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PubMed2005

Distinct segregation dynamics of the two Vibrio cholerae chromosomes.

The study of prokaryotic chromosome segregation has focused primarily on bacteria with single circular chromosomes. Little is known about segregation in bacteria with multipartite genomes. The human diarrhoeal pathogen Vibrio cholerae has two circular chromosomes of unequal sizes. Using static and time-lapse fluorescence microscopy, we visualized the localization and segregation of the origins of replication of the V. cholerae chromosomes. In all stages of the cell cycle, the two origins localized to distinct subcellular locations. In newborn cells, the origin of chromosome I (oriCIvc) was located near the cell pole while the origin of chromosome II (oriCIIvc) was at the cell centre. Segregation of oriCIvc occurred asymmetrically from a polar position, with one duplicated origin traversing the length of the cell towards the opposite pole and the other remaining relatively fixed. In contrast, oriCIIvc segregated later in the cell cycle than oriCIvc and the two duplicated oriCIIvc regions repositioned to the new cell centres. DAPI staining of the nucleoid demonstrated that both origin regions were localized to the edge of the visible nucleoid and that oriCIvc foci were often associated with specific nucleoid substructures. The differences in localization and timing of segregation of oriCIvc and oriCIIvc suggest that distinct mechanisms govern the segregation of the two V. cholerae chromosomes.

Chromosome Segregation↗

PubMed2006

Progressive segregation of the Escherichia coli chromosome.

We have followed the fate of 14 different loci around the Escherichia coli chromosome in living cells at slow growth rate using a highly efficient labelling system and automated measurements. Loci are segregated as they are replicated, but with a marked delay. Most markers segregate in a smooth temporal progression from origin to terminus. Thus, the overall pattern is one of continuous segregation during replication and is not consistent with recently published models invoking extensive sister chromosome cohesion followed by simultaneous segregation of the bulk of the chromosome. The terminus, and a region immediately clockwise from the origin, are exceptions to the overall pattern and are subjected to a more extensive delay prior to segregation. The origin region and nearby loci are replicated and segregated from the cell centre, later markers from the various positions where they lie in the nucleoid, and the terminus region from the cell centre. Segregation appears to leave one copy of each locus in place, and rapidly transport the other to the other side of the cell centre.

Cell Cycle↗

PubMed2000

DNA segregation in bacteria.

Segregation of DNA in bacterial cells is an efficient process that assures that every daughter cell receives a copy of genomic and plasmid DNA. In this review, we focus primarily on observations in recent years, including the visualization of DNA and proteins at the subcellular level, that have begun to define the events that separate DNA molecules. Unlike the process of chromosome segregation in higher cells, segregation of the bacterial chromosome is a continuous process in which chromosomes are separated as they are replicated. Essential to separation is the initial movement of sister origins to opposite ends of the cell. Subsequent replication and controlled condensation of DNA are the driving forces that move sister chromosomes toward their respective origins, which establishes the polarity required for segregation. Final steps in the resolution and separation of sister chromosomes occur at the replication terminus, which is localized at the cell center. In contrast to the chromosome, segregation of low-copy plasmids, such as Escherichia coli F, P1, and R1, is by mechanisms that resemble those used in eukaryotic cells. Each plasmid has a centromere-like site to which plasmid-specified partition proteins bind to promote segregation. Replication of plasmid DNA, which occurs at the cell center, is followed by rapid partition protein-mediated separation of sister plasmids, which become localized at distinct sites on either side of the division plane. The fundamental similarity between chromosome and plasmid segregation-placement of DNA to specific cell sites-implies an underlying cellular architecture to which both DNA and proteins refer.

Cell Compartmentation↗

PubMed2006

Involvement of synaptonemal complex proteins in sex chromosome segregation during marsupial male meiosis.

