Preprophase Band Of Microtubules Research Articles

Ultrastructure of cell division in the fusiform cells of the vascular cambium of Robinia pseudoacacia

The ultrastructure of periclinally dividing fusiform cells was studied in the vascular cambium of Robinia pseudoacacia. Fusiform cell division begins in April at Madison, Wisconsin, when the cambial cells still have many characteristics of a dormant cambium. Soon afterward, the cambial cells acquire the appearance typical of an active cambium. Sequential phases of the microtubule cycle were documented: cortical microtubules bordering the cell wall during interphase, perinuclear microtubules preceding formation of the mitotic spindle, spindle microtubules, and phragmoplast microtubules. A preprophase band of microtubules was not encountered. An extended phragmosome was not encountered in periclinally dividing fusiform cells. During cytokinesis, the phragmosome is represented by a broad cytoplasmic plate which precedes the developing phragmoplast and cell plate as they migrate toward the ends of the cell.

Nuclear and microtubular cycles in heterophasic multinuclearTriticum root-tip cells induced by caffeine

Nuclear and microtubular cycles were studied in large heterophasic multinuclear cells induced in root tips ofTriticum turgidum by caffeine treatment. Multinuclear cells and cells with polyploid nuclei exhibited various configurations of multiple and complex preprophase microtubule (Mt) bands (PPBs), including helical ones. The developmental stages of PPBs in some heterophasic cells did not comply with the cell cycle stages of the associated nuclei, a fact indicating that these events are not directly controlled by the associated nuclei. The heterophasic cells exhibited asynchronous nuclei at different stages of mitosis. In cells displaying prophase and interphase nuclei, the prophase spindle was either absent or developed around both of them or developed around the prophase nuclei earlier than around the interphase ones. During prometaphase-metaphase of the advanced nuclei the lagging interphase nuclei were induced to form prematurely condensed chromosomes (PCCs) along with spindle formation around them. These observations suggest that the mitotic transition in heterophasic cells is delayed but is ultimately achieved due to the effect of the advanced nuclei, which induces a premature mitotic entry of the lagging nuclei. Although kinetochore Mt bundles were found associated with PCCs, their metaphase and anaphase spindles were abnormal resulting in abnormal or abortive anaphases. In some heterophasic cells, metaphase-anaphase transition did not take place simultaneously in different chromosome groups, signifying that the cells do not exit from the mitotic state after anaphase initiation of the advanced nuclei. Asynchronous pace of mitosis of different chromosome groups was also observed during anaphase and telophase. Implications of these observations in understanding plant cell cycle regulation are discussed.

Cell division of binuclear cells induced by caffeine: Spindle organization and determination of division plane

Synchronously dividing binuclear cells were induced in root tips ofTriticum turgidum by caffeine treatment. Spindle and other microtubular configurations of such cells were studied using tubulin immunofluorescence and electron microscopy. The binuclear cells developed one, two or three preprophase microtubule bands longitudinally, transversely or rarely in a cross configuration. During the mitotic entry binuclear cells formed prophase spindles separately around each nucleus. When the nuclei were located fairly apart, their spindle structures developed independently throughout all mitotic phases. But when the nuclei were located closely together their metaphase and anaphase spindles shared a common polar region. However, the two spindles in such cells retained their functional autonomy. They display structurally independent minipoles in the common polar region. After anaphase the neighbouring nonsister chromosome groups of nuclei divided by a common polar region come to lie close together and in telophase, become enclosed by a common nuclear envelope. During cytokinesis of binuclear cells cell plates were formed only between sister nuclei. These cell plates may develop normally or may curve or branch giving rise to aberrant daughter cell walls. The peculiar mode of spindle and spindle polar region organization of binuclear cells and determination of the division plane in them are discussed.

