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1.
《Biophysical journal》2022,121(7):1246-1265
Cytotoxic T lymphocytes (T cells) and natural killer cells form a tight contact, the immunological synapse (IS), with target cells, where they release their lytic granules containing perforin/granzyme and cytokine-containing vesicles. During this process the cell repolarizes and moves the microtubule organizing center (MTOC) toward the IS. In the first part of our work we developed a computational model for the molecular-motor-driven motion of the microtubule cytoskeleton during T cell polarization and analyzed the effects of cortical-sliding and capture-shrinkage mechanisms. Here we use this model to analyze the dynamics of the MTOC repositioning in situations in which 1) the IS is in an arbitrary position with respect to the initial position of the MTOC and 2) the T cell has two IS at two arbitrary positions. In the case of one IS, we found that the initial position determines which mechanism is dominant and that the time of repositioning does not rise monotonously with the MTOC-IS distance. In the case of two IS, we observe several scenarios that have also been reported experimentally: the MTOC alternates stochastically (but with a well-defined average transition time) between the two IS; it wiggles in between the two IS without transiting to one of the two; or it is at some point pulled to one of the two IS and stays there. Our model allows one to predict which scenario emerges in dependency of the mechanisms in action and the number of dyneins present. We report that the presence of capture-shrinkage mechanism in at least one IS is necessary to assure the transitions in every cell configuration. Moreover, the frequency of transitions does not decrease with the distance between the two IS and is the highest when both mechanisms are present in both IS. 相似文献
2.
《Biophysical journal》2019,116(11):2079-2091
Upon contact with antigen-presenting cells, cytotoxic T lymphocytes (T cells) establish a highly organized contact zone denoted as the immunological synapse (IS). The formation of the IS implies relocation of the microtubule organizing center (MTOC) toward the contact zone, which necessitates a proper connection between the MTOC and the IS via dynamic microtubules (MTs). The efficiency of the MTs finding the IS within the relevant timescale is, however, still illusive. We investigate how MTs search the three-dimensional constrained cellular volume for the IS and bind upon encounter to dynein anchored at the IS cortex. The search efficiency is estimated by calculating the time required for the MTs to reach the dynein-enriched region of the IS. In this study, we develop simple mathematical and numerical models incorporating relevant components of a cell and propose an optimal search strategy. Using the mathematical model, we have quantified the average search time for a wide range of model parameters and proposed an optimized set of values leading to the minimal capture time. Our results show that search times are minimal when the IS formed at the nearest or at the farthest sites on the cell surface with respect to the perinuclear MTOC. The search time increases monotonically away from these two specific sites and is maximal at an intermediate position near the equator of the cell. We observed that search time strongly depends on the number of searching MTs and distance of the MTOC from the nuclear surface. 相似文献
3.
《Cell cycle (Georgetown, Tex.)》2013,12(14):2168-2174
Cell migration requires polarization of the cell into the leading edge and the trailing edge. Microtubules (MTs) are indispensable for polarized cell migration in the majority of cell types. To support cell polarity, MT network has to be functionally and structurally asymmetric. How is this asymmetry achieved? In interphase cells, MTs form a dynamic system radiating from a centrosome-based MT-organizing center (MTOC) to the cell edges. Symmetry of this radial array can be broken according to four general principles. Asymmetry occurs due to differential modulation of MT dynamics, relocation of existing MTs within a cell, adding an asymmetric nucleation site, and/or repositioning of a symmetric nucleation site to one side of a cell. Combinations of these asymmetry regulation principles result in a variety of asymmetric MT networks typical for diverse motile cell types. Importantly, an asymmetric MT array is formed at a non-conventional MT nucleation site, the Golgi. Here, we emphasize the contribution of this array to the asymmetry of MT network. 相似文献
4.
