scholarly journals pH gradients are not associated with tip growth in pollen tubes of Lilium longiflorum

1997 ◽  
Vol 110 (15) ◽  
pp. 1729-1740 ◽  
Author(s):  
M.D. Fricker ◽  
N.S. White ◽  
G. Obermeyer
Keyword(s):  
Pollen Tube ◽  
Tip Growth ◽  
Lilium Longiflorum ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Cytoplasmic Ph ◽  
Clear Zone ◽  
External Application ◽  
Ph Gradients ◽  
External Ph

The cytoplasmic pH of growing pollen tubes of Lilium longiflorum Thunb. was measured using the pH-sensitive fluorescent dye 2′,7′-bis-(carboxyethyl)-5(6′)-carboxyfl uorescein and confocal fluorescence ratio imaging. The average cytoplasmic pH in the clear zone of the pollen tube tip was pH 7.11, and no consistent pH gradients were detected in the clear zone, averaging around -1.00 milli pH unit microm(−1), or along the first 50 microm of the tube (3.62 milli pH units microm[-1]). In addition, no correlation was observed between the absolute tip cytoplasmic pH or the pH gradient and the pollen tube growth rates. Shifts of external pH to more acidic pH values (pH 4.5) caused a relatively small acidification by 0.18 pH units, whereas a more alkaline external pH >7.0 caused a dramatic increase in cytoplasmic pH and growth stopped immediately. Stimulation of the plasma membrane H+-ATPase by fusicoccin, resulted in an increase of tube growth but no change in cytoplasmic pH. On the other hand, vanadate (250–500 microM), a putative inhibitor of the pump, stopped tube growth and a slight cytoplasmic alkalinisation of 0.1 pH units was observed. Vanadate also arrested fusicoccin-stimulated growth and stimulated an increased alkalinisation of around 0.2 pH units. External application of CaCl2 (10 mM) caused a small acidification of less than 0.1 pH units in the clear zone, whilst LaCl3 (250 microM) caused slight and rather variable perturbations in cytoplasmic pH of no more than 0.1 pH units. Both treatments stopped growth. It was inferred from these data that tip-acid cytoplasmic pH gradients do not play a central role in the organisation or maintenance of pollen tube tip growth.

Pronounced cytoplasmic pH gradients are not required for tip growth in plant and fungal cells

1997 ◽  
Vol 110 (10) ◽  
pp. 1187-1198 ◽  
Author(s):  
R.M. Parton ◽  
S. Fischer ◽  
R. Malho ◽  
O. Papasouliotis ◽  
T.C. Jelitto ◽  
...  
Keyword(s):  
Tip Growth ◽  
Cell Types ◽  
Pollen Tubes ◽  
Fine Tuning ◽  
Cytoplasmic Ph ◽  
Dual Emission ◽  
Ph Gradients ◽  
Longitudinal Gradients ◽  
External Ph

The existence of pronounced cytoplasmic pH gradients within the apices of tip-growing cells, and the role of cytoplasmic pH in regulating tip growth, were investigated in three different cell types: vegetative hyphae of Neurospora crassa; pollen tubes of Agapanthus umbellatus; and rhizoids of Dryopteris affinis gametophytes. Examination of cytoplasmic pH in growing cells was performed by simultaneous, dual emission confocal ratio imaging of the pH-sensitive probe carboxy SNARF-1. Considerable attention was paid to the fine tuning of dye loading and imaging parameters to minimise cellular perturbation and assess the extent of dye partitioning into organelles. With optimal conditions, cytoplasmic pH was measured routinely with a precision of between +/−0.03 and +/−0.06 of a pH unit and a spatial resolution of 2.3 microm2. Based on in vitro calibration, estimated values of mean cytoplasmic pH for cells loaded with dye-ester were between 7.15 and 7.25 for the three cell types. After pressure injecting Neurospora hyphae with dextran-conjugated dye, however, the mean cytoplasmic pH was estimated to be 7.57. Dextran dyes are believed to give a better estimate of cytoplasmic pH because of their superior localisation and retention within the cytosol. No significant cytoplasmic pH gradient (delta pH of >0.1 unit) was observed within the apical 50 microm in growing cells of any of the three cell types. Acidification or alkalinisation of the cytoplasm in Neurospora hyphae, using a cell permeant weak acid (propionic acid at pH 7.0) or weak base (trimethylamine at pH 8.0), slowed down but did not abolish growth. However, similar manipulation of the cytoplasmic pH of Agapanthus pollen tubes and Dryopteris rhizoids completely inhibited growth. Modification of external pH affected the growth pattern of all cell types. In hyphae and pollen tubes, changes in external pH were found to have a small transient effect on cytoplasmic pH but the cells rapidly readjusted towards their original pH. Our results suggest that pronounced longitudinal gradients in cytoplasmic pH are not essential for the regulation of tip growth.

