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B Maro

Publications and source records attributed to B Maro.

At least 73 records · Page 4Linked to original sources
PubMed1986

Mechanism of polar body formation in the mouse oocyte: an interaction between the chromosomes, the cytoskeleton and the plasma membrane.

The influence of mouse oocyte chromosomes on their immediate environment has been investigated following their dispersal by dissolution of the metaphase spindle with nocodazole. Small clusters of chromosomes become redistributed around the egg cortex in a microfilament-dependent process. Each cluster has the capacity, on removal from nocodazole, to organize a spindle that rotates to yield a polar body. In this process of spindle formation, the chromosome clusters are able both to promote tubulin polymerization in their vicinity and to recruit microtubule-organizing centres (MTOCs) which organize the polymerized tubulin into spindles. In addition each oocyte chromosome cluster, as well as the non-dispersed sperm-derived haploid group of chromosomes, induces a focal accumulation of subcortical actin (corresponding to a filamentous area devoid of organelles) and a loss of surface Concanavalin A binding activity (corresponding to a loss of surface microvilli) in the overlying cortex. This induction ceases with the formation of pronuclei whether or not the pronuclei migrate centrally. Pronuclear formation is sensitive to the action of nocodazole for up to 2-4 h postinsemination, and pronuclear migration is totally sensitive to the drug. If pronuclei are blocked in a peripheral location by nocodazole they are associated with an elevation in Con A binding activity of the overlying membrane which corresponds to an area of the surface rich in blebby microvilli.

Animals↗
PubMed1986

Parthenogenesis and cytoskeletal organization in ageing mouse eggs.

The cytoskeletal organization of the mouse egg changes during ageing in vivo and in vitro. The earliest change observed is the disappearance of the microfilament-rich area overlying the meiotic spindle. This is followed by the migration of the spindle towards the centre of the egg. Finally the spindle breaks down and the chromosomes are no longer organized on a metaphase plate. This spindle disruption may result from changes in the microtubule nucleating material found at the spindle poles and from an increase in the critical concentration for tubulin polymerization. It is possible to correlate the changes in the cytoskeletal organization of the egg occurring during ageing with the different types of parthenogenetic embryos obtained after ethanol activation. These observations strengthen the hypothesis that the actin-rich cortical area that overlies the meiotic spindle forms a domain to which the meiotic cleavage furrow is restricted and provides some insights into the mechanisms by which different types of parthenogenetic embryos are generated.

Actins↗
PubMed1986

The generation of cell surface polarity in mouse 8-cell blastomeres: the role of cortical microfilaments analysed using cytochalasin D.

The mechanism by which a surface pole of microvilli is generated in mouse 8-cell blastomeres has been investigated. 4-cell and 8-cell embryos (or cell couplets) were incubated for precise times during their respective cell cycles in medium containing cytochalasin D (CCD) to disrupt the microfilament system. The blastomeres were analysed immediately for the distribution and state of organization of their microvilli, using three morphological techniques. The results indicate that the surface pole, characterized by microvilli containing CCD-resistant core filaments, is not generated by the gradual segregation of stable microvilli to the apical surface. An alternative model is proposed, based upon the stabilization of the apical cytocortex prior to the elongation of apical microvilli and the destabilization of cytocortical elements in contact areas and the shortening and loss of basolateral microvilli.

Actin Cytoskeleton↗
PubMed1986

The timing of compaction: control of a major developmental transition in mouse early embryogenesis.

The effect of protein synthesis inhibitors on compaction of the 8-cell mouse embryo has been investigated. The effects observed depended upon the duration and time of drug application and on the features of compaction scored. Continuous application from the late 2-cell or early 4-cell stages allowed cell flattening and surface polarization to occur in most embryos and advanced development of these features in many of them. Cell coupling developed only when drug addition was delayed until the mid 4-cell stage, and cytoplasmic polarization developed only when drug addition was delayed until the late 4-cell stage. We suggest that control over the timing of compaction is achieved at a post-translational level via a global permissive change within the blastomeres of the embryo.

Animals↗
PubMed1985

Fate of microtubule-organizing centers during myogenesis in vitro.

