Chaotization of division spindle, phragmoplast, and telophase chromosome groups in wheat x wheatgrass F1 hybrids meiosis

The paper describes the phenomenon of disorganization of completely formed subcellular structures: division spindle, phragmoplast and chromosome telophase groups. These structures disintegrate into their elements (cytoskeletal fibers, chromosomes) that transform into chaotic system. Chaotization of cytoskeleton structures such as prophase spindle in mitosis or perinuclear ring in meiosis is a normal step of wild type plant cell division. Disintegration of division spindle and phragmoplast presumably indicate the abnormality of temporal regulation of cytoskeleton cycle during meiosis. Disintegration of telophase chromosome groups and the migration of the chromosomes backward to the equatorial area might mean the abnormal start of some prometaphase mechanisms, in particular, chromokinesins activation.     

A new type of microtubular cytoskeleton in microsporogenesis of Lavatera thuringiaca L.

Summary. In Lavatera thuringiaca, kariokinesis and simultaneous cytokinesis during the meiotic division of microsporogenesis follow a procedure similar to that which takes place in the majority of members of the class Angiospermae. However, chondriokinesis occurs in a unique way found only in species from the family Malvaceae. Chondriokinesis in such species is well documented, but the relationship between the tubulin cytoskeleton and rearrangement of cell organelles during meiosis in L. thuringiaca has not been precisely defined so far. In this study, the microtubular cytoskeleton was investigated in dividing microsporocytes of L. thuringiaca by immunofluorescence. The meiotic stages and positions of cell organelles were identified by staining with 4′,6-diamidino-2-phenylindole. We observed that, during prophase I and II, changes in microtubular cytoskeleton configurations have unique features, which have not been described for other plant species. At the end of prophase I, organelles (mostly plastids and mitochondria) form a compact envelope around the nucleus, and the subsequent phases of kariokinesis take place within this arrangement. At this point of cell division, microtubules surround the organelle envelope and separate it from the peripheral cytoplasm, which is devoid of plastids and mitochondria. In telophase I, two newly formed nuclei are tightly surrounded by the cell organelle envelopes, and these are separated by the phragmoplast. Later, when the phragmoplast disappears, cell organelles still surround the nuclei but also move a little, starting to occupy the place of the disappearing phragmoplast. After the breakup of tetrads, the radial microtubule system is well developed, and cell organelles can still be observed as a dense envelope around the nuclei. At a very late stage of sporoderm development, the radial microtubule system disappears, and cell organelles become gradually scattered in the cytoplasm of the microspores. Using colchicines, specific inhibitors of microtubule formation, we investigated the relationship between the tubulin cytoskeleton and the distribution of cell organelles. Our analysis demonstrates that impairment of microtubule organization, which constitutes only a single component of the cytoskeleton, is enough to disturb typical chondriokinesis in L. thuringiaca. This indicates that microtubules (independent of microfilaments) are responsible for the reorganization of cell organelles during meiotic division. Correspondence: D. Tchórzewska, Department of Plant Anatomy and Cytology, Maria Curie-Skłodowska University, Akademicka 19, 20-033 Lublin, Poland.     

Dynamics of microtubular cytoskeleton during meiosis in higher plants. VIII. Comparison between successive and simultaneous cytokinesis

The abnormal cytoskeleton cycle in meiosis in pollen mother cells of cereal wide hybrids F reveals the role of polar microtubules in phragmoplast formation during successive cytokinesis. The cytoskeletal rearrangements during successive and simultaneous cytokinesis in higher plant meiosis are compared.     

Dynamics of microtubular cytoskeleton in higher plant meiosis. VI. Mechanisms of the successive cytokinesis

Intracellular morphological processes of successive cytokinesis in cereal pollen mother cells during normal and abnormal meiosis were studied. It was shown that the central spindle fiber system transforms into a phragmoplast at telophase. A model of centrifugal movement of the phragmoplast as a modification of B-anaphase has been proposed.     

The dynamics of the actin cytoskeleton during sporogenesis in Psilotum nudum L.