scholarly journals Signaling in Pollen Tube Growth: Beyond the Tip of the Polarity Iceberg

Plants ◽  
2019 ◽  
Vol 8 (6) ◽  
pp. 156 ◽  
Author(s):  
Nolan Scheible ◽  
Andrew McCubbin
Keyword(s):  
Pollen Tube ◽  
Signaling Pathways ◽  
Pollen Tube Growth ◽  
Tip Growth ◽  
Holistic Approach ◽  
Pollen Tubes ◽  
Small Gtpase ◽  
Tube Growth ◽  
Concerted Action ◽  
Current State

The coordinated growth of pollen tubes through floral tissues to deliver the sperm cells to the egg and facilitate fertilization is a highly regulated process critical to the Angiosperm life cycle. Studies suggest that the concerted action of a variety of signaling pathways underlies the rapid polarized tip growth exhibited by pollen tubes. Ca2+ and small GTPase-mediated pathways have emerged as major players in the regulation of pollen tube growth. Evidence suggests that these two signaling pathways not only integrate with one another but also with a variety of other important signaling events. As we continue to elucidate the mechanisms involved in pollen tube growth, there is a growing importance in taking a holistic approach to studying these pathways in order to truly understand how tip growth in pollen tubes is orchestrated and maintained. This review considers our current state of knowledge of Ca2+-mediated and GTPase signaling pathways in pollen tubes, how they may intersect with one another, and other signaling pathways involved. There will be a particular focus on recent reports that have extended our understanding in these areas.

scholarly journals Oxygen radicals and cytoplasm zoning in growing lily pollen tubes

2020 ◽  
Author(s):  
Alexandra Podolyan ◽  
Oksana Luneva ◽  
Maria Breygina
Keyword(s):  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Oxygen Radicals ◽  
Redox Regulation ◽  
Pollen Tubes ◽  
Negative Influence ◽  
Tube Growth ◽  
Differential Sensitivity ◽  
Growth Speed ◽  
Ph Gradients

AbstractRecently redox-regulation of tip growth has been extensively studied, but differential sensitivity of growing cells to particular ROS and their subcellular localization is still unclear. Here we used specific dyes to provide mapping of H2O2 and O•2− in short and long pollen tubes. We found apical accumulation of H2O2 and H2O2–producing organelles in the shank that were not colocalized with O•2−-producing mitochondria. Differential modulation of ROS content of the germination medium affected both growth speed and pollen tube morphology. Oxygen radicals affected ionic zoning: membrane potential and pH gradients. OH• caused depolarization all along the tube while O•2− provoked hyperpolarization and cytoplasm alkalinization. O•2− accelerated growth and reduced tube diameter, indicating that this ROS can be considered as pollen tube growth stimulator along with H2O2. Serious structural disturbances were observed upon exposure to OH• and H2O2 and O•2− quencher MnTMPP: pollen tube growth slowed down and ballooned tips formed in both cases, but in the presence of OH• membrane transport and organelle distribution was affected as well. OH•, thus, can be considered as a negative influence on pollen tubes which, presumably, have mechanisms for leveling it. The assumption was confirmed by EPR spectroscopy: pollen tubes actively reduce OH• content in the incubation medium.

scholarly journals Growing Pollen Tubes Possess a Constitutive Alkaline Band in the Clear Zone and a Growth-dependent Acidic Tip

1999 ◽  
Vol 144 (3) ◽  
pp. 483-496 ◽  
Author(s):  
J.A. Feijó ◽  
J. Sainhas ◽  
G.R. Hackett ◽  
J.G. Kunkel ◽  
P.K. Hepler
Keyword(s):  
Pollen Tube ◽  
Intracellular Ph ◽  
Pollen Tube Growth ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Actin Binding ◽  
Wide Field ◽  
Clear Zone ◽  
Direct Role ◽  
Acidic Domain