Jason Yi Xufeng Wu Andrew H. Chung James K. Chen Tarun M. Kapoor John A. Hammer 《The Journal of cell biology》2013,202(5):779-792
T cells rapidly reposition their centrosome to the center of the immunological synapse (IS) to drive polarized secretion in the direction of the bound target cell. Using an optical trap for spatial and temporal control over target presentation, we show that centrosome repositioning in Jurkat T cells exhibited kinetically distinct polarization and docking phases and required calcium flux and signaling through both the T cell receptor and integrin to be robust. In “frustrated” conjugates where the centrosome is stuck behind the nucleus, the center of the IS invaginated dramatically to approach the centrosome. Consistently, imaging of microtubules during normal repositioning revealed a microtubule end-on capture-shrinkage mechanism operating at the center of the IS. In agreement with this mechanism, centrosome repositioning was impaired by inhibiting microtubule depolymerization or dynein. We conclude that dynein drives centrosome repositioning in T cells via microtubule end-on capture-shrinkage operating at the center of the IS and not cortical sliding at the IS periphery, as previously thought. 相似文献
5.
Dynein-dependent motility of microtubules and nucleation sites supports polarization of the tubulin array in the fungus Ustilago maydis
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Microtubules (MTs) are often organized by a nucleus-associated MT organizing center (MTOC). In addition, in neurons and epithelial cells, motor-based transport of assembled MTs determines the polarity of the MT array. Here, we show that MT motility participates in MT organization in the fungus Ustilago maydis. In budding cells, most MTs are nucleated by three to six small and motile gamma-tubulin-containing MTOCs at the boundary of mother and daughter cell, which results in a polarized MT array. In addition, free MTs and MTOCs move rapidly throughout the cytoplasm. Disruption of MTs with benomyl and subsequent washout led to an equal distribution of the MTOC and random formation of highly motile and randomly oriented MTs throughout the cytoplasm. Within 3 min after washout, MTOCs returned to the neck region and the polarized MT array was reestablished. MT motility and polarity of the MT array was lost in dynein mutants, indicating that dynein-based transport of MTs and MTOCs polarizes the MT cytoskeleton. Observation of green fluorescent protein-tagged dynein indicated that this is achieved by off-loading dynein from the plus-ends of motile MTs. We propose that MT organization in U. maydis involves dynein-mediated motility of MTs and nucleation sites. 相似文献
6.
Microtubule detachment from the microtubule-organizing center as a key event in the complete turnover of microtubules in cells 总被引:6,自引:0,他引:6
Microtubule (MT) response to different steady state temperatures and to rapid shifts in temperature was studied quantitatively in large, thin cells (LT-cells) from the goldfish scale. MT number and total tubulin concentration per cell were found to be fairly constant in cells from the same fish, regardless of cell size but between fish, could differ by a factor of two. The total tubulin concentration was similar to that found in mammalian tissue culture cells and the proportion in MT form increased with increasing steady state temperature. Total MT length quickly and exponentially decreased when cells were rapidly chilled to approximately -3 degrees C. In contrast, the average length of the MTs bound to the MT organizing center (MTOC) did not significantly change. Free MTs were generated during chilling and had an average length roughly half that of bound MTs. These observations suggest that 1) there is a functional block to rapid depolymerization at the unattached end of the MTOC bound MTs and 2) depolymerization of the MT occurs from the originally bound end only after its release from the MTOC. The presence of free MTs in a wide variety of cells suggests that these two features may be characteristic of steady state MTs in other cells. When the temperature of the LT-cells was abruptly raised, the number of MTs initiated on the MTOC rapidly increased and reached a brief steady state long before the MTs completely elongated. Many MTs then apparently detached from the MTOC and depolymerized before a final steady state was reached. When cells containing newly polymerized MTs were chilled to approximately -3 degrees C, the MTs detached from the MTOC more rapidly than those starting from steady state. Furthermore, the block to depolymerization at the unattached end was not complete. These observations suggest that newly formed, non-steady state MTs are different from the older, steady state MTs. 相似文献
7.