A comparative study on stainability of preprophase bands by the PSTAIR antibody

The detectability of preprophase bands (PPBs) by antibody to PSTAIR sequence, which is found in cyclin-dependent kinases and is perfectly conserved in the p34cdc2 kinase from all known sources, was compared among root tip cells of 12 plant species and cultivars. Although more than 80% of prophase cells in all species examined had PPBs of microtubules (MTs), the detectability of PPBs by anti-PSTAIR varied from 0% to 88% depend on species examined. The detectability of PPBs by the antibody to PSTAIR was as high as that by antibody to tubulin inAllium cepa, A. fistulosum andA. tuberosum. InTriticum andPisum, the detectability varied greatly among cultivars. Only few faint PPBs could be detectable inChrysanthemum, and no PPBs were seen inHibiscus using anti-PSTAIR. The PSTAIR antibody recognized single (Hordeum, Triticum Zea) or multiple (Allium, Hibiscus, Pisum) bands around 34 kDa on protein blots of root tip exracts. PPBs of anti-PSTAIR cross reactive molecules were detectable in one fourth of the prophase cells ofPisum (cv. Snack) by the conventional method. However, the detectability of PPBs inPisum increased to 80% when the method for the PSTAIR-fluorescence was modified, suggesting that the low detectability of PPBs by anti-PSTAIR may not be due to genuine differences between species or cultivars, but may be the result of variable staining.

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.

PLANT MITOSIS PROMOTING FACTOR DISASSEMBLES THE MICROTUBULE PREPROPHASE BAND AND ACCELERATES PROPHASE PROGRESSION IN TRADESCANTIA

The regulation of mitosis in higher plant cells has been investigated by microinjecting protein kinase from the metaphase-arresting (met1) mutant of Chlamydomonas. Biochemical characterization of this enzyme complex confirms the presence of a p34 cdc2/cyclin B-like kinase. The enzyme was injected into living stamen hair cells of Tradescantia virginianain which microtubules (MTs) were visualized using fluorescent analogue cytochemistry and confocal laser scanning microscopy. Microinjection of this p34 cdc2/cyclin B-like kinase caused rapid disassembly of the preprophase band of MTs but not of interphase-cortical, spindle or phragmoplast MTs. Effects of the enzyme on the cytomorphology of live prophase cells were also monitored using video microscopy. We found that injection of this enzyme accelerated chromatin condensation and nuclear envelope breakdown. This indicates the presence and function in plants of an enzyme that can initiate nuclear division similar to the maturation or mitosis promoting factor (MPF) of animal cells. These studies provide the first direct evidence that the mitotically-active form of plant MPF can drive disassembly of preprophase band MTs, chromosome condensation and initiation of mitosis in plant cells.

Developmental features of protophloem sieve elements in roots of wheat (Triticum aestivum L.)

Protophloem sieve elements (PSEs) in roots of wheat (Triticum aestivum L.) are arranged in single vertical files. The number of PSEs within the files increases by symmetrical divisions, which take place after the completion of asymmetrical (formative) divisions and before the initiation of differentiation. The divisions are preceded by well defined pre-prophase bands (PPB) of microtubules, which surround the nucleus in an equatorial position. In the cytoplasmic region between the nuclear surface and the PPB, perinuclear and endoplasmic microtubules were observed. The perinuclear microtubules are considered as part of the developing spindle, while the endoplasmic ones interlink the perinuclear microtubules with the PPB. Dividing cells do not show any signs of incipient differentiation. The first and most reliable indication of a commencing differentiation is provided by the sieve-element plastids that begin to accumulate dense crystalloid inclusions in the very young PSEs. In mature PSEs plastids contain two kinds of crystalloid inclusions, dense and thin, in a translucent stroma. Depending on the plastid-inclusions criterion it was shown that: (a) the PSEs of a given root do not initiate differentiation at exactly the same stage, (b) the developmental sequence extends to a span of 7–9 actively differentiating PSEs arranged in a single vertical file, and (c) each PSE needs about 16–21 h to pass through the whole developmental sequence. In the last two differentiating PSEs of a file, mitochondria were found to be enveloped by single cisternae of ER. The association is temporary as it is lost in the first PSEs with an autolysed lumen. During differentiation, Golgi bodies were abundant and active in producing vesicles involved in cell wall development. Golgi vesicles were also found among the microtubules of the PPB, but no local thickening was observed. Golgi bodies disorganize in the last stages of autolysis and disappear in mature sieve elements.

