The effects of microtubule and microfilament disrupting agents on cytoskeletal arrays and wall deposition in developing cotton fibers

Summary The effects of various cytoskeletal disrupting agents (cholchicine, oryzalin, trifluralin, taxol, cytochalasins B and D) on microtubules, microfilaments and wall microfibril deposition were monitored in developing cotton fibers, using immunocytochemical and fluorescence techniques. Treatment with 10^–4 M colchicine, 10^–6 M trifluralin or 10^–6 M oryzalin resulted in a reduction in the number of microtubules, however, the drug-stable microtubules still appear to influence wall deposition. Treatment with 10^–5 M taxol increased the numbers of microtubules present within 15 minutes of application. New microtubules were aligned parallel to the existing ones, however, some evidence of random arrays was observed. Microtubules stabilized with taxol appeared to function in wall organization but do not undergo normal re-orientations during development. Microtubule disrupting agent had no detectable affect on the microfilament population. Exposure to either 4×10^–5 M cytochalasin B or 2×10^–6M cytochalasin D resulted in a disruption of microfilaments and a re-organization of microtubule arrays. Treatment with either cytochalasin caused a premature shift in the orientation of microtubules in young fibers, whereas in older fibers the microtubule arrays became randomly organized. These observations indicate that microtubule populations during interphase are heterogeneous, differing at least in their susceptibility to disruption by depolymerizing agents. Changes in microtubule orientation (induced by cytochalasin) indicate that microfilaments may be involved in regulating microtubule orientation during development.     

Hypersensitivity to cytoskeletal antagonists demonstrates microtubule-microfilament cross-talk in the control of root elongation in Arabidopsis thaliana

Elongation of diffusely expanding plant cells is thought to be mainly under the control of cortical microtubules. Drug treatments that disrupt actin microfilaments, however, can reduce elongation and induce radial swelling. To understand how microfilaments assist growth anisotropy, we explored their functional interactions with microtubules by measuring how microtubule disruption affects the sensitivity of cells to microfilament-targeted drugs. We assessed the sensitivity to actin-targeted drugs by measuring the lengths and diameters of expanding roots and by analysing microtubule and microfilament patterns in the temperature-sensitive Arabidopsis thaliana mutant microtubule organization 1 (mor1-1), along with other mutants that constitutively alter microtubule arrays. At the restrictive temperature of mor1-1, root expansion was hypersensitive to the microfilament-disrupting drugs latrunculin B and cytochalasin D, while immunofluorescence microscopy showed that low doses of latrunculin B exacerbated microtubule disruption. Root expansion studies also showed that the botero and spiral1 mutants were hypersensitive to latrunculin B. Hypersensitivity to actin-targeted drugs is a direct consequence of altered microtubule polymer status, demonstrating that cross-talk between microfilaments and microtubules is critical for regulating anisotropic cell expansion.     

Effects of four agents that modify microtubules and microfilaments (vinblastine,colchicine, lidocaine,and cytochalasin B) on macrophage-mediated cytotoxicity to tumor cells

Summary The effects of vinblastine, colchicine, lidocaine, and cytochalasin B on tumor cell killing by BCG-activated macrophages were examined. These four drugs were selected for their action on membrane-associated cytoskeletal components, microtubules, and microfilaments. Colchicine and vinblastine, which block microtubular synthesis, inhibit macrophage-mediated tumor-cell cytotoxicity at a concentration of 10^–6 M. Cytochalasin B, which disrupts microfilaments, enhances tumor cell lysis and stasis due to activated macrophages at a concentration of 10^–7 M. Lidocaine, which may induce the disappearance of both microtubules and microfilaments, has the same inhibiting effect as vinblastine at a concentration of 5×10^–7 M. Whereas vinblastine and lidocaine seem to act on the macrophage itself, cytochalasin B exerts its effect predominantly on the tumor cell. These results suggest that microtubules and microfilaments play a role in the destruction of tumor cells by activated macrophages.     

