(s):  
Uday K. Tirlapur ◽  
Monica Scali ◽  
Alessandra Moscatelli ◽  
Cecilia Del Casino ◽  
Gianpiero Cai ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Nicotiana Tabacum ◽  
Pollen Tube ◽  
Pollen Tubes ◽  
Spatial Variations ◽  
Tube Growth ◽  
Confocal Scanning Laser Microscopy ◽  
Pollen Hydration ◽  
Confocal Scanning Laser ◽  
High Level

SummaryUsing monoclonal anti-calmodulin antibodies in conjunction with confocal scanning laser microscopy we have analysed the spatial variations in the distribution pattern of calmodulin (CaM) during the sequential events of pollen hydration, germination and tube growth in Nicotiana tabacum. These immunocytochemical observations have been complemented by immunochemical studies wherein the anti-calmodulin antibody raised against pea CaM recognises a polypeptide of c. 18 kDa in the pollen extracts. Digitisation of confocally acquired optical sections of immunofluorescence images reveals that in hydrated pollen a high level of CaM is consistently present in the region of the germinal apertures. Subsequently, with the onset of germination a high CaM concentration was found associated with the plasma membrane of the germination bubble and in the cytoplasm in its vicinity, while in the vegetative cytoplasm a weak diffuse and intense punctate signal was registered. CaM immunostain was also detected in association with the plasma membrane of the tube tips in both short and long pollen tubes. Furthermore, the cytosol of the tubes invariably manifested an apically focused CaM gradient. We were, however, unable to detect any vacuolar association of CaM in the older regions of the pollen tubes. Although punctate immunostain was obvious across the pollen tube numerous punctate structures were invariably present in the extreme tip. The possible implications of these findings in development of cell polarity, polarised growth, maintenance of calcium homeostasis and CaM interactions with other mechanochemical motor proteins in effecting propulsion of organelles during pollen hydration, germination and pollen tube growth are discussed.

The influence of the detergent Triton X-100 on the growth and ultrastructure of tobacco pollen tubes

1990 ◽  
Vol 68 (5) ◽  
pp. 1131-1138 ◽  
Author(s):  
K. S. Rao ◽  
U. Kristen
Keyword(s):  
Pollen Tube ◽  
Growth Inhibition ◽  
Effective Dose ◽  
Toxic Substance ◽  
Pollen Tubes ◽  
Culture Technique ◽  
Tube Growth ◽  
Wall Thickening ◽  
Tobacco Pollen ◽  
Triton X

The influence of the detergent Triton X-100 on the growth and ultrastructure of tobacco pollen tubes was studied using a culture technique. Tube growth inhibition increased with increasing concentration of the detergent in the culture medium. At a concentration of 7.5 mg∙l−1 (effective dose) the detergent induced a 50% reduction of pollen tube growth. The effective dose is defined as the concentration of a toxic substance that causes a decrease of a metabolic process to 50% of the control. Growth inhibition was reflected by changes of tube ultrastructure and shape at the tip region. The ultrastructural alterations in the pollen tube following the cytotoxic effect of the detergent included (i) disturbance of the cytoplasmic zonation in the tip region of the tube, (ii) numerical decrease of stacked dictyosomes (reduction of Golgi activity), (iii) damage of the plasma membrane and deposition of vesicular structures in the periplasmic space, (iv) formation of concentric arrangements of rough ER, (v) damage of the mitochondrial matrix and cristae and (vi) tube wall thickening-and loosening. The Thiéry reaction revealed the polysaccharide nature of the vesicle contents and the glycolipid nature of the lipid bodies. Scanning electron microscopy demonstrated curvature and tip swelling of the detergent-treated pollen tubes. Most of these changes were likely due to the action of the detergent on membranes. Key words: Nicotiana sylvestris, pollen tubes, detergent, growth inhibition, ultrastructure, biomembranes.