Marsupial sex chromosomes break the rule that recombination during first meiotic prophase is necessary to ensure reductional segregation during first meiotic division. It is widely accepted that in marsupials X and Y chromosomes do not share homologous regions, and during male first meiotic prophase the synaptonemal complex is absent between them. Although these sex chromosomes do not recombine, they segregate reductionally in anaphase I. We have investigated the nature of sex chromosome association in spermatocytes of the marsupial Thylamys elegans, in order to discern the mechanisms involved in ensuring their proper segregation. We focused on the localization of the axial/lateral element protein SCP3 and the cohesin subunit STAG3. Our results show that X and Y chromosomes never appear as univalents in metaphase I, but they remain associated until they orientate and segregate to opposite poles. However, they must not be tied by a chiasma since their separation precedes the release of the sister chromatid cohesion. Instead, we show they are associated by the dense plate, a SCP3-rich structure that is organized during the first meiotic prophase and that is still present at metaphase I. Surprisingly, the dense plate incorporates SCP1, the main protein of the central element of the synaptonemal complex, from diplotene until telophase I. Once sex chromosomes are under spindle tension, they move to opposite poles losing contact with the dense plate and undergoing early segregation. Thus, the segregation of the achiasmatic T. elegans sex chromosomes seems to be ensured by the presence in metaphase I of a synaptonemal complex-derived structure. This feature, unique among vertebrates, indicates that synaptonemal complex elements may play a role in chromosome segregation.

Animals↗

PubMed2005

Dynein promotes achiasmate segregation in Schizosaccharomyces pombe.

Most organisms use crossovers (chiasmata) to maintain physical connections between homologous chromosomes that ensure their proper segregation at the first meiotic division. The fission yeast Schizosaccharomyces pombe has a residual ability to segregate homologous chromosomes in the absence of meiotic recombination (achiasmate segregation). Using cytologically tagged chromosomes, we established a role for the microtubule motor dynein in meiotic chromosome segregation. Dhc1, the motor subunit of dynein, is required for chromosome segregation in both the presence and the absence of recombination. Dlc1, a member of the Tctex-1 dynein light-chain family, preferentially affects the segregation of achiasmate chromosomes. Dlc1 is the first identified protein, outside of Drosophila, that preferentially affects achiasmate chromosome segregation. We discuss possible roles of the dynein motor in this process.

Aneuploidy↗

PubMed1992

There are two mechanisms of achiasmate segregation in Drosophila females, one of which requires heterochromatic homology.

There are numerous examples of the regular segregation of achiasmate chromosomes at meiosis I in Drosophila melanogaster females. Classically, the choice of achiasmate segregational partners has been thought to be independent of homology, but rather made on the basis of availability or similarities in size and shape. To the contrary, we show here that heterochromatic homology plays a primary role in ensuring the proper segregation of achiasmate homologs. We observe that the heterochromatin of chromosome 4 functions as, or contains, a meiotic pairing site. We show that free duplications carrying the 4th chromosome pericentric heterochromatin induce high frequencies of 4th chromosome nondisjunction regardless of their size. Moreover, a duplication from which some of the 4th chromosome heterochromatin has been removed is unable to induce 4th chromosome nondisjunction. Similarly, in the absence of either euchromatic homology or a size similarity, duplications bearing the X chromosome heterochromatin also disrupt the segregation of two achiasmate X chromosome centromeres. Although heterochromatic regions are sufficient to conjoin nonexchange homologues, we confirm that the segregation of heterologous chromosomes is determined by size, shape, and availability. The meiotic mutation Axs differentiates between these two processes of achiasmate centromere coorientation by disrupting only the homology-dependent mechanism. Thus there are two different mechanisms by which achiasmate segregational partners are chosen. We propose that the absence of diplotene-diakinesis during female meiosis allows heterochromatic pairings to persist until prometaphase and thus to co-orient homologous centromeres. We also propose that heterologous disjunctions result from a separate and homology-independent process that likely occurs during prometaphase. The latter process, which may not require the physical association of segregational partners, is similar to those observed in many insects, in Saccharomyces cerevisiae and in C. elegans males. We also suggest that the physical basis of this process may reflect known properties of the Drosophila meiotic spindle.