The actin cytoskeleton (microfilaments, MFs) accompanies the tubulin cytoskeleton (microtubules) during the meiotic division of the cell, but knowledge about the scope of their physiological competence and cooperation is insufficient. To cast more light on this issue, we analysed the F-actin distribution during the meiotic division of the Psilotum nudum sporocytes. Unfixed sporangia of P. nudum were stained with rhodamine-phalloidin and 4′,6-diamidino-2-phenylindole dihydrochloride, and we monitored the changes in the actin cytoskeleton and nuclear chromatin throughout sporogenesis. We observed that the actin cytoskeleton in meiotically dividing cells is not only part of the kariokinetic spindle and phragmoplast but it also forms a well-developed network in the cytoplasm present in all phases of meiosis. Moreover, in telophase I F-actin filaments formed short-lived phragmoplast, which was adjacent to the plasma membrane, exactly at the site of future cell wall formation. Additionally, the meiocytes were pre-treated with cytochalasin-B at a concentration that causes damage to the MFs. This facilitated observation of the effect of selective MFs damage on the course of meiosis and sporogenesis of P. nudum. Changes were observed that occurred in the cytochalasin-treated cells: the daughter nuclei were located abnormally close to each other, there was no formation of the equatorial plate of organelles and, consequently, meiosis did not occur normally. It seems possible that, if the actin cytoskeleton only is damaged, regular cytokinesis will not occur and, hence, no viable spores will be produced.     

Dynamics of microtubular cytoskeleton in higher plant meiosis. IX. The cycle accomplishment. Transition from phragmoplast to radial microtubule system

In this study we analysed the terminal step of cytoskeleton cycle in higher plant meiosis: transition from phragmoplast to radial interphase configuration. Wild type meiosis in a range of mono- and dicotyledonous species was studied. A number of cytoskeleton abnormalities on this stage was described in meiotic mutants, haploids and wide hybrids of various species. We described processes of cytoskeleton rearrangements on this stage: disjunction of phragmoplast MTs, their shortening and the role of daughter cell membranes. The independence of the interphase radial MT system formation from the previous steps of cytoskeleton cycle and from nuclear envelope cycle is proposed.     

洋葱小孢子母细胞减数分裂过程中微管分布变化

应用间接免疫荧光标记技术和激光共聚焦扫描显微镜成像技术观察洋葱小孢子母细胞减数分裂过程中微管分布变化。减数分裂之前,小孢子母细胞中的微管较短,呈辐射状,由细胞核表面向四周扩散。减数分裂开始后,细胞质中的一部分微管蛋白聚集成纺锤体微管,控制染色体的分布。进入减数分裂I后期,纺锤体微管变为牵引染色体移向两极的着丝粒微管和连接纺锤体两极的极丝微管。之后,所有微管集中在两个核之间,构成成膜体。然后,微管解聚成微管蛋白弥散在细胞质中。减数分裂I完成后,二分体2个子细胞中的微管蛋白又聚集成2个纺锤体微管,开始减数分裂II过程。经过减数分裂II中期,2个二分体细胞中的微管再次集中在2个细胞核之间形成成膜体,隔离2个细胞核。此后,微管蛋白解聚,弥散分布在小孢子细胞质中。     

Accessory phragmoplast corrects abnormal cytokinesis in wheat x rye hybrids

The compensation for phragmoplast dysfunction in the male meiosis of F1 wheat × rye hybrids was described. In pollen mother cells (PMCs), he transition from central spindle fibers (forming a solid bundle) to phragmoplast (hollow cylinder) was blocked. This blockage suppresses the centrifugal movement of the phragmoplast and cell-plate formation. As a result, cells become binucleate. Sometimes, two nuclei fuse and form one restitution nucleus. In PMCs of the wheat × rye F1 hybrid D-144 gp 06 year (T. aestivum n. 93-60 t 9 × S. cereale n. Saratovskaya 7) with this phenotype, an additional phragmoplast is formed at the late telophase. This occurs by a common mechanism for the development of the immobile phragmoplast in the meiosis in bicotyledons; new phragmoplasts arise as a result of microtubule polymerization starting from the spindle poles. The accessory phragmoplast facilitates a new cell plate assembly and achievement of cytokinesis.     