Late-acting self-incompatibility in tea plant (Camellia sinensis)

Biologia ◽  
2012 ◽  
Vol 67 (2) ◽  
Author(s):  
Xuan Chen ◽  
Shan Hao ◽  
Li Wang ◽  
Wanping Fang ◽  
Yuhua Wang ◽  
...  
Keyword(s):  
Pollen Tube ◽  
Camellia Sinensis ◽  
Pollen Tubes ◽  
Tea Plant ◽  
The Self ◽  
Self Incompatibility ◽  
Cross Pollination ◽  
Self Pollination ◽  
Hand Pollination ◽  
Pollen Tube Elongation

AbstractThe self-incompatibility of tea plant (Camellia sinensis (L.) O. Kuntze) was studied with the methods of aniline blue fluorescence assay and paraffin sections. The characteristics of pollen tube elongation after hand pollination was analyzed in 4 tea cultivars, including ‘Keemenzhong’, ‘Longjing-changye’, ‘Fuding-dabaicha’ and ‘Yabukita’, under self-pollination and cross-pollination, respectively. Although there were some difference among cultivars, pollen tubes elongated through the style and reach the ovary successfully at 48 h after pollination for both cross- and self-pollen tubes in all the four cultivars of tea. Pollen tubes entered into the ovule micropyles, however, only for cross-pollination, but not for self-pollination. Pollen tubes of selfing plants, failed in fertilizing, seemed have some difficulties to enter the ovule. All of which indicated that the self-incompatibility of tea plant is a late-acting self-incompatibility system (LSI) or an ovarian sterility (OS), in which the self incompatibility was due to none self pollen tube penetrating into the ovule and no fertilization.

scholarly journals Plasma membrane nano-organization specifies phosphoinositide effects on Rho-GTPases and actin dynamics in tobacco pollen tubes

2020 ◽  
Author(s):  
Marta Fratini ◽  
Praveen Krishnamoorthy ◽  
Irene Stenzel ◽  
Mara Riechmann ◽  
Kirsten Bacia ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Trafficking ◽  
Rho Gtpases ◽  
Rho Gtpase ◽  
Pollen Tubes ◽  
Actin Dynamics ◽  
Tobacco Pollen ◽  
Dual Distribution ◽  
Cytoskeletal Dynamics

AbstractPollen tube growth requires coordination of cytoskeletal dynamics and apical secretion. The regulatory phospholipid, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) is enriched in the subapical plasma membrane of pollen tubes and can influence both actin dynamics and secretion. How alternative PtdIns(4,5)P2-effects are specified is unclear. Spinning disc microscopy (SD) reveals dual distribution of a fluorescent PtdIns(4,5)P2-reporter in dynamic plasma membrane nanodomains vs. apparent diffuse membrane labelling, consistent with spatially distinct coexisting pools of PtdIns(4,5)P2. Several PI4P 5-kinases (PIP5Ks) can generate PtdIns(4,5)P2 in pollen tubes. Despite localizing to one membrane region, AtPIP5K2 and NtPIP5K6 display distinctive overexpression effects on cell morphologies, respectively related to altered actin dynamics or membrane trafficking. When analyzed by SD, AtPIP5K2-EYFP associated with nanodomains, whereas NtPIP5K6-EYFP localized diffusely. Chimeric AtPIP5K2 and NtPIP5K6 variants with reciprocally swapped membrane-associating domains evoked reciprocally shifted effects on cell morphology upon overexpression. Overall, PI4P 5-kinase variants targeted to nanodomains stabilized actin, suggesting a specific function of PtdIns(4,5)P2-nanodomains. A distinct role of nanodomain-associated AtPIP5K2 in actin regulation is further supported by proximity to and interaction with the Rho-GTPase NtRac5, and by functional interplay with elements of ROP-signalling. Plasma membrane nano-organization may thus aid the specification of PtdIns(4,5)P2-functions to coordinate cytoskeletal dynamics and secretion in pollen tubes.

scholarly journals The Arabidopsis thaliana Class II Formin FH13 Modulates Pollen Tube Growth