Microtubule organization and nucleation were studied during in vitro human myogenesis by immunocytology that used monoclonal and polyclonal antitubulin antibodies and a rabbit nonimmune serum that reacts with human centrosomes. In myoblasts, we observed a classical microtubule network centered on juxtanuclear centrosomes. Myotubes possessed numerous microtubules organized in parallel without any apparent nucleation centers. Centrosomes in these cells were not associated one to each nucleus but were often clustered in the vicinity of nuclei groups. They were significantly smaller than those of the mononucleated cells. The periphery of each nucleus in myotubes was labeled with the serum that labels centrosomes suggesting a profound reorganization of microtubule-nucleating material. Regrowth experiments after Nocodazole treatment established that microtubules were growing from the periphery of the nuclei. The redistribution of nucleating material was shown to take place early after myoblast fusion. Such a phenomenon appears to be specific to myogenic differentiation in that artificially induced polykaryons behaved differently: the centrosomes aggregated to form only one or a few giant nucleating centers and the nuclei did not participate directly in the nucleation of microtubules. The significance of these results is discussed in relation to the possible role of the centrosome in establishing cell polarity.

Cell Division↗
PubMed1985

Non-spindle microtubule organizing centers in metaphase II-arrested mouse oocytes.

A human autoantiserum (5051) directed against pericentriolar material (PCM) was used to study the distribution of microtubule-organizing centers (MTOCs) in the oocyte and during the first cell cycle of mouse development. In oocytes, the PCM was found not only at the poles of the barrel-shaped metaphase II spindle but also at many discrete loci around the cytoplasm near the cell cortex. The spindle poles were also composed of several PCM foci. In metaphase-arrested eggs only the PCM foci located near the chromosomes acted as MTOCs. However, after reduction of the critical concentration for tubulin polymerization by taxol, the cytoplasmic PCM foci were also found to be associated with nucleation of microtubules. After fertilization the cortical PCM foci remained in a peripheral position until the end of the S phase, when they appeared to migrate centrally towards the pronuclei. At prometaphase of the first mitotic division, numerous MTOCs were found around the two sets of chromosomes; these MTOCs then aligned to form two bands on either side of the metaphase plate of the first mitosis.

Alkaloids↗
PubMed1985

A dissection of the mechanisms generating and stabilizing polarity in mouse 8- and 16-cell blastomeres: the role of cytoskeletal elements.

Pairs of 8-cell or 16-cell blastomeres were cultured for up to 9 h after their formation from isolated 1/4 or 1/8 blastomeres respectively. Blastomeres were examined for the incidence and orientation of their surface polarity, as assessed by binding of FITC-Con A and by distribution of microvilli, and of their cytoplasmic polarity, as assessed by distribution of cytoplasmic actin, clathrin and a 100 kD antigen associated with the lysosomal/acid vesicle fraction of membranous organelles. The effect on polarity of incubating the pairs of cells in taxol, nocodazole, cytochalasin D or in a combination of nocodazole plus cytochalasin D for different parts of the incubation period was examined. Neither the development nor the stability of the surface polarity in 8-cell blastomeres was blocked by any treatment and only the use of CCD in combination with nocodazole affected the incidence of surface polarity appreciably. However, with some treatments, the form and position of the surface poles were modified. In the presence of microtubule inhibitors surface poles extended over a larger area of the cell surface, while exposure to CCD led to poles that were not opposite to the contact point between cells. In contrast to surface polarity, the development of cytoplasmic polarity was suppressed by both microtubule- and microfilament-inhibiting drugs, which also reversed it rapidly. In polar 16-cell blastomeres surface polarity was influenced in a similar manner to that of 8-cell blastomeres, only the combined use of cytochalasin D and nocodazole having any major effect. Polarization of clathrin in polar 16-cell blastomeres was inhibited almost completely by all drug treatments applied including cytochalasin D. The focal concentration of lysosomal antigen that occurs during the 16-cell stage was reduced only in the continuous presence of nocodazole plus cytochalasin D, but once established was not reversed appreciably by any drug. However, the localization of the lysosomal antigen to the basal region of polarized cells did not seem to occur in the presence of any drug. The dissociation of surface and cytoplasmic polarity revealed in these experiments leads us to conclude that either surface polarity is a prerequisite for the organization of cytoplasmic polarity, and mediates the latter via the cytoskeleton, or surface and cytoplasmic polarity develop by parallel but separate mechanisms.

