scholarly journals Cytoplasmic microtubules and fungal morphogenesis: ultrastructural effects of methyl benzimidazole-2-ylcarbamate determined by freeze-substitution of hyphal tip cells.

1980 ◽  
Vol 87 (1) ◽  
pp. 55-64 ◽  
Author(s):  
R J Howard ◽  
J R Aist
Keyword(s):  
Intracellular Transport ◽  
Freeze Substitution ◽  
Ultrastructural Level ◽  
Spindle Pole ◽  
Voltage Electron ◽  
Spindle Pole Bodies ◽  
Fungal Morphogenesis ◽  
Cytoplasmic Microtubules ◽  
Tip Cells ◽  
Hyphal Tip

The effects of methyl benzimidazole-2-ylcarbamate (MBC), one of only a few agents that are active against microtubules of fungi, were analyzed at the ultrastructural level in freeze-substituted hyphal tip cells of Fusarium acuminatum. Nontreated and control cells had numerous microtubules throughout. After just 10 min of exposure to MBC, almost no cytoplasmic microtubules were present, except near spindle pole bodies. After 45 min of exposure to MBC, no microtubules were present in hyphal tip cells, but they were present in the relatively quiescent subapical cells. These observations suggested that there are different rates of turnover for cytoplasmic microtubules in apical and subapical cells and for microtubules near spindle pole bodies and that MBC acts by inhibiting microtubules assembly. A statistical analysis of the distribution of intracytoplasmic vesicles in thick sections of cells treated with MBC, D2O or MBC + D2O was obtained by use of a high-voltage electron microscope. More than 50% of the vesicles in the apical 30 micrometers of control cells were found to lie within 2 micrometers of the tip cell apex. MBC treatment caused this vesicle distribution to become uniform, resulting in a substantial increase in the number of vesicles in subapical regions. The reduction in the number of cytoplasmic microtubules, induced by MBC, apparently inhibited intracellular transport of these vesicles and rendered random the longitudinal orientation of mitochondria. In most cases, D2O appeared capable of preventing these MBC-effects through stabilization of microtubules. These observations support the "vesicle hypothesis" of tip growth and establish a transport role for cytoplasmic microtubules in fungal morphogenesis.

scholarly journals Mitosis in the fission yeast Schizosaccharomyces pombe as revealed by freeze-substitution electron microscopy

1986 ◽  
Vol 80 (1) ◽  
pp. 253-268
Author(s):  
K. Tanaka ◽  
T. Kanbe
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Mitotic Spindle ◽  
Nuclear Division ◽  
Freeze Substitution ◽  
Spindle Pole ◽  
Nuclear Space ◽  
Spindle Pole Bodies ◽  
Transmission Electron ◽  
Cytoplasmic Microtubules ◽  
Spindle Microtubules

Nuclear division in Schizosaccharomyces pombe has been studied in transmission electron micrographs of sections of cells fixed by a method of freeze-substitution. We have found cytoplasmic microtubules in the vicinity of the spindle pole bodies and two kinds of microtubules, short discontinuous ones and long, parallel ones in the intranuclear mitotic spindle. For most of the time taken by nuclear division the spindle pole bodies face each other squarely across the nuclear space but early in mitosis they briefly appear twisted out of alignment with each other, thereby imparting a sigmoidal shape to the bundle of spindle microtubules extending between them. This configuration is interpreted as indicating active participation of the spindle in the initial elongation of the dividing nucleus. It is proposed that mitosis is accompanied by the shortening of chromosomal microtubules simultaneously with the elongation of the central pole-to-pole bundle of microtubules of the intranuclear spindle. Daughter nuclei are separated by the sliding apart of interdigitating microtubules of the spindle at telophase. Some of the latter bear dense knobs at their ends.

Independent nuclear motility and hyphal tip growth

1995 ◽  
Vol 73 (S1) ◽  
pp. 122-125 ◽  
Author(s):  
James R. Aist
Keyword(s):  
Tip Growth ◽  
Spindle Pole ◽  
Life Cycles ◽  
Microtubule Organizing Center ◽  
Hyphal Tip Growth ◽  
Spindle Pole Bodies ◽  
Organizing Center ◽  
Key Words Actin ◽  
Tip Cells ◽  
Hyphal Tip

Independent nuclear motility is involved in many important aspects of fungal life cycles, including the following: nuclear division; population of hyphal tip cells, branches, and spores with nuclei; dikaryotization; and karyogamy. Spindle pole bodies are almost constantly in motion during all phases of the nuclear cycle, and they have been linked to most instances of independent nuclear motility. A role for microtubules in this process is now well established, and research is being focused on which set of them, astral or cytoplasmic, is utilized as well as on the microtubule-associated motor proteins that may generate the force. In some cases, F-actin may interact with the microtubules or even provide an alternative cytoskeleton supporting nuclear migration. Hyphal tip growth and independent nuclear motility are coordinated and interrelated processes, making the elucidation of the signals, processes, and structures involved an attractive area for further research. Key words: actin, microtubule, microtubule associated protein, microtubule organizing center, motility, nucleus.

