Development of subprotoplasts from in vitro-grown tobacco pollen tubes

Tobacco pollen tubes were used as a standard in vitro system to investigate cell growth aberrations caused by some of the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) programme chemicals and other toxic compounds. Changes in cytoskeletal pattern were observed in the tube cells by using tubu-lin immunofluorescence and rhodamin–phalloidin fluorescence for the localisation of microtubules and actin filaments, respectively. Four different types of cell malformation were found: screw-like growth, isodiametric tip swelling, hook formation, and pollen grain enlargement. We suggest that these malformations resulted from an interference by the chemicals with the cytosolic calcium gradient which controls tip growth and the orientation of the pollen tube. The results may contribute to a general understanding of toxicity-based cell malformations.

Download Full-text

Toxicity of some herbicides to in vitro growing tobacco pollen tubes (the pollen test)

Environmental and Experimental Botany ◽

10.1016/0098-8472(91)90073-w ◽

1991 ◽

Vol 31 (2) ◽

pp. 217-222 ◽

Cited By ~ 14

Author(s):

K. Strube ◽

D. Janke ◽

R. Kappler ◽

U. Kristen

Keyword(s):

Pollen Tubes ◽

Tobacco Pollen

Download Full-text

Cytosolic proteins from tobacco pollen tubes that crosslink microtubules and actin filaments in vitro are metabolic enzymes

Cytoskeleton ◽

10.1002/cm.20483 ◽

2010 ◽

Vol 67 (12) ◽

pp. 745-754 ◽

Cited By ~ 10

Author(s):

Silvia Romagnoli ◽

Claudia Faleri ◽

Luca Bini ◽

Tobias I. Baskin ◽

Mauro Cresti

Keyword(s):

Actin Filaments ◽

Metabolic Enzymes ◽

Pollen Tubes ◽

Cytosolic Proteins ◽

Tobacco Pollen

Download Full-text

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

Biochemical Journal ◽

10.1042/bj20080693 ◽

2008 ◽

Vol 414 (1) ◽

pp. 43-52 ◽

Cited By ~ 24

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.

Download Full-text

Localization of profilin- and actin-like immunoreactivity in in vitro-germinated tobacco pollen tubes by electron microscopy after special water-free fixation techniques

Sexual Plant Reproduction ◽

10.1007/bf00242264 ◽

1995 ◽

Vol 8 (3) ◽

Cited By ~ 16

Author(s):

Monika Grote ◽

Ines Swoboda ◽

RichardB. Meagher ◽

Rudolf Valenta

Keyword(s):

Electron Microscopy ◽

Pollen Tubes ◽

Tobacco Pollen ◽

Fixation Techniques

Download Full-text

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

The Protein Journal ◽

10.1007/s10930-021-09972-x ◽

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.

Download Full-text

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

Horticulture Research ◽

10.1038/s41438-021-00578-z ◽

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.

Download Full-text

Control of the developmental pathway of tobacco pollen in vitro

Planta ◽

10.1007/bf00392260 ◽

1986 ◽

Vol 168 (4) ◽

pp. 427-432 ◽

Cited By ~ 121

Author(s):

M. Kyo ◽

H. Harada

Keyword(s):

Developmental Pathway ◽

Tobacco Pollen

Download Full-text

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

Journal of Cell Science ◽

10.1242/jcs.110.10.1187 ◽

1997 ◽

Vol 110 (10) ◽

pp. 1187-1198 ◽

Cited By ~ 3

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.

Download Full-text