Root hairs: Specialized tubular cells extending root surfaces

Root hairs are tubular extensions of epidermal cells that have their origin either in any protoderm cell or in specialized protoderm cells called trichoblasts. These latter cells are the result of an asymmetric cytokinesis determined by the positioning of a pre-prophase band of microtubules. The smaller sibling cell is the trichoblast and specializes physiologically and structurally prior to root hair outgrowth. Several genes are involved in the initiation and outgrowth of root hairs. Elongation of root hairs is by tip growth, and, correlated with this, cytoplasmic organelles and cytoskeletal elements show a polarized distribution; the apical dome consists of numerous vesicles, many associated with cell wall synthesis. The relationship between cellulose microfibril deposition and the pattern of cortical microtubules has received considerable attention, as has the role of the cytoskeleton and calcium in controlling cytoplasmic streaming. Root hairs extend the absorbing surface of the root and therefore have been studied in terms both of physiological characteristics of the plasma membrane and uptake of water and of various ions in the soil solution. Many plant species develop soil sheaths (rhizosheaths) which protect the root surface from desiccation and harbour various microorganisms; root hairs are intimately involved in these sheaths. Various growth regulators have been studied in terms of their effect on the structure and function of root hairs. Root hairs play a significant role in the interaction between plants and nitrogen-fixing microorganisms (e.g.,Rhizobium, Frankia) and symbiotic mycorrhizal fungi.

The plant cell cycle: conserved and unique features in mitotic control.

Somatic plant cells can use a hormone checkpoint in late G2 phase. Here cytokinin stimulates removal of phosphotyrosine from p34cdc2 kinase and concurrently capacity for activation of the kinase by Cdc25 phosphatase declines while activity of the kinase increases and cells enter mitosis. Processes unique to plant mitosis are driven by the mitotically active kinase since the enzyme taken from plant cells in metaphase, when injected, can disassemble the preprophase band microtubules that form in G2 phase at the site of the future cross wall. This action is specific, since microtubules are not depolymerised when in interphase cytoplasmic array, or spindle, or phragmoplast. Plant metaphase kinase acts as MPF by accelerating chromosome condensation and nuclear envelope breakdown.

Pattern and process of wall formation in developing endosperm

AbstractEndosperm is emerging as a system for investigating the genetic control of wall placement and deposition in plant development. Development of endosperm progresses in distinct stages from a wall‐less syncytial stage to a cellular stage that is entirely typical of plant meristems where the division plane is predicted by a preprophase band of microtubules (PPB) and cytokinesis is completed by formation of a cell plate in association with a phragmoplast. Four developmentally different types of walls, each associated with a different microtubule system, are sequentially produced: (1) free growing walls deposited in the absence of mitosis and phragmoplasts; (2) walls guided by cytoplasmic phragmoplasts formed adventitiously in the absence of mitosis; (3) walls formed by interzonal phragmoplasts in a cell cycle that lacks PPBs; and (4) wall deposition driven by interzonal phragmoplasts in a cycle that includes PPBs. We are using methods of differential screening to isolate cDNA clones corresponding in temporal and spatial pattern to the types of wall development, and are studying mutants for clues to the genetic controls of wall development.

F-actin redistributions at the division site in livingTradescantia stomatal complexes as revealed by microinjection of rhodamine-phalloidin

Microinjection of rhodamine-phalloidin into living cells of isolatedTradescantia leaf epidermis and visualisation by confocal microscopy has extended previous results on the distribution of actin in mitotic cells of higher plants and revealed new aspects of actin arrays in stomatal cells and their initials. Divisions in the stomatal guard mother cells and unspecialised epidermal cells are symmetrical. Asymmetrical divisions occur in guard mother precursor cells and subsidiary mother cells. Each asymmetrical division is preceded by migration of the nucleus and the subsequent accumulation of thick bundles of anticlinally oriented actin filaments localised to the area of the anticlinal wall closest to the polarised nucleus. During prophase, in all cell types, a subset of cortical actin filaments coaligns to form a band, which, like the preprophase band of microtubules, accurately delineates the site of insertion of the future cell wall. Following the breakdown of the nuclear envelope, F-actin in these bands disassembles but persists elsewhere in the cell cortex. Thus, cortical F-actin marks the division site throughout mitosis, firstly as an appropriately positioned band and then by its localised depletion from the same region of the cell cortex. This sequence has been detected in all classes of division inTradescantia leaf epidermis, irrespective of whether the division is asymmetrical or symmetrical, or whether the cell is vacuolate or densely cytoplasmic. Taken together with earlier observations on stamen hair cells and root tip cells it may therefore be a general cytoskeletal feature of division in cells of higher plants.

Colchicine-induced Paracrystals in Root Cells of Wheat (Triticum aestivum L.)