Alkaloids↗
PubMed1985

Changes in the distribution of membranous organelles during mouse early development.

The unfertilized oocyte, fertilized egg and early embryo (2-cell to 16-cell) of the mouse have been examined immunocytochemically for the distribution of antigens associated with the endoplasmic reticulum, the lysosomal and acidic vesicle fraction (100 kD antigen), Golgi apparatus (135 kD antigen) and coated vesicles (clathrin). The distribution of these antigens has also been examined in isolated 8-cell and 16-cell-stage blastomeres of various ages and phenotypes. Endoplasmic reticulum is detected only weakly in the oocyte and egg, but is seen abundantly at later stages both in association with the nuclear membrane and evenly distributed throughout the cytoplasm, except in regions of cell:cell apposition from which it is excluded. Intracellular clathrin is associated with the spindle in mitotic and meiotic cells. During interphase, clathrin is distributed throughout the cell until the mid-8-cell stage when it is concentrated into the apical region of the cell under the region of membrane at which a surface pole of microvilli will form subsequently. Thus, the cytoplasmic polarization of clathrin precedes overt polarization at the surface. At mitosis, the clathrin relocates to the spindle and is distributed to both daughter cells. It resumes an apical location beneath the surface pole of microvilli in polar daughter 1/16 cells, but remains dispersed in apolar daughter 1/16 cells. Both the lysosomal and Golgi antigens are distributed throughout the cytoplasm until the early 16-cell stage. In pairs of 16-cell blastomeres both antigens aggregate in a single cluster and do so whether the surface phenotype of the blastomeres is polar or apolar. The position of this cluster is not consistently related to the point of contact with the other cell in the pair but there is a suggestion that in cells with a polar surface phenotype the polar foci of Golgi/lysosomal antigens are located between the nucleus and the surface pole at earlier time points, but shift to a position between the basolateral membrane and the nucleus at the later time point. In intact 16-cell embryos also, the aggregated Golgi/lysosomal antigens of polar cells appear to localize to the basal region. The distributions of these various organelles in embryonic cells reported here show a number of differences from those reported previously for mature, differentiated cells.

Animals↗
PubMed1985

Meiosis II, mitosis I and the linking interphase: a study of the cytoskeleton in the fertilised mouse egg.

Data concerning the organisation and the role of microtubules and microfilaments during the 20 h following fertilisation in vitro of the mouse oocyte is reviewed. This period covers the fertilisation-induced completion of meiosis, from second meiotic metaphase through the first mitotic cell cycle, and involves the union of the haploid parental genomes. Various antibodies were used against actin, tubulin and PCM to describe the localisation of the cytoskeletal components and the two cytoskeleton-disrupting drugs nocodazole and CCD to assess the functional importance of the cytoskeleton.

Animals↗
PubMed1984

Vimentin filaments and centrosomes. Are they associated?

HeLa cells were examined by immunofluorescence using anti-vimentin and anti-centrosphere anti-bodies, and by transmission electron microscopy (TEM), after vimentin redistribution induced by the action of nocodazole or taxol. A redistribution of vimentin bundles in the centriolar area was observed after nocodazole treatment, although no direct interaction between centrioles and vimentin filaments could be detected. After taxol treatment, the juxtanuclear accumulation of vimentin filaments and the centrioles were rarely observed in the same area. Our results do not support the concept of a direct association between centrioles and vimentin filaments.

Alkaloids↗
PubMed1984

Microtubules influence compaction in preimplantation mouse embryos.

The role of microtubules during compaction of the 8-cell-stage mouse embryo was investigated using the drugs Taxol (which leads to a non-controlled polymerization of tubulin) and Nocodazole (which causes depolymerization of microtubules). Taxol inhibits compaction in most non-compacted embryos and reverses it in already compacted embryos. These effects were observed on both cell flattening (as judged by phase-contrast microscopy) and on cell surface polarization (as judged by scanning electron microscopy and the surface binding of fluorescent concanavalin A). In contrast Nocodazole does not inhibit cell flattening, but rather accelerates its completion. Nocodazole influences the detailed organization of the surface pole and appears to reduce the incidence of surface polarization but does not reverse polarity once established to a significant extent. We conclude that microtubules exercise a constraining role during compaction, influencing cell shape, cell organization and the time at which compaction takes place.