scholarly journals Role of fungal dynein in hyphal growth, microtubule organization, spindle pole body motility and nuclear migration

1998 ◽  
Vol 111 (11) ◽  
pp. 1555-1566 ◽  
Author(s):  
S. Inoue ◽  
B.G. Turgeon ◽  
O.C. Yoder ◽  
J.R. Aist
Keyword(s):  
Spindle Pole Body ◽  
Nuclear Migration ◽  
Cytoplasmic Dynein ◽  
Spindle Pole ◽  
Wild Type ◽  
Interphase Nuclei ◽  
Spindle Pole Bodies ◽  
Body Motility ◽  
Tip Cells ◽  
Hyphal Tip

Cytoplasmic dynein is a microtubule-associated motor protein with several putative subcellular functions. Sequencing of the gene (DHC1) for cytoplasmic dynein heavy chain of the filamentous ascomycete, Nectria haematococca, revealed a 4,349-codon open reading frame (interrupted by two introns) with four highly conserved P-loop motifs, typical of cytoplasmic dynein heavy chains. The predicted amino acid sequence is 78.0% identical to the cytoplasmic dynein heavy chain of Neurospora crassa, 70.2% identical to that of Aspergillus nidulans and 24.8% identical to that of Saccharomyces cerevisiae. The genomic copy of DHC1 in N. haematococca wild-type strain T213 was disrupted by inserting a selectable marker into the central motor domain. Mutants grew at 33% of the wild-type rate, forming dense compact colonies composed of spiral and highly branched hyphae. Major cytological phenotypes included (1) absence of aster-like arrays of cytoplasmic microtubules focused at the spindle pole bodies of post-mitotic and interphase nuclei, (2) limited post-mitotic nuclear migration, (3) lack of spindle pole body motility at interphase, (4) failure of spindle pole bodies to anchor interphase nuclei, (5) nonuniform distribution of interphase nuclei and (6) small or ephemeral Spitzenkorper at the apices of hyphal tip cells. Microtubule distribution in the apical region of tip cells of the mutant was essentially normal. The nonuniform distribution of nuclei in hyphae resulted primarily from a lack of both post-mitotic nuclear migration and anchoring of interphase nuclei by the spindle pole bodies. The results support the hypothesis that DHC1 is required for the motility and functions of spindle pole bodies, normal secretory vesicle transport to the hyphal apex and normal hyphal tip cell morphogenesis.

scholarly journals TINA Interacts with the NIMA Kinase in Aspergillus nidulans and Negatively Regulates Astral Microtubules during Metaphase Arrest

2003 ◽  
Vol 14 (8) ◽  
pp. 3169-3179 ◽  
Author(s):  
Aysha H. Osmani ◽  
Jonathan Davies ◽  
C. Elizabeth Oakley ◽  
Berl R. Oakley ◽  
Stephen A. Osmani
Keyword(s):  
Aspergillus Nidulans ◽  
Synthetic Lethality ◽  
Spindle Pole ◽  
Dependent Manner ◽  
Metaphase Arrest ◽  
Enhanced Production ◽  
Spindle Pole Bodies ◽  
In Series ◽  
A Cell ◽  
Cytoplasmic Microtubules

The tinA gene of Aspergillus nidulans encodes a protein that interacts with the NIMA mitotic protein kinase in a cell cycle-specific manner. Highly similar proteins are encoded in Neurospora crassa and Aspergillus fumigatus. TINA and NIMA preferentially interact in interphase and larger forms of TINA are generated during mitosis. Localization studies indicate that TINA is specifically localized to the spindle pole bodies only during mitosis in a microtubule-dependent manner. Deletion of tinA alone is not lethal but displays synthetic lethality in combination with the anaphase-promoting complex/cyclosome mutation bimE7. At the bimE7 metaphase arrest point, lack of TINA enhanced the nucleation of bundles of cytoplasmic microtubules from the spindle pole bodies. These microtubules interacted to form spindles joined in series via astral microtubules as revealed by live cell imaging. Because TINA is modified and localizes to the spindle pole bodies at mitosis, and lack of TINA causes enhanced production of cytoplasmic microtubules at metaphase arrest, we suggest TINA is involved in negative regulation of the astral microtubule organizing capacity of the spindle pole bodies during metaphase.