Microtubules regulate aortic endothelial cell actin microfilament reorganization in intact and repairing monolayers

To understand the role of microtubules and microfilaments in regulating endothelial monolayer integrity and repair, and since microtubules and microfilaments show some co-alignment in endothelial cells, we tested the hypothesis that microtubules organize microfilament distribution. Disruption of microtubules with colchicine in resting confluent aortic endothelial monolayers resulted in disruption of microfilament distribution with a loss of dense peripheral bands, an increase in actin microfilament bundles, and an associated increase of focal adhesion proteins at the periphery of the cells. However, when microfilaments were disrupted with cytochalasin B, microtubule distribution did not change. During the early stages of wound repair of aortic endothelial monolayers, microtubules and microfilaments undergo a sequential series of changes in distribution prior to cell migration. They are initially distributed randomly relative to the wound edge, then align parallel to the wound edge and then elongate perpendicular to the wound edge. When microtubules in wounded cultures were disrupted, dense peripheral bands and lamellipodia formation were lost with increases in central stress fibers. However, following microfilament disruption, microtubule redistribution was not disrupted and the microtubules elongated perpendicular to the wound edge similar to non-treated cultures. Microtubules may organize independently of microfilaments while microfilaments require microtubules to maintain normal organization in confluent and repairing aortic endothelial monolayers.     

Formation and maintenance of viroplasmic centers in Tipula iridescent virus-infected mosquito cells with deranged cytoskeletons

We have examined the role of cytoskeletal elements with respect to the formation and maintenance of viroplasmic centers (VCs) in Tipula iridescent virus (TIV)-infected mosquito Aedes albopictus (C6/36) cells. Filamentous systems consisting of microtubules and microfilaments were detected by immunofluorescence microscopy. Inoculation of cells with TIV resulted in an alteration of microtubule and microfilament organization whether or not VCs developed. The formation of short arrays of microtubules induced by taxol or the depolymerization of microtubules by colchicine, as observed by immunofluorescence microscopy, had no apparent effect upon the development of VCs as detected by Hoechst staining and electron microscopy. The dissolution of the actin-containing filamentous system by cytochalasin B also had no effect upon development. We conclude from these results that microtubules and microfilaments are not involved in the formation or maintenance of VCs in TIV-infected A. albopictus (C6/36) cells.     

Reactivation of Ca2+-dependent cytoplasmic contraction in permeabilized cell models of the heliozoon Echinosphaerium akamae

Permeabilized cell models of the large heliozoon Echinosphaerium akamae were prepared by treatment with 100 mM EGTA or 1% Triton X-100. When > 10(-6) M Ca(2+) was added to the EGTA-permeabilized cells, axopodial cytoplasm became contracted and several swellings were formed along the axopodial length. Axonemal microtubules remained intact, while higher concentration of Ca(2+) (> 10(-4) M) induced microtubule disassembly and complete breakdown of the axopodia. In Triton-permeabilized cells, cytoplasmic contraction and relaxation of the cell body were induced repeatedly by successive addition and removal of Ca(2+). The contraction did not require ATP, and was not inhibited by cytochalasin B. Electron microscopy showed, in EGTA-permeabilized axopodia, contractile tubules became granulated by the addition of Ca(2+). From these observations, it is strongly suggested that Ca(2+)-dependent granulation of the contractile tubules is responsible for the axopodial contraction.     

The distribution and requirements of microtubules and microfilaments in bovine oocytes during in vitro maturation

Microtubules and microfilaments are major cytoskeletal components and important modulators for chromosomal movement and cellular division in mammalian oocytes. In this study we observed microtubule and microfilament organisation in bovine oocytes by laser scanning confocal microscopy, and determined requirements of their assembly during in vitro maturation. After germinal vesicle breakdown, small microtubular asters were observed near the condensed chromatin. The asters appeared to elongate and encompass condensed chromatin particles. At the metaphase stage, microtubules were observed in the second meiotic spindle at the metaphase stage. The meiotic spindle was a symmetrical, barrel-shaped structure containing anastral broad poles, located peripherally and radially oriented. Treatment with nocodazole did not inhibit germinal vesicle breakdown. However, progression to metaphase failed to occur in oocytes treated with nocodazole. In contrast, microfilaments were observed as a relatively thick uniform area around the cell cortex and overlying chromatin following germinal vesicle breakdown. Treatment with cytochalasin B inhibited microfilament polymerisation but did not prevent either germinal vesicle breakdown or metaphase formation. However, movement of chromatin to the proper position was inhibited in oocytes treated with cytochalasin B. These results suggest that both microtubules and microfilaments are closely associated with reconstruction and proper positioning of chromatin during meiotic maturation in bovine oocytes.     