The formin mDia regulates GSK3beta through novel PKCs to promote microtubule stabilization but not MTOC reorientation in migrating fibroblasts
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In migrating cells, external signals polarize the microtubule (MT) cytoskeleton by stimulating the formation of oriented, stabilized MTs and inducing the reorientation of the MT organizing center (MTOC). Glycogen synthase kinase 3beta (GSK3beta) has been implicated in each of these processes, although whether it regulates both processes in a single system and how its activity is regulated are unclear. We examined these issues in wound-edge, serum-starved NIH 3T3 fibroblasts where MT stabilization and MTOC reorientation are triggered by lysophosphatidic acid (LPA), but are regulated independently by distinct Rho GTPase-signaling pathways. In the absence of other treatments, the GSK3beta inhibitors, LiCl or SB216763, induced the formation of stable MTs, but not MTOC reorientation, in starved fibroblasts. Overexpression of GSK3beta in starved fibroblasts inhibited LPA-induced stable MTs without inhibiting MTOC reorientation. Analysis of factors involved in stable MT formation (Rho, mDia, and EB1) showed that GSK3beta functioned upstream of EB1, but downstream of Rho-mDia. mDia was both necessary and sufficient for inducing stable MTs and for up-regulating GSK3beta phosphorylation on Ser9, an inhibitory site. mDia appears to regulate GSK3beta through novel class PKCs because PKC inhibitors and dominant negative constructs of novel PKC isoforms prevented phosphorylation of GSK3beta Ser9 and stable MT formation. Novel PKCs also interacted with mDia in vivo and in vitro. These results identify a new activity for the formin mDia in regulating GSK3beta through novel PKCs and implicate novel PKCs as new factors in the MT stabilization pathway. 相似文献
8.
Palazzo AF Joseph HL Chen YJ Dujardin DL Alberts AS Pfister KK Vallee RB Gundersen GG 《Current biology : CB》2001,11(19):1536-1541
In migrating adherent cells such as fibroblasts and endothelial cells, the microtubule-organizing center (MTOC) reorients toward the leading edge [1-3]. MTOC reorientation repositions the Golgi toward the front of the cell [1] and contributes to directional migration [4]. The mechanism of MTOC reorientation and its relation to the formation of stabilized microtubules (MTs) in the leading edge, which occurs concomitantly with MTOC reorientation [3], is unknown. We show that serum and the serum lipid, lysophosphatidic acid (LPA), increased Cdc42 GTP levels and triggered MTOC reorientation in serum-starved wounded monolayers of 3T3 fibroblasts. Cdc42, but not Rho or Rac, was both sufficient and necessary for LPA-stimulated MTOC reorientation. MTOC reorientation was independent of Cdc42-induced changes in actin and was not blocked by cytochalasin D. Inhibition of dynein or dynactin blocked LPA- and Cdc42-stimulated MTOC reorientation. LPA also stimulates a Rho/mDia pathway that selectively stabilizes MTs in the leading edge [5, 6]; however, activators and inhibitors of MTOC reorientation and MT stabilization showed that each response was regulated independently. These results establish an LPA/Cdc42 signaling pathway that regulates MTOC reorientation in a dynein-dependent manner. MTOC reorientation and MT stabilization both act to polarize the MT array in migrating cells, yet these processes act independently and are regulated by separate Rho family GTPase-signaling pathways. 相似文献
9.