scholarly journals Ultrastructure of Mitosis and Spindle Pole Bodies in the Zygomycetous Fungus Coemansia reversa Using Conventional Fixation and Freeze Substitution

2014 ◽  
Vol 20 (S3) ◽  
pp. 1292-1293 ◽  
Author(s):  
R.A. Healy ◽  
G.J. Celio ◽  
T.K.A. Kumar ◽  
R.W. Roberson ◽  
D.J. McLaughlin
Keyword(s):  
Freeze Substitution ◽  
Spindle Pole ◽  
Spindle Pole Bodies

Ultrastructural analysis of hyphal tip cell growth in fungi: Spitzenkorper, cytoskeleton and endomembranes after freeze-substitution

1981 ◽  
Vol 48 (1) ◽  
pp. 89-103
Author(s):  
R.J. Howard
Keyword(s):  
Freeze Substitution ◽  
Hyphal Growth ◽  
Coated Vesicle ◽  
Fusarium Acuminatum ◽  
Tip Cell ◽  
Transmission Electron ◽  
Cytoplasmic Vesicles ◽  
Cytoskeletal Elements ◽  
Tip Cells ◽  
Hyphal Tip

The ultrastructure of freeze-substituted tip cells of Fusarium acuminatum was analysed by conventional and high-voltage transmission electron microscopy (HVEM). At least 2 morphologically distinct types of Golgi-like endomembrane cisternae were observed, each existing as single, fenestrated sheets and tubular elements that were often very closely associated with mitochondria. From HVEM observations of thick (0.25 and 0.5 micron) sections, the Spitzenkorper appeared to correspond to an apical mass of vesicles. A network of microfilaments was identified among component vesicles of the Spitzenkorper and adjacent to developing septa. Microtubules were oriented primarily parallel to the direction of hyphal growth and were located in all areas of the cytoplasm, including the tip cell apex. Cytoplasmic vesicles were closely associated with these microtubules. From these observations it is suggested that cytoskeletal elements play important roles in localized cell wall formation. The filasome, a previously unreported type of coated vesicle in fungi, might also be involved in wall synthesis.

Hyphal tip growth and cytoplasmic characters of Conidiobolus coronatus (Zoopagomycota, Entomophthoromycotina)

Mycologia ◽  
2018 ◽  
Vol 110 (1) ◽  
pp. 31-38 ◽  
Author(s):  
Fisher ◽  
Romberger ◽  
Lowry ◽  
Shange ◽  
Roberson
Keyword(s):  
Tip Growth ◽  
Spindle Pole ◽  
Evolutionary Relationships ◽  
Polarized Growth ◽  
Conidiobolus Coronatus ◽  
Hyphal Tip Growth ◽  
Spindle Pole Bodies ◽  
Transmission Electron ◽  
Tem Microscopy ◽  
Hyphal Tip

Characteristics of hyphal structure and growth can provide insights into the mechanisms of polarized growth and support investigations of fungal phylogeny. To assist with the resolution of evolutionary relationships of the zygomycetes, the authors used comparative bioimaging methods (light [LM] and transmission electron [TEM] microscopy) to describe selected subcellular characters of hyphal tips of Conidiobolus coronatus. Growing hyphae of C. coronatus contain Spitzenkörper (Spk). Spk are most commonly present in hyphae of Dikarya (Ascomycota and Basidiomycota) and are rarely reported in zygomycete hyphae, which possess an apical vesicle crescent (AVC). Such findings raise questions regarding the evolution of the Spk and its relationship with the AVC. Descriptions of additional subcellular characters (e.g., mitotic-phase spindle pole bodies, cytoplasmic behavior, organelle structure) are also presented.

Fine structure of an organelle associated with the nucleus and cytoplasmic microtubules in the cellular slime mould Polysphondylium violaceum

1975 ◽  
Vol 18 (2) ◽  
pp. 315-326
Author(s):  
U.P. Roos
Keyword(s):  
Mitotic Spindle ◽  
Spindle Pole ◽  
Interphase Nuclei ◽  
Spindle Pole Bodies ◽  
Nucleation Sites ◽  
Cellular Slime ◽  
Granular Matrix ◽  
Cytoplasmic Microtubules ◽  
Spindle Microtubules ◽  
Opaque Body

Polysphondylium violaceum was grown in association with Escherichia coli. Vegetative amoebae and pseudoplasmodia were fixed under different conditions and processed for electron microscopy. An electron-opaque body (nucleus-associated body, NAB) lies in the cytoplasm near the tapered end of interphase nuclei. The NAB consists of a disk-shaped, multilayered core, approximately 200 nm in diameter and 150 nm thick, embedded in a granular matrix from which electron-opaque nodules protrude. The nodules are termination points of microtubules radiating from the NAB into the cytoplasm or running along the nucleus. On the average there are 16 nodules per NAB. One or two microtubules terminate in each nodule. Spindle pole bodies, arising by duplication of the NAB at the beginning of mitosis, are unstructured foci for spindle microtubules in mitotic cells. It is suggested that cytoplasmic microtubules do not determine cell shape, but they probably cause the tapering deformation of the nucleus. They may, furthermore, represent a storage form of subunits for utilization during the formation of the mitotic spindle. The nodules of the NAB are potential nucleation sites of cytoplasmic microtubules during interphase. Spindle pole bodies presumably acquire a microtubule organizing capability by integration of the decondensed nodules.