Tubulin conformations other than microtubules in the meristematic cells of wheat roots grown in the presence of 2 mM colchicine solution were investigated by immunofluorescence and electron microscopy. In the affected cells microtubules disappeared and were replaced by tubulin fluorescent strands that occurred in the cortical cytoplasm. With increasing time of exposure to colchicine the tubulin strands became better organized and occurred also in the subcortical cytoplasm and finally they were restricted to the area around the nucleus. In prophase and preprophase cells thick strands occupied the cortical cytoplasmic zone where in normal cells a preprophase microtubule band (PMB) was expected to be assembled.In the colchicine-treated cells electron microscopy revealed an accumulation of paracrystalline aggregates, which initially occurred along the cell wall and later deeper in the cytoplasm, in the perinuclear regions and the cytoplasmic invaginations of the nucleus. In transverse planes the paracrystalline strands appear to consist of hexagonal subunits in a 'honeycomb' arrangement, while in longitudinal and oblique sections they exhibit variable images. Since their distribution coincides with that of the tubulin strands visualized by immunofluorescence, they are considered to be the same structure. Therefore, the paracrystals consist of, or at least contain, tubulin. They are most likely to be polymers of tubulin-colchicine complexes.

The effect of taxol onTriticum preprophase root cells: preprophase microtubule band organization seems to depend on new microtubule assembly

To examine whether preprophase microtubule band (PPB) organization occurs by rearrangement of pre-existing, or by assembly of new microtubules (Mts), we treated root cells ofTriticum turgidum with taxol, which stabilizes pre-existing Mts by slowing their depolymerization. With taxol early preprophase cells failed to form a normal PPB and PPB narrowing was prevented in cells that had already formed a wide one. The PPB became persistent in prometaphase cells and the formation of multipolar prophase-prometaphase spindles was induced. These data favour the suggestion that PPB formation and narrowing, as well as prophase spindle development, are dynamic processes depending on continuous Mt assembly at the PPB site and in the perinuclear cytoplasm.

A cytoskeletal 50 kDa protein in higher plants that forms intermediate-sized filaments and stabilizes microtubules

Filamentous structures of 7–10 nm in diameter were regenerated in vitro from a soluble 50 kDa protein (p50) that had been isolated from mung bean seedlings and from cultured tobacco cells. The polymerization of p50 in vitro was dependent on the presence of guanosine nucleotides, in particular, guanosine monophosphate (GMP). Unlike tubulin, p50 is a stable basic protein with the ability to polymerize even after heat treatment for 1 min at 70 °C. Furthermore, the freeze-dried powder of p50 retained the ability to regenerate filamentous structures after it had been dissolved in polymerization buffer to which GMP was then added. Two monoclonal antibodies against p50 were obtained. These antibodies stained the filamentous structures that extended from the surface of the nucleus to the cell periphery in interphase tobacco cells. They stained spindles and phragmoplasts as did tubulin-specific antibodies. They also stained fibrillar structures that were present around the spindle poles and the telophase daughter nuclei in which no microtubules were present. These results suggest that a part of the cell's complement of p50 may be associated with microtubules in dividing cells while the rest may itself form unique fibrillar structures. The antibodies against p50 did not stain cortical microtubules or the pre-prophase band of microtubules. The antibody against p50 also stained intermediate filament-like structures in cultured animal cells. The formation of microtubules in vitro was markedly stimulated and the assembled microtubules were greatly stabilized by p50. Further investigation of p50 is indispensable for the understanding of properties and function of intermediate-sized filaments in higher plant cells.