Summary A 49 kDa protein in tobacco BY-2 cells has been found to be cross-reactive with antibodies raised against a 51 kDa protein that was isolated from sea urchin centrosomes and identified as a microtubule-organizing center (MTOC) in animal cells. Tracing the fate of the 49 kDa protein during progression of the cell cycle in highly synchronized tobacco BY-2 cells revealed that this protein was colocalized with plant microtubules (MTs): the location of the 49 kDa protein coincided with preprophase bands (PPBs), mitotic spindles and phragmoplasts. Furthermore, between the M and G1 phases, the 49 kDa protein was observed in the perinuclear regions, in which the initials of MTs are organizing to form cortical MTs. At the G1 phase the location of the 49 kDa protein in the cell cortex coincided with that of the cortical MTs. It appeared that the 49 kDa protein in the cell cortex was transported as granules from the perinuclear regions. Thus, it is highly probable that the 49 kDa protein, which reacts with antibodies against the 51 kDa protein in sea urchin centrosomes, plays the role of an MTOC in plant cells. Thus, the mechanisms for organizing MTs in higher organisms appear to share a common protein, even though the organization of MTs is superficially very different in plant and animal cells.Abbreviations DAPI
4,6-diamidino-2-phenyl indole
- MT
microtubule
- MTOC
microtubule-organizing center
- PAGE
polyacrylamide gel electrophoresis
- PBS
phosphate-buffered saline
- PPB
preprophase band
- SDS
sodium dodecylsulfate 相似文献
10.
Reorientation of the microtubule-organizing center and the Golgi apparatus in cloned cytotoxic lymphocytes triggered by binding to lysable target cells 总被引:13,自引:0,他引:13
By immunofluorescence observations with cell couples of cloned murine cytotoxic T lymphocytes (CTL) and target cells, evidence is presented for a rapid reorientation of the microtubule-organizing center (MTOC) and the Golgi apparatus (GA) in the effector cell (but not in the target cell) toward the contact area with the target. The reorientation of the MTOC/GA and the cytotoxic activity of the CTL were inhibited reversibly by nocodazole, a microtubule-disrupting agent. In lectin-formed cell couples of CTL and neuraminidase-treated target cells, the MTOC in essentially all of the CTL was oriented toward the effector-target contact area of a lysable target cell, but was left randomly oriented with a nonlysable target cell. A similar random orientation of the effector-MTOC was also observed in cell couples of cloned natural killer cells and nonlysable targets. These findings indicate that the repositioning of the MTOC and the GA, which is shared by CTL and natural killer cells, is an essential and early event in the onset of the cytolytic mechanism. It is suggested that this reorientation serves the purpose of directing to the bound target cell secretory vesicles derived from the GA that contain cytotoxic substances. 相似文献
11.
Osteoclasts are highly specialized, multinucleated cells responsible for the selective resorption of the dense, calcified bone matrix. Microtubules (MTs) contribute to the polarization and trafficking events involved in bone resorption by osteoclasts; however, the origin of these elaborate arrays is less clear. Osteoclasts arise through cell fusion of precursor cells. Previous studies have suggested that centrosome MT nucleation is lost during this process, with the nuclear membrane and its surrounding Golgi serving as the major MT organizing centers (MTOCs) in these cells. Here we reveal that precursor cell centrosomes are maintained and functional in the multinucleated osteoclast and interestingly form large MTOC clusters, with the clusters organizing significantly more MTs compared with individual centrosomes. MTOC cluster formation requires dynamic MTs and minus-end directed MT motor activity. Inhibition of these centrosome clustering elements had a marked impact on both F-actin ring formation and bone resorption. Together these findings show that multinucleated osteoclasts employ unique centrosomal clusters to organize the extensive MTs during bone attachment and resorption. 相似文献
12.