The spindle pole body cycle, meiosis, and basidial cytology of the smut fungus Microbotryum violaceum

1991 ◽  
Vol 69 (8) ◽  
pp. 1795-1803 ◽  
Author(s):  
Mary L. Berbee ◽  
Robert Bauer ◽  
F. Oberwinkler
Keyword(s):  
Spindle Pole Body ◽  
Freeze Substitution ◽  
Ustilago Maydis ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Smut Fungus ◽  
Microbotryum Violaceum ◽  
Pole Body ◽  
Spindle Pole Bodies ◽  
Middle Piece

Freeze-substituted basidia of the smut fungus Microbotryum violaceum (Ustilaginales, Basidiomycotina) were examined electron microscopically with particular attention to the meiotic spindle pole body cycle and cytoplasmic characters of phylogenetic significance. Prophase basidia contained a subapical cluster of vesicles and tubules. During prophase, the spindle pole body consisted of two globular elements connected by a middle piece. The spindle pole body had an electron-opaque layer near the nucleus, and each globular element was bisected by an electron-opaque disk. The meiosis I spindle extended between two monoglobular, disc-containing spindle pole bodies. During interphase I and II, septa lacking pores divided the basidium between daughter nuclei. In interphase I, a putative new spindle pole body appeared between the nuclear envelope and the monoglobular spindle pole body residual from the first division. In meiosis II, a spindle was again established between two monoglobular spindle pole bodies, each of which again contained an electron-opaque disc. The cytoplasmic characters of M. violaceum are compared with those of Ustilago maydis and Sphacelotheca polygoni-serrulati. Key words: Microbotryum violaceum, basidiomycete, Ustilaginales, spindle pole body, freeze-substitution, ultrastructure.

The dynamic behaviour of microtubules and their contributions to hyphal tip growth in Aspergillus nidulans

Microbiology ◽  
2005 ◽  
Vol 151 (5) ◽  
pp. 1543-1555 ◽  
Author(s):  
Karina Sampson ◽  
I. Brent Heath
Keyword(s):  
Aspergillus Nidulans ◽  
Tip Growth ◽  
Spindle Pole ◽  
Hyphal Tip Growth ◽  
Complex Processes ◽  
Spindle Pole Bodies ◽  
Tubulin Subunit ◽  
Apical Cells ◽  
Tip Extension ◽  
Hyphal Tip

Creating and maintaining cell polarity are complex processes that are not fully understood. Fungal hyphal tip growth is a highly polarized and dynamic process involving both F-actin and microtubules (MTs), but the behaviour and roles of the latter are unclear. To address this issue, MT dynamics and subunit distribution were analysed in a strain of Aspergillus nidulans expressing GFP–α-tubulin. Apical MTs are the most dynamic, the bulk of which move tipwards from multiple subapical spindle pole bodies, the only clear region of microtubule nucleation detected. MTs populate the apex predominantly by elongation at rates about three times faster than tip extension. This polymerization was facilitated by the tipward migration of MT subunits, which generated a tip-high gradient. Subapical regions of apical cells showed variable tubulin subunit distributions, without tipward flow, while subapical cells showed even tubulin subunit distribution and low MT dynamics. Short MTs, of a similar size to those reported in axons, also occasionally slid into the apex. During mitosis in apical cells, MT populations at the tip varied. Cells with less distance between the tip and the first nucleus were more likely to loose normal MT populations and dynamics. Reduced MTs in the tip, during mitosis or after exposure to the MT inhibitor carbendazim (MBC), generally correlated with reduced, but continuing growth and near-normal tip morphology. In contrast, the actin-disrupting agent latrunculin B reduced growth rates much more severely and dramatically distorted tip morphology. These results suggest substantial independence between MTs and hyphal tip growth and a more essential role for F-actin. Among MT-dependent processes possibly contributing to tip growth is the transportation of vesicles. However, preliminary ultrastructural data indicated a lack of direct MT–organelle interactions. It is suggested that the population of dynamic apical MTs enhance migration of the ‘cytomatrix’, thus ensuring that organelles and proteins maintain proximity to the constantly elongating tip.

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