2021 ◽  
Vol 12 ◽  
Author(s):  
Eva Kollárová ◽  
Anežka Baquero Forero ◽  
Fatima Cvrčková
Keyword(s):  
Arabidopsis Thaliana ◽  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Tip Growth ◽  
Pollen Tubes ◽  
Class Ii ◽  
Membrane Dynamics ◽  
Tube Growth ◽  
Class I ◽  
Pollen Tube Elongation

Formins are a large, evolutionarily conserved family of actin-nucleating proteins with additional roles in regulating microfilament, microtubule, and membrane dynamics. Angiosperm formins, expressed in both sporophytic and gametophytic tissues, can be divided into two subfamilies, Class I and Class II, each often exhibiting characteristic domain organization. Gametophytically expressed Class I formins have been documented to mediate plasma membrane-based actin assembly in pollen grains and pollen tubes, contributing to proper pollen germination and pollen tube tip growth, and a rice Class II formin, FH5/RMD, has been proposed to act as a positive regulator of pollen tube growth based on mutant phenotype and overexpression data. Here we report functional characterization of the Arabidopsis thaliana pollen-expressed typical Class II formin FH13 (At5g58160). Consistent with published transcriptome data, live-cell imaging in transgenic plants expressing fluorescent protein-tagged FH13 under the control of the FH13 promoter revealed expression in pollen and pollen tubes with non-homogeneous signal distribution in pollen tube cytoplasm, suggesting that this formin functions in the male gametophyte. Surprisingly, fh13 loss of function mutations do not affect plant fertility but result in stimulation of in vitro pollen tube growth, while tagged FH13 overexpression inhibits pollen tube elongation. Pollen tubes of mutants expressing a fluorescent actin marker exhibited possible minor alterations of actin organization. Our results thus indicate that FH13 controls or limits pollen tube growth, or, more generally, that typical Class II formins should be understood as modulators of pollen tube elongation rather than merely components of the molecular apparatus executing tip growth.

Molecular control of the glucan synthase-like protein NaGSL1 and callose synthesis during growth of Nicotiana alata pollen tubes

2008 ◽  
Vol 414 (1) ◽  
pp. 43-52 ◽  
Author(s):  
Lynette Brownfield ◽  
Sarah Wilson ◽  
Ed Newbigin ◽  
Antony Bacic ◽  
Steve Read
Keyword(s):  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Nicotiana Alata ◽  
Molecular Control ◽  
Intracellular Location ◽  
Glucan Synthase ◽  
Tobacco Pollen

The protein NaGSL1 (Nicotiana alata glucan synthase-like 1) is implicated in the synthesis of callose, the 1,3-β-glucan that is the major polysaccharide in the walls of N. alata (flowering tobacco) pollen tubes. Here we examine the production, intracellular location and post-translational processing of NaGSL1, and relate each of these to the control of pollen-tube callose synthase (CalS). The 220 kDa NaGSL1 polypeptide is produced after pollen-tube germination and accumulates during pollen-tube growth, as does CalS. A combination of membrane fractionation and immunoelectron microscopy revealed that NaGSL1 was present predominantly in the endoplasmic reticulum and Golgi membranes in younger pollen tubes when CalS was mostly in an inactive (latent) form. In later stages of pollen-tube growth, when CalS was present in both latent and active forms, a greater proportion of NaGSL1 was in intracellular vesicles and the plasma membrane, the latter location being consistent with direct deposition of callose into the wall. N. alata CalS is activated in vitro by the proteolytic enzyme trypsin and the detergent CHAPS, but in neither case was activation associated with a detectable change in the molecular mass of the NaGSL1 polypeptide. NaGSL1 may thus either be activated by the removal of a few amino acids or by the removal of another protein that inhibits NaGSL1. These findings are discussed in relation to the control of callose biosynthesis during pollen germination and pollen-tube growth.

scholarly journals Ovary Signals for Pollen Tube Guidance in Chalazogamous Mangifera indica L.