Chaotization of division spindle,phragmoplast, and telophase chromosome groups in wheat wheatgrass F1 hybrids meiosis

The phenomenon of the disorientation of completely formed systemic cytoskeleton structures, i.e., the division spindle and phragmoplast, into the constituent elements and their transformation into a network of disoriented fibers in the course of cell division is described. The phenomenon of the disintegration and dispersion in the cytoplasm of completely formed telophase chromosome groups, which is not associated with the chaotization of the cytoskeleton structures, is also described. These abnormalities are revealed in the meiosis of pollen mother cells of the first generation of wheat-wheatgrass hybrids. The chaotization of cytoskeleton structures is a only normal phenomenon in plant-cell division in late prophase-early prometaphase, whereas, at stages of metaphase and telophase, it can indicate a disturbance in the time regulation of the cytoskeleton cycle in the course of meiotic division. The disintegration of the chromosome telophase groups and their movement backwards to the spindle equator can indicate the untimely involvement of processes of prometaphase, specifically the activation of chromokinesins. The significance of the process of cytoskeleton chaotization in the biology of a plant cell is discussed.     

Formation and function of phragmoplast during successive cytokinesis stages in higher plant meiosis

Cytoskeletal rearrangements were studied during meiotic telophase in a number of monocotyledonous plant species. Wild type and abnormal meiosis (in wide cereal hybrids, meiotic mutants and allolines) was analyzed. It was found that central spindle fibers that move centrifugally, along with newly-formed MTs, are the basis of phragmoplast formation and function in PMCs of monocotyledonous plant species with successive cytokinesis stages. A model for centrifugal movement of the meiotic phragmoplast is proposed; this model is a modification of the corresponding process during B-anaphase.     

Microfilament Distribution in Maize Meiotic Mutants Correlates with Microtubule Organization

Microtubules and microfilaments often codistribute in plants; their presumed interaction can be tested with drugs although it is not always clear that these are without side effects. In this study, we exploited mutants defective in meiotic cell division to investigate in a noninvasive way the relationship between the two cytoskeletal elements. By staining unfixed, permeabilized cells with rhodamine-phalloidin, spatial and temporal changes in microfilament distribution during maize meiosis were examined. In wild-type microsporocytes, a microtubule array that radiates from the nucleus disappeared during spindle formation and returned at late telophase. This result differed from the complex cytoplasmic microfilament array that is present at all stages, including karyokinesis and cytokinesis. During division, a second class of microfilaments also was observed in the spindle and phragmoplast. To analyze this apparent association of microtubules and microfilaments, we examined several meiotic mutants known to have stage-specific disruptions in their microtubule arrays. Two mutations that altered the number or form of meiotic spindles also led to a dramatic reorganization of F-actin. In contrast, rearrangement of nonspindle, cytoplasmic microtubules did not lead to concomitant changes in F-actin distribution. These results suggested that microtubules and microfilaments interact in a cell cycle-specific and site-specific fashion during higher plant meiosis.     

Identification of a phragmoplast-associated kinesin-related protein in higher plants

The phragmoplast executes cytokinesis in higher plants. The major components of the phragmoplast are microtubules, which are arranged in two mirror-image arrays perpendicular to the division plane [1]. The plus ends of these microtubules are located near the site of the future cell plate. Golgi-derived vesicles are transported along microtubules towards the plus ends to deliver materials bound for the cell plate [2] [3]. During cell division, rapid microtubule reorganization in the phragmoplast requires the orchestrated activities of microtubule motor proteins such as kinesins. We isolated an Arabidopsis cDNA clone of a gene encoding an amino-terminal motor kinesin, AtPAKRP1, and have determined the partial sequence of its rice homolog. Immunofluorescence experiments with two sets of specific antibodies revealed consistent localization of AtPAKRP1 and its homolog in Arabidopsis and rice cells undergoing anaphase, telophase and cytokinesis. AtPAKRP1 started to accumulate along microtubules towards the spindle midzone during late anaphase. Once the phragmoplast microtubule array was established, AtPAKRP1 conspicuously localized to microtubules near the future cell plate. Our results provide evidence that AtPAKRP1 is a hitherto unknown motor that may take part in the establishment and/or maintenance of the phragmoplast microtubule array.     

Altering of phragmoplast orientation as a result of excessive centrifugal movement in the absence of cell plate

The inability of phragmoplast to stop its centrifugal movement after reaching the mother cell membrane is described in abnormal meiosis with the arrest of cell plate formation. The excess of phragmoplast expansion leads to rotation of the whole telophase figure (phragmoplast with daughter nuclei) within the cell through 90 degrees. It has been suggested that this phenomenon may occur because of a the lack of signal stopping cytokinesis. Such a signal arises due to formation of daughter cell membranes.     