Microfilaments and microtubules in calcium ionophore-induced secretion of lysosomal enzymes from human polymorphonuclear leukocytes.

Human peripheral blood leukocytes (PMN) are induced to release lysosomal enzymes by the calcium ionophore A23187 in the presence but not the absence of extracellular Ca++. Whereas secretion induced by particulate or immune stimuli is accompanied by an increase in visible microtubules and is inhibitable by colchicine, secretion induced by A23187 and Ca++ was not accompanied by an increase in microtubule numbers and was not inhibited by colchicine. Ca++ did not appear to regulate microtubule assembly in these cells since resting PMN had a mean of 22.3 +/- 2.0 microtubules in the centriolar region as compared to 22.3 +/- 1.1 in ionophore-treated cells and 24.9 +/- 1.5 in cells exposed to ionophore and 1 mM Ca++. Bipolar filaments, 10 nm thick and 300--400 nm long, were numerous in the pericortical cytoplasm of cells exposed to both reagents. Microtubules in these cells were decorated with an electron-opaque fibrillar material. PMN exposed to A23187 and Ca++ were contracted in two directions at right angles to each other: (a) Contractions parallel to the plasma membrane resulted in extensive plication of the cell membrane. The cytoplasm subjacent to the plicae contained dense filamentous webs. Plication was prevented by cytochalasin B or reversed by subsequent exposure to an endocytic stimulus such as zymosan. (b) Contractions perpendicular to the plasma membrane, toward the cytocenter, resulted in the formation of vacuoles in normal PMN and of membrane invaginations in cytochalasin B-treated PMN. Whereas contractions parallel to the plasma membrane could occur in the absence of enzyme release (ionophore alone) and enzyme release could occur in the absence of such contractions (ionophore plus calcium plus cytochalasin B), contraction toward the cytocenter occurred in all experimental conditions in which significant enzyme release was obtained. Thus, lysosomal enzyme secretion in PMN involves contractile movements in the plasma membrane toward the lysosomes rather than the reverse. These calcium-mediated contractile events are mediated by cytochalasin B-insensitive microfilaments but not by microtubule assembly.     

10 nm filaments in normal and transformed cells.

An antibody was raised against an electrophoretically homogeneous protein from cultured fibroblasts and shown to be directed against 10 nm filaments. The antiserum did not stain microtubules or actin microfilaments. The distribution of 10 nm filaments in normal cells was studied during growth, spreading, locomotion, mitosis, and after treatment with colchicine and cytochalasin B. The 58,000 dalton subunit protein is apparently all polymerized in the filaments which are insoluble in nonionic detergent. The distribution of 10 nm filaments is altered by colchicine treatments which disrupt microtubules. The organization of 10 nm filaments is altered in transformed cells.     

Phorbol myristate acetate stimulates microtubule and 10-nm filament extension and lysosome redistribution in mouse macrophages