Re-formation of microtubules in Closterium ehrenbergii Meneghini after cold-induced depolymerization
T. Hogetsu 《Planta》1986,167(4):437-443
Immunofluorescence microscopy was used to examine the re-formation of microtubules (MT), after cold-induced depolymerization, in Closterium ehrenbergii. The C. ehrenbergii cells undergo cell division followed by semicell expansion in the dark period of daily light-dark cycles. Five types of MTs, namely the MT ring, hair-like MTs around the nuclei, spindle MTs, radially arranged MTs and transverse wall MTs, appeared and disappeared sequentially during and following cell division. The wall MTs were distributed transversely only in the expanding new semicells. When cells were chilled in ice water, wall MTs in expanding cells were fragmented, and then disappeared as did the other types of MTs, within 5 min. When cells were warmed at 20°C after 2 h chilling, wall MTs and the other types of MTs re-formed. At the early stage of wall-MT re-formation in expanding cells, small, star-like MTs were formed, and then randomly oriented MTs developed in both the expanding new and the old semicells. The MT ring was also re-formed at the boundary between the new and old semicells. There were no obvious MT-organizing centers in the random arrangement. As time passed, the randomly oriented wall MTs in the old semicells disappeared and those in the expanding new semicells gradually assumed a transverse orientation. These results indicate that wall MTs can be rearranged transversely after they have been re-formed and that nucleation of wall MTs is separable from the mechanism for ordering them.Abbreviations MT(s)
microtubule(s)
- MTOC(s)
microtubule-organizing center(s) 相似文献
13.
The idea of microtubules (MTs) as of passive railway tracks, along which transport vesicles travel by use of motor proteins, is widely accepted. In the present work the organization of MT system during EGF-receptor endocytosis was investigated by indirect double immunofluorescence in HeLa and A431 cell lines. Stimulation of cells with EGF resulted in formation of EGF receptor-containing peripheral vesicular endosomes. During time course of endocytosis the endosomes tended to concentrate in juxtranuclear region close to MTOC. This translocation was dependent on MTs since nocodazole treatment resulted in endosomes' scattering throughout the cytoplasm. Parallel staining of the cells with tubulin antibody has revealed significant remodeling of MTs organization during endocytosis. At early stages MTs demonstrated slight retraction at the cell periphery and the increasing intensity of tubulin fluorescence in the juxtranuclear region. Later on, long individual MTs disappeared and peripheral cytoplasm show diffuse staining in combination with a meshwork of short MT fragments. This stage correlated with EGFR localization in juxtranuclear endosomes. Disappearance of EGFR-positive staining due to its lysosomal degradation occurred in parallel to reestablishment of radial MT system. Possible functional significance of described alterations in organization of tubulin cytoskeleton is discussed. 相似文献
14.
Vinogradova T Paul R Grimaldi AD Loncarek J Miller PM Yampolsky D Magidson V Khodjakov A Mogilner A Kaverina I 《Molecular biology of the cell》2012,23(5):820-833
Assembly of an integral Golgi complex is driven by microtubule (MT)-dependent transport. Conversely, the Golgi itself functions as an unconventional MT-organizing center (MTOC). This raises the question of whether Golgi assembly requires centrosomal MTs or can be self-organized, relying on its own MTOC activity. The computational model presented here predicts that each MT population is capable of gathering Golgi stacks but not of establishing Golgi complex integrity or polarity. In contrast, the concerted effort of two MT populations would assemble an integral, polarized Golgi complex. Indeed, while laser ablation of the centrosome did not alter already-formed Golgi complexes, acentrosomal cells fail to reassemble an integral complex upon nocodazole washout. Moreover, polarity of post-Golgi trafficking was compromised under these conditions, leading to strong deficiency in polarized cell migration. Our data indicate that centrosomal MTs complement Golgi self-organization for proper Golgi assembly and motile-cell polarization. 相似文献
15.
In this paper, we construct a novel nonlocal transport model that describes the evolution of microtubules (MTs) as they interact with stationary distributions of motor proteins. An advection term accounts for directed MT transport (sliding due to motor protein action), and an integral term accounts for reorientation of MTs due to their interactions with cross-linking motor proteins. Simulations of our model show how MT patterns depend on boundary constraints, as well as model parameters that represent motor speed, cross-linking capability (motor activity), and directionality. In large domains, and using motor parameter values consistent with experimentally-derived values, we find that patterns such as asters, vortices, and bundles are able to persist. In vivo, MTs take on aster patterns during interphase and they form bundles in neurons and polarized epithelial cells. Vortex patterns have not been observed in vivo, however, are found in in vitro experiments. In constrained domains, we find that similar patterns form (asters, bundles, and vortices). However, we also find that when two opposing motors are present, anti-parallel bundles are able to form, resembling the mitotic spindle during cell division. This model demonstrates how MT sliding and MT reorientation are sufficient to produce experimentally observed patterns. 相似文献
16.