Aluminium Effects on Microtubule Organization in Dividing Root-Tip Cells of Triticum turgidum. II. Cytokinetic Cells

Triticum turgidum were examined, using tubulin immunolabeling and electron microscopy. In cells, which at the beginning of the treatment were at a transitional stage between anaphase and telophase, the transformation of the interzonal microtubule (Mt) system into a phagmoplast was delayed. In cells treated at a telophase/ cytokinetic stage, the lateral phragmoplast expansion towards the cell periphery was delayed or inhibited. Besides, in cells entering telophase through an abnormal mitosis, Al inhibited phragmoplast formation and induced the organization of atypical tubulin bundles. The latter formed a network around the reassembling polyploid nucleus. The Al-effects resulted in the disturbance of cytokinesis and the formation of binuclear or polyploid cells, which lacked typical Mts. Instead of them, the post-telophase cells displayed atypical tubulin aggregations. In addition, Al affects cell plate development. Dividing cells, encompassing early interphase daughter nuclei, contained incomplete, atypical cell plates. The latter were quite thick, wavy and perforated, showing large “islands”, which contained electron transparent material. In some cells, the atypical cell plates gave rise to incomplete daughter walls, but in some others they were dismantled. The aberrant cell plates as well as the young daughter cell walls fluoresced intensely after aniline blue staining, an observation suggesting that they contain significant quantities of callose. The above findings combined with those derived from the study of the Al-effects on the mitotic spindle show that Mt cytoskeleton is a target site of Al toxicity in dividing cells. Received 24 October 2000/ Accepted in revised form 19 January 2001     

Heat stress affects the organization of microtubules and cell division in Nicotiana tabacum cells

To investigate the effects of heat stress on the plant cytoskeleton, the structure of microtubule arrays in N. tabacum suspension cells incubated at 38 or 42°C was analysed. Whilst incubation at 42 °C resulted in the disruption of the majority of cellular microtubules after 30 min, in cells exposed to 38 °C all the microtubule arrays were preserved even after 12 h of incubation, although their organization was altered. The most susceptible were the microtubules of the mitotic spindle and the phragmoplast. Several abnormalities were observed: (i) splitting of the spindle into several parts; (ii) elongation of the spindles; (iii) formation of microtubule asters in mitotic cells, and (iv) elongation of phragmoplast microtubules. Exposure of cells to 38 °C caused a decrease in the mitotic index but an accumulation of telophase cells. The recovery of normal microtubule organization occurred after 12 h. Treatment of the cells subjected to heat stress conditions with an inhibitor of protein synthesis, cycloheximide, did not prevent either the alterations of microtubule organization or accumulation of cells containing phragmoplasts. Therefore, heat shock proteins do not seem to be directly responsible for the microtubule disorganization induced by heat stress.     

{gamma}-Tubulin and microtubule organization during meiosis in the liverwort Ricciocarpus natans (Ricciaceae)

Extant liverworts are "living fossils" considered sister to all other plants and as such provide clues to the evolution of the microtubule organizing center (MTOC) in anastral cells. This report is the first on microtubule arrays and their γ-tubulin-nucleating sites during meiosis in a member of the Ricciales, a specialized, species-rich group of complex thalloid (marchantioid) liverworts. In meiotic prophase, γ-tubulin becomes concentrated at several sites adjacent to the nuclear envelope. Microtubules organized at these foci give rise to a multipolar prometaphase spindle. By metaphase I, the spindle has matured into a bipolar structure with truncated poles. In both first and second meiosis, γ-tubulin forms box-like caps at the spindle poles. γ-Tubulin moves from spindle poles to the proximal surfaces of telophase chromosomes where interzonal microtubules are nucleated. Although a phragmoplast is organized, no cell plate is deposited, and second division occurs simultaneously in the undivided sporocyte. γ-Tubulin surrounds each of the tetrad nuclei, and phragmoplasts initiated between both sister and nonsister nuclei direct simultaneous cytokinesis. The overall pattern of meiosis (unlobed polyplastidic sporocytes, nuclear envelope MTOC, multipolar spindle origin, spindles with box-like poles, and simultaneous cytokinesis) more closely resembles that of Conocephalum than other marchantiod liverworts.     