2021 ◽  
Vol 11 ◽  
Author(s):  
Jorge Lora ◽  
Veronica Perez ◽  
Maria Herrero ◽  
Jose I. Hormaza
Keyword(s):  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Mangifera Indica ◽  
Embryo Sac ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Transmitting Tissue ◽  
Egg Apparatus ◽  
Pollen Tube Elongation

Most flowering plants show porogamy in which the pollen tubes reach the egg apparatus through the micropyle. However, several species show chalazogamy, an unusual pollen tube growth, in which the pollen tubes reach the embryo sac through the chalaza. While ovary signals for pollen tube growth and guidance have been extensively studied in porogamous species, few studies have addressed the process in chalazogamous species such as mango (Mangifera indica L.), one of the five most important fruit crops worldwide in terms of production. In this study, we characterize pollen–pistil interaction in mango, paying special attention to three key players known to be involved in the directional pollen tube growth of porogamous species such as starch, arabinogalactan proteins (AGPs), and γ-aminobutyric acid (GABA). Starch grains were observed in the style and in the ponticulus at anthesis, but their number decreased 1 day after anthesis. AGPs, revealed by JIM8 and JIM13 antibodies, were homogenously observed in the style and ovary, but were more conspicuous in the nucellus around the egg apparatus. GABA, revealed by anti-GABA antibodies, was specifically observed in the transmitting tissue, including the ponticulus. Moreover, GABA was shown to stimulate in vitro mango pollen tube elongation. The results support the heterotrophic growth of mango pollen tubes in the style at the expense of starch, similarly to the observations in porogamous species. However, unlike porogamous species, the micropyle of mango does not show high levels of GABA and starch, although they were observed in the ponticulus and could play a role in supporting the unusual pollen tube growth in chalazogamous species.

scholarly journals Effects of Polyamines on Self-incompatibility-like Responses in Pollen Tubes of Citrus Cultivars, Banpeiyu and Hyuganatsu

2015 ◽  
Vol 140 (2) ◽  
pp. 183-190 ◽  
Author(s):  
Yiran Li ◽  
Asuka Uchida ◽  
Akiha Abe ◽  
Akihiro Yamamoto ◽  
Tomonari Hirano ◽  
...  
Keyword(s):  
Pollen Tube ◽  
High Performance ◽  
Crude Protein ◽  
Pollen Tubes ◽  
Self Incompatibility ◽  
Morphological Alterations ◽  
Tube Content ◽  
Citrus Maxima ◽  
Pollen Tube Elongation ◽  
Inflation Tube

Citrus is a genus with gametophytic self-incompatibility, which prevents fertilization by self-pollination. Polyamines (PAs), as a class of active small molecules, widely participate in various cellular activities. To investigate the effects of the changes in PA contents [putrescine, spermidine (SPD), and spermine] on pollen tube growth in self-incompatible (SI) plants, we cultured pollen of Citrus maxima ‘Banpeiyu’ and C. tamurana ‘Hyuganatsu’ in a Citrus mature pollen culture system and detected the PA content changes in pollen by high-performance liquid chromatography (HPLC) analysis during germination and after treatment with crude protein extracts of compatible or incompatible styles. We profiled the PA content changes during the germination course in pollen of ‘Banpeiyu’ and ‘Hyuganatsu’ by HPLC, and identified increases of free and perchloric-acid-insoluble SPD contents in pollen tubes in the incompatible treatment. Exogenous SPD treatment obviously increased the SPD contents in the pollen tube, inhibited pollen tube elongation, and resulted in some abnormal morphological alterations, such as tip inflation, tube content leakage and tip twist, which were observed in both ‘Banpeiyu’ and ‘Hyuganatsu’ pollen tubes exposed to incompatible treatment. This finding implied that SPD might participate in the SI response in Citrus.

Sign in / Sign up

Export Citation Format

Share Document