M Hauser 《European journal of cell biology》1986,42(2):295-304
Short and long-term effects of the antitumor drug taxol on the microtubular axonemes and on the microtubule-organizing centroplast of the centrolhelidian Heterophrys marina have been investigated. Short-term treatment reveals that general aspects of cell structure remain substantially unaffected and that additional microtubule (MT) assembly in individual axonemes is accompanied by disassembly in others. However, the interaction between MTs, via cross-bridging associated proteins, is seriously affected as indicated by the numerous softly-bent axopods with disturbed arrays of the normally hexagonal pattern. Stabilization of MTs becomes evident by the reexpansion of axopods during low temperature incubation and also by the rapid inhibition of the saltatory movement of the extrusive organelles. Rapidly reexpanding axonemes of cells incubated at higher temperatures and in high taxol concentrations arise asymmetrically from the microtubule-organizing centrosomal structure (centroplast) and form a single thick and thorn-like axopodium, indicating a certain disarrangement of the centrally located microtubule-organizing center (MTOC), which obviously is severely damaged after long-term treatment. With increasing disorganization of the centroplast's structure, these cells reveal themselves unable to sustain their regular microtubular axonemal cytoskeleton. Paradoxically, polymerization of free microtubules from the tubulin pool does not take place. Instead, paracrystalline arrays of twisted filaments appear within the cytoplasm. It is concluded that heliozoan MTs can only persist if stabilized by additional factors, such as permanent interaction with the intact centroplast, and that even in the presence of taxol, MTs unattached to such an MTOC will be intrinsically unstable. 相似文献
17.
Summary Examination was made of the structure and assembly of the cortical microtubule (MT) cytoskeleton in the coenocytic green algaBoodlea coacta (Dickie) Murray et De Toni by immunofluorescence microscopy. Cortical MTs inBoodlea protoplasts are arranged randomly but some show a meridional arrangement within 6 h after protoplast formation. At 6–9 h such MTs become highly concentrated and parallel to each other in certain areas. At 12 h the concentration is uniformly high throughout the cell, indicating the completion of high density meridional arrangement of cortical MTs. Cortical MTs exhibiting a high density, meridional arrangement show characteristic disassembly by treatment with 10 M amiprophos-methyl (APM) or cold treatment (0 °C). Disassembly occurs by each MT unit at positions skipping 30–40 m in the transverse direction, and neighboring MTs subsequently disassemble to form MT groups. Each group becomes slender and then disappears completely within the following 24 h. The meridional arrangement of cortical MTs is disrupted by N-ethylmaleimide (NEM) accompanied by a remarkable reduction in density. The remaining MTs form groups at 30–40 m intervals from each other, as also occurs with drug or cold treatment, but disruption and density return to normal levels following removal of NEM. It appears that there are meridionally oriented channels, anchor-rich and anchor-poor, in the plasma membrane. The channels could be distributed alternately and anchors could be deposited in a cross-linking manner with cortical MTs to form a stable cortical MT-cytoskeleton. MTs comprising the cortical MT cytoskeleton could be oriented by meridionally oriented channels of anchors which are distributed following establishment of cell polarity.Abbreviations APM
amiprophos-methyl
- MT
microtubule
- MTOC
microtubule organizing center
- NEM
N-ethylrnaleimide 相似文献
18.