Phorbol myristate acetate (PMA) stimulates cell spreading and fluid- phase pinocytosis in mouse peritoneal macrophages. Colchicine (10(-5) M) and cytochalasin B (10(-5) M) abolish PMA stimulated pinocytosis but have little effect on cellular spreading (Phaire-Washington et al., 1980, J. Cell Biol., 86:634-640). We report here that PMA also alters the organization of the cytoskeleton and the distrubution of organelles in these cells. Neither control nor PMA-treated macrophages contain actin cables. PMA-treated resident thioglycolate-elicited macrophages exhibit beneath their substrate-adherent membranes many randomly distributed punctate foci that stain brightly for actin. The appearance and distribution of these actin-containing foci are not altered by colchicine (10(-5) M) or cytochalasin B (10(-5) M). In thioglycolate- elicited macrophages PMA causes the extension and radial organization of microtubules and 10-nm filaments and promotes the movement of secondary lysosomes from their perinuclear location to the peripheral cytoplasm. Depending upon the concentration of PMA used, 45-71% of thioglycolate-elicited macrophages and 32-44% of proteose-peptone- elicited macrophages and numerous lysosomes, radiating from the centrosphere region, arranged linearly along microtubule and 10-nm filament bundles. Colchicine (10(-5) M) and podophyllotoxin (10(-5) M) prevent the radial redistribution of microtubules, 10-nm filaments, and lysosomes in these cells. Cytochalasins B and D (10(-5) M) have no inhibitory effects on these processes. These findings indicate that microtubules and 10-nm filaments respond in a coordinated fashion to PMA and to agents that inhibit microtubule function; they suggest that these cytoskeletal elements regulate the movement and distribution of lysosomes in the macrophage cytoplasm.     

Rearrangement of the actin filament and microtubule cytoskeleton during induction of microspore embryogenesis inBrassica napus L. cv. Topas

Summary Changes in the actin filament and microtubule cytoskeleton were examined during heat- and cytochalasin D-induced embryogenesis in microspores ofBrassica napus cv. Topas by rhodamine phalloidin and immunofluorescence labelling respectively. The nucleus was displaced from its peripheral to a more central position in the cell, and perinuclear actin microfilaments and microtubules extended onto the cytoplasm. Heat treatment induced the formation of a preprophase band of microtubules in microspores; preprophase bands are not associated with the first pollen mitosis. Actin filament association with the preprophase band was not observed. The orientation and position of the mitotic spindle were altered, and it was surrounded with randomly oriented microfilaments. The phragmoplast contained microfilaments and microtubules, as in pollen mitosis I, but it assumed a more central position. Cytoskeletal reorganisation also occurred in microspores subjected to a short cytochalasin D treatment, in the absence of a heat treatment. Cytochalasin D treatment of microspores resulted in dislocated mitotic spindles, disrupted phragmoplasts, and symmetric divisions and led to embryogenesis, confirming that a normal actin cytoskeleton has a role in preventing the induction of embryogenesis.Abbreviations CD cytochalasin D - MF actin microfilament - MT microtubule - PPB preprophase band     

Actin microfilaments and fibroin secretion in the silkgland cells of Bombyx mori : Effects of cytochalasin B

Bombyx mori posterior silkgland cells exhibit an impressive microfilament apparatus located at the cellular apex. It consists of bundles of packed, long microfilaments of 50–70 Å diameter running along circumferences delimiting the lumen of the gland, perpendicularly to the flow of luminal silk. Microfilaments are closely associated with microtubules of the cytoplasmic ‘radial microtubule system’. Immunolabelling with purified antihuman actin antibodies was used to demonstrate their actin-like nature. Apical microfilaments are sensitive to cytochalasin B (CB) which selectively inhibits the secretion of fibroin. Following the removal of the drug, microfilaments recover their normal morphology and secretion resumes. The possible implication of contraction of microfilaments in the process of secretion is discussed.     

The Cytoskeleton and Nuclear Disassembly during Germinal Vesicle Breakdown in Starfish Oocytes