Cytokinesis in Spirogyra: Integration of Cleavage and Cell-Plate Formation

In Spirogyra, cytokinesis is initiated by a cleavage furrow. When the furrow contacts the elongating interzonal fibers of the telophase spindle, a small phragmoplast appears at the zone of contact, inside of which a cell-plate appears to complete cytokinesis. The use of two sequential mechanisms for achieving cytokinesis may be an intermediate, and possibly a transitional, state in the evolution of the phragmoplast found in higher plants. We tested the relative contribution of the two types of cytokinetic mechanisms by observing the effects of certain drugs on living cells, recorded in time-lapse. The antimicrotubule drug oryzalin causes the spindle to break down rapidly. When applied during early to midcytokinesis, the interzonal spindle collapses onto the cleavage furrow, which continues to grow inward. However, cytokinesis is not complete; examination of such cells 8-12 h later, after the nuclei separate, reveals a small aperture in the center of the cross-wall. This result shows that the phragmoplast/cell-plate is essential for complete cytokinesis, which apparently cannot be accomplished by cleavage alone, and that inward growth of the cleavage furrow is not dependent on microtubules. The antiactin drug cytochalasin D stops cleavage quite rapidly, and the eventual consequences of treatment depend on the stage reached in cleavage. If the cytochalasin is applied early in cytokinesis, cell division is unable to proceed further. Between telophase nuclei the cell does generate a large mass of cytoplasm that contains typical proliferating phragmoplast fibers; cell-plate formation is initiated in this mass but is never able to proceed further, and no coalescence of material into a cross-wall has been recorded. If cytochalasin is applied later, after the cleavage furrow has contacted the interzonal spindle fibers, cleavage stops but now cell-plate formation proceeds and cytokinesis is completed, albeit slowly. This result demonstrates that an interaction between the cleavage furrow and the forming phragmoplast is necessary for the latter to operate normally. These results support our suggestion that Spirogyra illustrates an intermediate stage in the evolution of the phragmoplast, since both cleavage and cell-plate formation are required for cytokinesis. (This suggestion does not require Spirogyra to be on the line of evolution leading to higher plants; rather, Spirogyra appears to be undergoing an evolutionary process similar to what might have occurred in their progenitors.) Furthermore, interaction of the cleavage furrow with the forming cell-plate/phragmoplast is required for cross-wall completion. These results may have significance in considering mechanisms by which higher plants locate their cellplates properly, an ability associated with the evolution and possible function(s) of the preprophase band of microtubules.

Endosperm Development in Barley: Microtubule Involvement in the Morphogenetic Pathway.

An immunofluorescence study of sectioned barley endosperm imaged by confocal laser scanning microscopy provided three-dimensional data on the relationship of microtubules to the cytoplasm, nuclei, and cell walls during development from 4 to 21 days after pollination (DAP). Microtubules play an important role throughout endosperm ontogeny. The syncytium is organized into units of nuclear-cytoplasmic domains by nuclear-based radial microtubule systems that appear to control the pattern of the first anticlinal walls at 5 to 6 DAP. After 7 DAP, phragmoplasts of two origins (interzonal and cytoplasmic) guide wall formation. Large compartments formed by the "free growing" walls in association with cytoplasmic phragmoplasts formed adventitiously at interfaces of opposing microtubule systems are subsequently subdivided by interzonal phragmoplast/cell plates to give rise to the starchy endosperm. During development of the aleurone layer from 8 to 21 DAP, the microtubule cycle is typical of plant histogenesis; cortical microtubules are hooplike, and preprophase bands of microtubules predict the division plane.

Embryo sac development in Arabidopsis thaliana II. The cytoskeleton during megagametogenesis

The microtubular and actin cytoskeletons have been investigated during megagametogenesis in Arabidopsis thaliana using immunofluorescence labelling of isolated coenocytic and mature embryo sacs. We found both actin and microtubules (MTs) to occur in abundance throughout megagametogenesis and in all constituent cells of the mature embryo sac. During many stages, the patterns of distribution of these cytoskeletal elements are congruent and may prove to be co-aligned. Many changes in the arrays of MTs and microfilaments take place and indicate varying roles of the cytoskeleton in the different stages and cell types of megagametogenesis. Two major populations of MTs recur throughout embryo sac formation: (1) Elaborate nuclear-based networks are found during the two-nucleate and four-nucleate developmental stages as well as in the egg cell. These arrays may function in positioning the nuclei. (2) Cytoplasmic MTs in longitudinal orientation in the two-nucleate embryo sac, synergids and part of the egg cell, or in a reticulate pattern in the four-nucleate embryo sac, egg and central cell probably participate in organization of the cytoplasm. Synergid MTs converge at the filiform apparatus. Preprophase bands of MTs are absent throughout megagametogenesis but phragmoplast arrays occur during cellularization of the embryo sac. Well developed arrays of cortical MTs are restricted to the antipodal cells. A large concentration of MTs in the part of the egg cell adjacent to the synergids is well placed for being involved with sperm cell movement within the degenerative synergid. On the basis of the morphology of the cytoskeleton, we concur with views that the shape of megagametophyte is largely determined by the surrounding tissues, including the integumentary tapetum.