Efficient intracellular transport of the capsid of alphaherpesviruses, such as herpes simplex virus 1 (HSV-1), is known to be dependent upon the microtubule (MT) network. Typically, the MT network radiates from an MT-organizing center (MTOC), which is, in most cases, the centrosome. During herpesvirus egress, it has been assumed that capsids travel first from the nucleus to the centrosome and then from the centrosome to the site of envelopment. Here we report that the centrosome is no longer a primary MTOC in HSV-1-infected cells, but it retains this function in cells infected by another alphaherpesvirus, pseudorabies virus (PrV). As a result, MTs formed at late times after infection with PrV grow from a major, centralized MTOC, while those formed after HSV-1 infection arise from dispersed locations in the cytoplasm, indicating the presence of alternative and minor MTOCs. Thus, loss of the principal MT nucleating center in cells following HSV-1 infection raises questions about the mechanism of HSV-1 capsid egress. It is possible that, rather than passing via the centrosome, capsids may travel directly to the site of envelopment after exiting the nucleus. We suggest that, in HSV-1-infected cells, the disruption of centrosomal functions triggers reorganization of the MT network to favor noncentrosomal MTs and promote efficient viral spread. 相似文献
19.
Martín-Cófreces NB Sancho D Fernández E Vicente-Manzanares M Gordón-Alonso M Montoya MC Michel F Acuto O Alarcón B Sánchez-Madrid F 《Journal of immunology (Baltimore, Md. : 1950)》2006,176(7):4201-4207
The translocation of the microtubule-organizing center (MTOC), its associated signaling complex, and the secretory apparatus is the most characteristic early event that involves the tubulin cytoskeleton of T or NK cells after their interaction with APC or target cells. Our results show that Fyn kinase activity is essential for MTOC reorientation in an Ag-dependent system. Moreover, T cells from Fyn-deficient mice are unable to rearrange their tubulin cytoskeleton in response to anti-CD3-coated beads. Analysis of conjugates of T cells from transgenic OT-I mice with dendritic cells revealed that an antagonist peptide induces translocation of the MTOC, and that this process is impaired in T cells from Fyn(-/-) OT-I mice. In addition, Fyn deficiency significantly affects the MTOC relocation mediated by agonist peptide stimulation. These results reveal Fyn to be a key regulator of tubulin cytoskeleton reorganization in T cells. 相似文献
20.
Donghua Liu Ping Xue Qingmin Meng Jing Zou Jiegang Gu Wusheng Jiang 《Plant cell reports》2009,28(4):695-702
The effects of lead and copper on the arrangement of microtubule (MT) cytoskeleton in root tip cells of Allium sativum L. were investigated. Batch cultures of garlic were carried out under defined conditions in the presence 10−4 M Pb/Cu of various duration treatments. With tubulin immunolabelling and transmission electron microscopy (TEM), we found
four different types of MT structures depending on the cell cycle stage: the interphase array, preprophase band, mitotic spindle
and phragmoplast were typical for the control cells. Pb/Cu affected the mechanisms controlling the organization of MT cytoskeleton,
and induces the following aberrations in interphase and mitotic cells. (1) Pb/Cu induced the formation of atypical MT arrays
in the cortical cytoplasm of the interphase cells, consisting of skewed, wavy MT bundles, MT fragments and ring-like tubulin
aggregations. (2) Pb/Cu disordered the chromosome movements carried out by the mitotic spindle. The outcome was chromosome
aberrations, for example, chromosome bridges and chromosome stickiness, as well as inhibition of cells from entering mitosis.
(3) Depending on the time of exposure, MTs disintegrated into shorter fragments or they completely disappeared, indicating
MT depolymerization. (4) Different metals had different effects on MT organization. MTs were more sensitive to the pressure
of Cu ions than Pb. Moreover, TEM observations showed that the MTs were relatively short and in some places wavy when exposed
to 10−4 M Pb/Cu solutions for 1–2 h. In many sections MTs were no longer visible with increasing duration of treatment (>4 h). Based
on these results, we suggested that MT cytoskeleton is primarily responsible for Pb/Cu-associated toxicity and tolerance in
plants. 相似文献