Alkaloids↗
PubMed1984

The distribution of cytoplasmic actin in mouse 8-cell blastomeres.

Three non-homogeneous patterns of cytoplasmic actin distribution have been demonstrated in pairs of 8-cell blastomeres. Newly formed blastomeres showed an actin distribution associated with the remnant of the previous mitotic spindle. Subsequently blastomeres showed a zonal clearing of actin from regions of intercellular contact, the extent of the clearing increasing with the extent of contact. A polarized distribution of actin was evident from the early to mid 8-cell stage and coincided with the movement of nuclei towards the point of intercellular contact. The detection of polar actin preceded by 2-4 h the detection of a surface polarity as assessed by the FITC-Con A binding pattern and the distribution of cortical microvillous actin. However, once a surface pole of microvilli had formed, it persisted under conditions which led to loss of polar cytoplasmic actin. Incubation in cytochalasin D (CCD) resulted in a dispersed homogeneous pattern of actin distribution but did not prevent the formation of surface poles as assessed both by the Con A binding pattern and detection of polar microvilli. However, the poles formed were less clearly defined and the density and length of microvilli within them was variable. Moreover, when CCD was added early during the 8-cell stage the position of the poles was frequently not on an axis perpendicular to the point of intercellular contact. Cytochalasin D also affected the movement of the nucleus that occurs during the process of polarization. On the basis of these experiments, we conclude that actin is likely to be involved in the events of polarization, but that its precise role remains to be determined.

Actins↗
PubMed1984

Changes in actin distribution during fertilization of the mouse egg.

Unfertilized mouse oocytes and eggs 1-8 h after fertilization in vitro were examined at the light microscope level for structural changes, distribution of actin (as assessed by both antiactin antibodies and NBD-phallicidin), surface binding of concanavalin A (Con A) and chromosomal distribution and condensation. The influence of cytochalasin D on these events was also assessed. Changes in actin distribution were associated with rotation of the second anaphase spindle, formation of the second polar body, the events following incorporation of the sperm nucleus, and formation and migration of the pronuclei. Cytochalasin D prevented spindle rotation, polar body formation, pronuclear migration and the restoration of Con A binding over the site of sperm entry and the site of polar body formation, but did not affect sperm fusion and entry, the loss of Con A binding at the site of sperm entry and pronuclear formation.

Actins↗
PubMed1982

In vivo and in vitro effects of the mitochondrial uncoupler FCCP on microtubules.

FCCP (carbonylcyanide-p-trifluoromethoxyphenylhydrazone), a potent uncoupler of oxidative phosphorylation, induces the complete disruption of cellular microtubules. A further analysis of this effect on BHK21 cells has shown that a decrease in the number of microtubules can be observed 15 min after adding FCCP and there is complete disruption after 60 min. Regrowth of microtubules was initiated 30 min after removal of FCCP, in marked contrast with the rapid reversion observed when microtubules are disrupted by nocodazole. A similar delay was required for the recovery of mitochondrial function as assessed by rhodamine 123 labelling. The effect of FCCP on microtubules was partially inhibited by preincubation of the cells with NaN3, suggesting that FCCP acts on microtubules through mitochondria. FCCP did not depolymerize microtubules of cells permeabilized with Triton X-100. In vitro polymerisation of microtubule protein was only slightly diminished by concentrations of FCCP which provoke complete disassembly in vivo. SDS-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the microtubules polymerized in vitro in the presence of FCCP showed a reduced amount of high mol. wt. proteins, mainly MAP 2, associated with them. In an attempt to reproduce the mitochondrial effects of FCCP in vitro, we checked the effects of alkaline pH and calcium on microtubule protein polymerization in the presence of FCCP. FCCP did not influence the calcium inhibitory effect but did significantly increase the inhibitory effect of alkaline pH. We conclude that FCCP could depolymerise microtubules in vivo through a dual operation: increasing the intracellular pH by the disruption of the mitochondrial H+ gradient and decreasing the stability of microtubules by impairing the binding of microtubule-associated proteins.

Animals↗