Using both the proton selective vibrating electrode to probe the extracellular currents and ratiometric wide-field fluorescence microscopy with the indicator 2′,7′-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF)-dextran to image the intracellular pH, we have examined the distribution and activity of protons (H+) associated with pollen tube growth. The intracellular images reveal that lily pollen tubes possess a constitutive alkaline band at the base of the clear zone and an acidic domain at the extreme apex. The extracellular observations, in close agreement, show a proton influx at the extreme apex of the pollen tube and an efflux in the region that corresponds to the position of the alkaline band. The ability to detect the intracellular pH gradient is strongly dependent on the concentration of exogenous buffers in the cytoplasm. Thus, even the indicator dye, if introduced at levels estimated to be of 1.0 μM or greater, will dissipate the gradient, possibly through shuttle buffering. The apical acidic domain correlates closely with the process of growth, and thus may play a direct role, possibly in facilitating vesicle movement and exocytosis. The alkaline band correlates with the position of the reverse fountain streaming at the base of the clear zone, and may participate in the regulation of actin filament formation through the modulation of pH-sensitive actin binding proteins. These studies not only demonstrate that proton gradients exist, but that they may be intimately associated with polarized pollen tube growth.

scholarly journals Arabidopsis ABCG28 is required for the apical accumulation of reactive oxygen species in growing pollen tubes

2019 ◽  
Vol 116 (25) ◽  
pp. 12540-12549 ◽  
Author(s):  
Thanh Ha Thi Do ◽  
Hyunju Choi ◽  
Michael Palmgren ◽  
Enrico Martinoia ◽  
Jae-Ung Hwang ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Tip Growth ◽  
Ectopic Expression ◽  
Reactive Oxygen ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Oxygen Species ◽  
Ros Accumulation

Tip-focused accumulation of reactive oxygen species (ROS) is tightly associated with pollen tube growth and is thus critical for fertilization. However, it is unclear how tip-growing cells establish such specific ROS localization. Polyamines have been proposed to function in tip growth as precursors of the ROS, hydrogen peroxide. The ABC transporter AtABCG28 may regulate ROS status, as it contains multiple cysteine residues, a characteristic of proteins involved in ROS homeostasis. In this study, we found that AtABCG28 was specifically expressed in the mature pollen grains and pollen tubes. AtABCG28 was localized to secretory vesicles inside the pollen tube that moved toward and fused with the plasma membrane of the pollen tube tip. Knocking out AtABCG28 resulted in defective pollen tube growth, failure to localize polyamine and ROS to the growing pollen tube tip, and complete male sterility, whereas ectopic expression of this gene in root hair could recover ROS accumulation at the tip and improved the growth under high-pH conditions, which normally prevent ROS accumulation and tip growth. Together, these data suggest that AtABCG28 is critical for localizing polyamine and ROS at the growing tip. In addition, this function of AtABCG28 is likely to protect the pollen tube from the cytotoxicity of polyamine and contribute to the delivery of polyamine to the growing tip for incorporation into the expanding cell wall.

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.

The distribution of calcium in the grass pollen tube

1985 ◽  
Vol 225 (1240) ◽  
pp. 315-327 ◽  
Keyword(s):  
Pollen Tube ◽  
Pollen Dispersal ◽  
Lilium Longiflorum ◽  
Pollen Tubes ◽  
Distal Region ◽  
Tube Growth ◽  
Growth Physiology ◽  
Large Numbers ◽  
The Difference ◽  
Apical Organization

The distribution of calcium in the terminal regions of actively extending pollen tubes of two Gramineae, Zea mays and Pennisetum americanum , has been investigated by chlorotetracycline (CTC) fluorescence and by energy-dispersive X-ray analysis. Neither method reveals a concentration gradient declining from the tip towards the older parts of the tube comparable with that reported from the pollen tubes of Lilium longiflorum . The difference evidently arises from dissimilarities in pollen-tube growth physiology and the apical organization of the tube. Growth is achieved by the insertion of dictyosome-derived vesicles carrying wallprecursor materials at the tube tip. In L . longiflorum these are produced in a subapical zone of the tube, and a characteristic zonation of cytoplasmic organelles in the distal region develops during growth. In the grasses, large numbers of the wall precursor bodies (‘P-particles ’) are produced before pollen dispersal, and are stored in the grain; they are distributed throughout the tube during early growth, and the organelle zonation in the apex is less pronounced. CTC-induced fluorescence is strongly associated with mitochondria, membranes and P-particles, suggesting that the observed distribution of calcium may reflect mainly the distribution of the element held in organelles and membranes rather than mobile Ca 2+ in the cytosol.

scholarly journals Protein Analysis of Pollen Tubes after the Treatments of Membrane Trafficking Inhibitors Gains Insights on Molecular Mechanism Underlying Pollen Tube Polar Growth