In response to maturation-inducing hormone, prophase-arrested oocytes of the starfish Pisaster ochraceus resume meiosis and undergo nuclear disassembly during a process referred to as germinal vesicle breakdown (GVBD). Time-lapse video recordings of maturing oocytes reveal that the nucleus lengthens along the animal-vegetal axis of the oocyte directly prior to GVBD. Neither taxol (10 μM) nor microtubule-depolymerizing agents [colcemid (50 μM), colchicine (250 μM), or nocodazole (1 μM)] prevent the pre-GVBD changes in nuclear shape from occurring, although correlative microscopical studies demonstrate that microtubules are nucleated (taxol) or depolymerized (colcemid, colchicine, nocodazole) at the concentrations listed above. The microtubule-altering drugs also do not affect the time at which GVBD begins or ends. A 10 μM solution of the microfilament-disrupting drug cytochalasin B (CB), on the other hand, essentially eliminates the pre-GVBD elongation of the nucleus. CB also slightly delays the onset of GVBD and significantly lengthens the time required to complete GVBD. Such studies suggest that: (i) drug-sensitive microtubules are not required for GVBD to proceed in a normal fasion; (ii) the pre-GVBD changes in nuclear shape involve microfilament-mediated events; and (iii) cytochalasin-induced depolymerization of microfilaments retards the normal timing of GVBD.     

The cytoskeleton and rat granulosa cell steroidogenesis: possible involvement of microtubules and microfilaments

The participation of both microtubules and microfilaments in granulosa cell steroidogenesis was assessed by monitoring the effects of colchicine (0-250 microM) and/or cytochalasin B (0-10 micrograms/ml) or dihydrocytochalasin B (0-2.0 micrograms/ml) on cellular morphology and production of progestins during 24 h of culture. Both colchicine and the cytochalasins increased granulosa cell production of progesterone and of 20 alpha-hydroxy-pregn-4-en-3-one (20 alpha-OH-progesterone) in a dose-dependent manner. The largest increase in steroidogenesis (about 2- to 3-fold) was observed at 4-250 microM colchicine and at 2-10 micrograms/ml cytochalasin. Those concentrations of the inhibitors of microtubule or microfilament polymerization that stimulated basal progestin production also markedly influenced cell spreading. Whereas cells cultured for 24 h in medium alone became very flattened with numerous cytoplasmic extensions, those cultured with colchicine (0.2-250 microM) or cytochalasin (0.4-2 micrograms/ml) were much less spread and progressively became more rounded and regular in outline. These changes in cell morphology were reflected by decreases in the mean area occupied by the cells on the culture surface of up to 60-65% and reductions in mean contour index values from 5.7 +/- 0.1 (control) to 3.9 +/- 0.1 (250 microM colchicine), 4.2 +/- 0.1 (2 micrograms/ml cytochalasin B), or 4.1 +/- 0.1 (2 micrograms/ml dihydrocytochalasin B). Cultures containing both colchicine and cytochalasin B exhibited a greater steroidogenic response than that elicited by either inhibitor alone. For example, granulosa cell progesterone production was stimulated almost 2-fold by 4 microM colchicine or 2 microM/ml cytochalasin B, but 5.5-fold by 4 microM colchicine plus 2 micrograms/ml cytochalasin B.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of cytochalasin B and colchicine on the morphogenesis of mitochondria in Drosophila hydei during meiosis and early spermiogenesis

Summary Morphogenesis of mitochondria in male germ cells in cultivated cytocysts begins in early prophase I at which time mitochondria thicken and become ordered along the spindle apparatus during meiosis. At the end of the second meiotic division they aggregate to form the Nebenkern.In the presence of colchicine or cytochalasin B mitochondria are able to begin differentiation, although the correct course of meiosis is not guaranteed. In medium supplemented with colchicine they undergo normal thickening but do not aggregate, in a pattern known from untreated cultures. This may indicate that microtubules are involved in the aggregation process of mitochondria as colchicine is known to inhibit microtubule formation. Moreover, in cell cultures treated with cytochalasin B mitochondrial aggregation does occur; it is concluded that microfilaments, which are sensitive to cytochalasin B, do not play a detectable role in the aggregation of mitochondria.     