Morphogenetic Plastid Migration and Microtubule Arrays in Mitosis and Cytokinesis in the Green Alga Coleochaete orbicularis

This study provides data on cell division in Coleochaete orbicularis, an important taxon in evolutionary theories deriving land plants from green algae. Vegetative growth in discoid species of Coleochaete results from marginal cell division in two planes—radial and circumferential. Like many algae and certain of the simple land plants, Coleochaete is monoplastidic. Prior to mitosis, the single plastid migrates to a position where it will divide and be distributed into the daughter cells. Unlike monoplastidic cell division in hornworts, mosses, and lycopsids; microtubule nucleation is not intimately associated with the plastids. Instead, microtubule organization is associated with centriolar centrosomes throughout the cell cycle, as is common in algae. The cytokinetic apparatus lacks preprophase bands of microtubules, but includes typical phragmoplasts consisting of brushlike arrays of microtubules on either side of a dark zone. However, the origin and role of phragmoplasts is unusual. Phragmoplasts appear to develop among microtubules that emanate from the polar centrosomes rather than from nuclear envelopes and/or plastids. The function of phragmoplasts in Coleochaete is unclear, as the process of cytokinesis is not strictly centrifugal. Some infurrowing occurs in radial division, and cytokinesis appears to be entirely centripetal by infurrowing in circumferential division. The cortical arrays of microtubules differ from those typical of land plants in that they develop as a network in association with centrosomes after mitosis.

Interphase and preprophase microtubule organization in some polarized cell types of the liverwort Marchantia paleacea Bert.

SUMMARYThe organization of interphase and preprophase‐prophase microtubules was studied in photosynthetic filament mother cells (PFMCs), photosynthetic filament cells (PFCs), mucilage papilla mother cells (MPMCs) and mucilage papillae (MP) of Marchantia paleacea Bert. These cell types exhibit polarized growth resulting, (a) in the formation of cell outgrowths followed by asymmetrical division (PFMCs and MPMCs), or (b) in the development of a tubular cell shape (PFCs and MP). The above cell types display an interphase cortical microtubule ring perpendicular to the axis of growth. In PFMCs and MPMCs it resides at the base of the cell outgrowth, while in PFCs and MP it has a median location. This microtubule ring is involved in the deposition of transverse, circumferentially aligned, cellulose microfibrils, defines the site where cell protrusion formation occurs and affects cell morphogenesis. In differentiated MP microtubules are rearranged in longitudinal or oblique cortical arrays. PFCs and MP also contain a prominent system of endoplasmic microtubules which may be involved in cytoplasmic polarization. In PFMCs, PFCs and MPMCs the interphase microtubule ring seems to function as a preprophase microtubule band (PPB). In the asymmetrically dividing PFMCs and MPMCs the PPB is complete but in symmetrically dividing PFCs it may be interrupted. The cell plate meets the parent wall at sites adjacent to the PPB cortical zone. These observations reveal further peculiarities in cortical microtubule organization and particularly in the PPBs of M. paleacea. Possible factors underlying the formation of complete or interrupted PPBs are discussed.

Cell Cycle in Potentially Dedifferentiating Cereal Mesophyll Protoplasts Cultured in vitro II. Behaviour of the Cytoskeleton

Summary Following up on the recent finding that mesophyll protoplasts of wheat and oats were able to reenter the cell cycle when cultured in vitro , the behaviour of the cytoskeleton was studied. The aim was to elucidate whether changes in the cytoskeletal patterns correlate with the abnormal progression through the cycle observed previously. Protoplasts freshly isolated from mature mesophyll tissue (type B) exhibited very few, fragmented microtubules, whereas mesophyll protoplasts isolated from immature leaf tissue (type A) showed an abundant network fo microtubules. Type B protoplasts of wheat, which were able to reenter but recalcitrant to complete S-phase, failed to reestablish a microtubular network. Type A wheat protoplasts capable of progressing from G1 to G2 but incapable of entering mitosis lost microtubules during culturing and eventually resembled type B protoplasts. In contrast to wheat, type A protoplasts of oats retained microtubules in culture and type B protoplasts reestablished an abundant network of microtubules after about 5 days in culture. Both types of protoplasts were capable of going through mitosis. Microtubule configurations typical of mitotic cells were observed. However, several unusual features became apparent. No preprophase bands of microtubules were found. The spindles were short and blurred and did not form distinct poles. Phragmoplast microtubules were observed, but the deposition of cell plate material seemed to be slow. The phragmoplasts disappeared and a cortical network of microtubles was reestablished, apparently before completion of cytokinesis. Actin microfilaments were found in all freshly isolated and cultured premitotic protoplasts. They were not detected during the mitosis of oat protoplasts, but were found again in phragmoplasts. They appeared to be associated in abundance with presumptive phragmosomes, even after disappearance of phragmoplast microtubules. The cell cycle abnormalities observed indicate a possible cause for the cereal protoplast recalcitrance.