2021 ◽  
Vol 40 (2) ◽  
pp. 205-222
Author(s):  
Monica Scali ◽  
Alessandra Moscatelli ◽  
Luca Bini ◽  
Elisabetta Onelli ◽  
Rita Vignani ◽  
...  
Keyword(s):  
Pollen Tube ◽  
Actin Cytoskeleton ◽  
Membrane Trafficking ◽  
Tip Growth ◽  
Male Gametophyte ◽  
Pollen Tubes ◽  
Structural Features ◽  
Two Dimensional Gel Electrophoresis ◽  
Depth Analysis

AbstractPollen tube elongation is characterized by a highly-polarized tip growth process dependent on an efficient vesicular transport system and largely mobilized by actin cytoskeleton. Pollen tubes are an ideal model system to study exocytosis, endocytosis, membrane recycling, and signaling network coordinating cellular processes, structural organization and vesicular trafficking activities required for tip growth. Proteomic analysis was applied to identifyNicotiana tabacumDifferentially Abundant Proteins (DAPs) after in vitro pollen tube treatment with membrane trafficking inhibitors Brefeldin A, Ikarugamycin and Wortmannin. Among roughly 360 proteins separated in two-dimensional gel electrophoresis, a total of 40 spots visibly changing between treated and control samples were identified by MALDI-TOF MS and LC–ESI–MS/MS analysis. The identified proteins were classified according to biological processes, and most proteins were related to pollen tube energy metabolism, including ammino acid synthesis and lipid metabolism, structural features of pollen tube growth as well modification and actin cytoskeleton organization, stress response, and protein degradation. In-depth analysis of proteins corresponding to energy-related pathways revealed the male gametophyte to be a reliable model of energy reservoir and dynamics.

scholarly journals Hyperoside promotes pollen tube growth by regulating the depolymerization effect of actin-depolymerizing factor 1 on microfilaments in okra

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Biying Dong ◽  
Qing Yang ◽  
Zhihua Song ◽  
Lili Niu ◽  
Hongyan Cao ◽  
...  
Keyword(s):  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Pollen Germination ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Actin Depolymerizing Factor ◽  
Specific Mechanism ◽  
Promoting Effect ◽  
New Research

AbstractMature pollen germinates rapidly on the stigma, extending its pollen tube to deliver sperm cells to the ovule for fertilization. The success of this process is an important factor that limits output. The flavonoid content increased significantly during pollen germination and pollen tube growth, which suggests it may play an important role in these processes. However, the specific mechanism of this involvement has been little researched. Our previous research found that hyperoside can prolong the flowering period of Abelmoschus esculentus (okra), but its specific mechanism is still unclear. Therefore, in this study, we focused on the effect of hyperoside in regulating the actin-depolymerizing factor (ADF), which further affects the germination and growth of pollen. We found that hyperoside can prolong the effective pollination period of okra by 2–3-fold and promote the growth of pollen tubes in the style. Then, we used Nicotiana benthamiana cells as a research system and found that hyperoside accelerates the depolymerization of intercellular microfilaments. Hyperoside can promote pollen germination and pollen tube elongation in vitro. Moreover, AeADF1 was identified out of all AeADF genes as being highly expressed in pollen tubes in response to hyperoside. In addition, hyperoside promoted AeADF1-mediated microfilament dissipation according to microfilament severing experiments in vitro. In the pollen tube, the gene expression of AeADF1 was reduced to 1/5 by oligonucleotide transfection. The decrease in the expression level of AeADF1 partially reduced the promoting effect of hyperoside on pollen germination and pollen tube growth. This research provides new research directions for flavonoids in reproductive development.

scholarly journals Testing of self-(in)compatibility in apricot cultivars from European breeding programmes

2013 ◽  
Vol 40 (No. 2) ◽  
pp. 65-71 ◽  
Author(s):  
D. Milatović ◽  
D. Nikolić ◽  
B. Krška
Keyword(s):  
Fluorescence Microscopy ◽  
Pollen Tube ◽  
Pollen Tube Growth ◽  
Reliable Method ◽  
Pollen Tubes ◽  
Tube Growth ◽  
Self Incompatibility ◽  
Breeding Programmes

Self-(in)compatibility was tested in 40 new apricot cultivars from European breeding programmes. Pollen-tube growth in pistils from laboratory pollinations was analysed using the fluorescence microscopy. Cultivars were considered self-compatible if at least one pollen tube reached the ovary in the majority of pistils. Cultivars were considered self- incompatible if the growth of pollen tubes in the style stopped along with formation of characteristic swellings. Of the examined cultivars, 18 were self-compatible and 22 were self-incompatible. Fluorescence microscopy provides a relatively rapid and reliable method to determine self-incompatibility in apricot cultivars.      

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