Structural analysis of human neutrophil migration: Centriole, microtubule, and microfilament orientation and function during chemotaxis

Orientation of nucleus, centriole, microtubules, and microfilaments within human neutrophils in a gradient of chemoattractant (5 percent Escherichia coli endotoxin-activated serum) was evaluated by electron microscopy. Purified neutropils (hypaque-Ficoll) were placed in the upper compartment of chemotactic chambers. Use of small pore (0.45 μm) micropore filters permitted pseudopod penetration, but impeded migration. Under conditions of chemotaxis with activated serum beneath the filter, the neutrophil population oriented at the filter surface with nuclei located away from the stimulus, centrioles and associated radial array of microtubules beneath the nuclei, and microfilament-rich pseudopods penetrating the filter pores. Reversal of the direction of the gradient of the stimulus (activated serum above cells) resulted in a reorientation of internal structure which preceded pseudopod formation toward the activated serum and migration off the filter. Coordinated orientation of the entire neutrophil population did not occur in buffer (random migration) or in a uniform concentration of activated serum (activated random migration). Conditions of activated random migration resulted in increased numbers of cells with locomotory morphology, i.e. cellular asymmetry with linear alignment of nucleus, centriole, microtubule array, and pseudopods. Thus, activated serum increased the number of neutrophils exhibiting locomotory morphology, and a gradient of activated serum induced the alignment of neutrophils such that this locomotory morphology was uniform in the observed neutrophil populayion. In related studies, cytochalasin B and colchicines were used to explore the role of microfilaments and microtubules in the neutrophil orientation and migration response to activated serum. Cytochalasin B (3.0 μg/ml) prevented migration and decreased the microfilaments seen, but allowed normal orientation of neutrophil structures. In an activated serum gradient, colchicines, but not lumicolchicine, decreased the orientation of nuclei and centrioles, and caused a decrease in centriole-associated microtubules in concentrations as low as 10(-8) to 10(-7) M. These colchicines effects were associated with the rounding of cells and impairment of pseudopod formation. The impaired pseudopod formation was characterized by an inability to form pseudopods in the absence of a solid substrate, a formation of narrow pseudopods within a substrate, and a defect in pseudopod orientation in an activated serum gradient. Functional studies of migration showed that colchicines, but not lumicolchicine, minimally decreased activated random migration and markedly inhibited directed migration, but had not effect on random migration. These studies show that, although functioning microfilaments are probably necessary for neutrophil migration, intact microtubules are essential for normal pseudopod formation and orientation, and maximal unidirectional migration during chemotaxis.     

Actin microfilaments facilitate the retrograde transport from the Golgi complex to the endoplasmic reticulum in mammalian cells

The morphology and subcellular positioning of the Golgi complex depend on both microtubule and actin cytoskeletons. In contrast to microtubules, the role of actin cytoskeleton in the secretory pathway in mammalian cells has not been clearly established. Using cytochalasin D, we have previously shown that microfilaments are not involved in the endoplasmic reticulum–Golgi membrane dynamics. However, it has been reported that, unlike botulinum C2 toxin and latrunculins, cytochalasin D does not produce net depolymerization of actin filaments. Therefore, we have reassessed the functional role of actin microfilaments in the early steps of the biosynthetic pathway using C2 toxin and latrunculin B. The anterograde endoplasmic reticulum-to-Golgi transport monitored with the vesicular stomatitis virus-G protein remained unaltered in cells treated with cytochalasin D, latrunculin B or C2 toxin. Conversely, the brefeldin A-induced Golgi membrane fusion into the endoplasmic reticulum, the Golgi-to-endoplasmic reticulum transport of a Shiga toxin mutant form, and the subcellular distribution of the KDEL receptor were all impaired when actin microfilaments were depolymerized by latrunculin B or C2 toxin. These findings, together with the fact that COPI-coated and uncoated vesicles contain β/γ-actin isoforms, indicate that actin microfilaments are involved in the endoplasmic reticulum/Golgi interface, facilitating the retrograde Golgi-to-endoplasmic reticulum membrane transport, which could be mediated by the orchestrated movement of transport intermediates along microtubule and microfilament tracks.     