Nuclear cytoplasmic domains,microtubules and organelles in microsporocytes of the slipper orchid Cypripedium californicum A. Gray dividing by simultaneous cytokinesis

Microsporocytes of the slipper orchidCypripedium californicum A. Gray divide simultaneously after second meiosis. The organization and apportionment of the cytoplasm throughout meiosis are functions of nuclear-based radial microtubule systems (RMSs) that define domains of cytoplasm - a single sporocyte domain before meiosis, dyad domains within the undivided cytoplasm after first meiosis, and four spore domains after second meiosis. Organelles migrate to the interface of dyad domains in the undivided cytoplasm after first meiotic division, and second meiotic division takes place simultaneously on both sides of the equatorial organelle band. Microtubules emanating from the telophase II nuclei interact to form columnar arrrays that interconnect all four nuclei, non-sister as well as sister. Cell plates are initiated in these columns of microtubules and expand centrifugally along the interface of opposing RMSs, coalescing in the center of the sporocyte and joining with the original sporocyte wall at the periphery to form the tetrad of microspores. Organelles are distributed into the spore domains in conjunction with RMSs. These data, demonstrating that cytokinesis in microsporogenesis can occur in the absence of both components of the typical cytokinetic apparatus (the preprophase band of microtubules which predicts the division site and the phragmoplast which controls cell-plate deposition), suggest that plant nuclei have an inherent ability to establish a domain of cytoplasm via radial microtubule systems and to regulate wall deposition independently of the more complex cytokinetic apparatus of vegetative cells.     

Microtubule organization during successive microsporogenesis in Allium cepa and simultaneous cytokinesis in Nicotiana tabacum

Microtubule cytoskeleton organization during microspore mother cell (MMC) meiosis in Allium cepa L. and microsporogenesis in Nicotiana tabacum L. was examined. The MMC microtubules (MTs) were short and well dispersed in the cytoplasm of both taxa. As the MMCs of both species entered metaphase of meiosis I, the MTs constructed a spindle that facilitated the chromosomes to orient in the meridian plane. At anaphase of meiosis I, the spindle MTs differentiated into two types: one MT type became short, pulled the chromosomes toward the two poles, and was designated as centromere MTs; the second type of MT connected the two poles, and was designated as pole MTs. In A. cepa, where successive cytokinesis was observed, pole MTs assumed a tubbish shape. Some new short MTs aggregated in the meridian plane and constricted to form a phragmoplast, which developed into a cell plate, divided the cytoplasm into two parts and produced a dyad. However, in tobacco, a phragmoplast was not generated in anaphase of meiosis I and II and cytokinesis did not occur. The spindle MTs depolymerized and reorganized the radial arrangement of MTs from the nucleate surface to the periplasm during anaphase. Following telophase of meiosis II, the cytoplasm produced centripetal furrows, which met in the center of the cell and divided it into four parts, serving as a form of cytokinesis. In this process, MTs appeared to bear no relationship to cytokinesis.     

The kinesin-like calmodulin binding protein is differentially involved in cell division

The kinesin-like calmodulin (CaM) binding protein (KCBP), a minus end-directed microtubule motor protein unique to plants, has been implicated in cell division. KCBP is negatively regulated by Ca(2)+ and CaM, and antibodies raised against the CaM binding region inhibit CaM binding to KCBP in vitro; therefore, these antibodies can be used to activate KCBP constitutively. Injection of these antibodies into Tradescantia virginiana stamen hair cells during late prophase induces breakdown of the nuclear envelope within 2 to 10 min and leads the cell into prometaphase. However, mitosis is arrested, and the cell does not progress into anaphase. Injection of antibodies later during cell division has no effect on anaphase transition but causes aberrant phragmoplast formation and delays the completion of cytokinesis by approximately 15 min. These effects are achieved without any apparent degradation of the microtubule cytoskeleton. We propose that during nuclear envelope breakdown and anaphase, activated KCBP promotes the formation of a converging bipolar spindle by sliding and bundling microtubules. During metaphase and telophase, we suggest that its activity is downregulated.