Effect of cytochalasin B and colchicine on the stimulation of -amylase release from rat parotid tissue slices

A role for microfilaments and microtubules in the secretion of α-amylase is indicated since cytochalasin B and colchicine inhibited the stimulation of α-amylase release by epinephrine (30 or 15 μM) but only cytochalasin B inhibited the stimulation by N⁶, O^2′ dibutyryl adenosine 3′,5′monophosphate (1.0 mM). It was necessary to incubate the parotid tissue slices in the presence of cytochalasin B (1 hr.) or colchicine (4 hrs.) before adding the agonist in order to see the inhibitory effects.     

Changes of the actin filament system in the green algaMicrasterias denticulata induced by different cytoskeleton inhibitors

Summary Two different techniques have been adapted forMicrasterias denticulata to depict the actin cytoskeleton of both untreated and inhibitor-treated developing cells: the quickstaining method, where the cells are fixed in a mixture of glutaraldehyde and formaldehyde followed by staining with phalloidin without embedding, and the methacrylate method, where the cells are also fixed by aldehydes and where the embedding medium is removed prior to incubation with an actin antibody. Both methods produce sufficient preservation and visualization of actin microfilaments (MFs) and confirm earlier observations on the presence of a cortical actin MF network in both the growing and the nongrowing semicell as well as of a basketlike MF arrangement around the migrating nucleus. The results show that a network of actin MFs is essential for the proper development of the young lobes ofM. denticulata. Early developmental stages expanding uniformly at the beginning of growth lack any netlike actin MF arrangement. The actin cytoskeleton in developing cells treated with the actin-targeting agents cytochalasin D and latrunculin B is markedly influenced. Cytochalasin D, which produces the most pronounced effects, causes a breakdown of the network of actin MFs, resulting in bright actin clusters as well as in short and abnormally thick actin fragments particularly in cortical cell regions. In latrunculin B-treated cells remnants of the former actin MF network are still visible, yet most of the actin cytoskeleton appears collapsed and is reduced to short filament pieces. The disturbance of the actin MF system visualized in the present study correlates with the severe morphological and ultrastructural changes occurring in desmid cells as a consequence of both drugs. The dinitroanilin herbicide oryzalin, known to deploymerize cytoplasmic microtubules, causes also an impairment of the actin cytoskeleton inM. denticulata though not sufficient to influence normal cell growth and differentiation.Abbreviations CB cytochalasin B - CD cytochalasin D - DMSO dimethyl sulfoxide - FA formaldehyde - GA glutaraldehyde - LAT-A latrunculin A - LAT-B latrunculin B - MFs microfilaments - MT microtubule Dedicated to Professor Walter Gustav Url on the occasion of his 70th birthday     

Influence of microfilament and microtubule inhibitors applied by immersion and microinjection on circulation streaming in the staminal hairs ofTradescantia blossfeldiana

Summary Reaction of cytoplasmic streaming inTradescantia staminal hairs to microfilament and microtubule specific inhibitors, applied either by conventional immersion or by microinjection, indicates that both the actin-myosin and the microtubule system may be involved in driving the particle stream. Cytoplasmic streaming was stopped at relatively high drug concentrations when the cells were immersed in the inhibitor solution. Microinjection of defined concentrations of inhibitor into single, selected cells were effective at concentrations at least two orders of magnitude lower. Further reduction of inhibitor concentrations, however, enhanced streaming up to 100%. When a mixture of cytochalasin D and oryzalin were injected at concentrations that had previously been shown to enhance particle movement, very efficient inhibition occurred and streaming rapidly stopped. Adjacent cells on both sides of the injected cell were also affected: within a few minutes of the injection of microfilament inhibitors the basal cell reacted, followed slightly later by the apical one; microtubule inhibitors caused a reaction in the apical cell earlier than in the basal cell. The results are discussed with respect to the involvement of actin and myosin microfilaments, as well as microtubules, as force generating systems of particle movement.Abbreviations CB cytochalasin B - CD cytochalasin D - Cys cysteine - DMSO dimethylsulfoxid - DTT dithiothreitol - MI microinjection - NBD 7-nitrobenz-2-oxa-1,3-diazole - NEM N-ethylmaleimide Nocodazole methyl [5-(2-thienylcarbonyl)-1 H-benzimidazol-2-yl]carbamate