Animals↗

PubMed1989

The claret locus in Drosophila encodes products required for eyecolor and for meiotic chromosome segregation.

The claret (ca) locus in Drosophila encodes products that are needed both for wild-type eyecolor and for correct meiotic chromosome segregation. Mutants described previously provide evidence that two mutationally independent coding regions are present at ca. We have recovered six new P element-induced and one spontaneous ca mutant. Four of these new mutants affect both eyecolor and chromosome segregation. The high frequency of co-mutation of these two functions suggests that the corresponding genes are closely adjacent to one another. We recovered genomic DNA sequences corresponding to the ca locus by chromosome walking, and showed using revertant analysis that the cloned region encodes ca+. Transformation experiments demonstrate that the mutant effect resulting in meiotic chromosome non-disjunction (nd) and loss is fully rescued by DNA from the cloned region. Two RNAs of 7.4 and 2.2 kb have been identified by Northern blot analysis as the putative eyecolor and segregational products. Expression of the RNAs with respect to males and females, and their presence or absence in ca and nd mutants indicate that the 7.4 kb RNA corresponds to the product needed for wild-type eyecolor and the 2.2 kb RNA is the product required for normal chromosome segregation. These RNAs are transcribed in opposite directions to one another. Alleles that affect both eyecolor and chromosome segregation are deletion mutants that affect both transcripts. Thus, the putative eyecolor and segregational products are encoded by separate genes. Mutants that affect both eyecolor and chromosome segregation apparently do so because they delete essential regions of both genes.

Alleles↗

PubMed1975

Non-mendelian segregation in hybrids between chinese hamster cells.

Mechanisms of segregation have been examined in hybrids between Chinese hamster cells, where chromosome loss in comparison to other systems is minimal. Hybrid cells were grown in HAT medium and subjected to back selection with bromodeoxyuridine (BUDR) or azaguanine (AZG). In AZG or BUDR at 30 mug/ml, segregation began with a random high frequency event that gave rise to cells capable of growth in both HAT and back selection medium, unlike the precursor hybrid or original parental cell types. BUDR-resistant segregants were propagated serially in the presence of BUDR, and were examined by clonal analysis for changes in plating properties during long term culture. Over a period of 300 days the HAT/BUDR plating ratio for sergregant cells declined continuously. A parallel decrease was observed in the rate of H3-thymidine incorporation, along with a drop in thymidine kinase activity. These shifts took place only in the presence of BUDR, and could be reversed by altered selection in HAT medium. Clonal studies showed that the evolution of segregant properties occurred in most if not all cells of the population, and did not arise from variation and selection of minority cell types. These properties of the segregating system are not consistent with models based on gene mutation, chromosome rearrangements, or chromosome loss. The evolution of segregants resembles more closely a sorting-out progress, taking place by intracellular selection over many generations. The segregating units may conceivably be cytoplasmic determinants linked functionally to nuclear genes, and which serve to modulate the events of phenotypic expression. Several lines of evidence which bear on this concept are discussed.

Azaguanine↗

PubMed1977

Chromosome loss is responsible for segregation at the HPRT locus in Chinese hamster cell hybrids.

The phenomenon of segregation of gene expression has been examined in intraspecific somatic cell hybrids. Specifically, segregation at the hypoxanthine guanine phosphoribosyltransferase (HPRT) locus has been studied in hybrids of Chinese hamster cell lines. The role of chromosome segregation, or other chromosomal events has been assessed by detailed comparison of karyotypes in the 6-thioguanine resistant segregants with those of the parental hybrid lines. The results clearly demonstrate that loss of an entire X chromosome is the primary event responsible for segregation at the HPRT locus, while deletion of a portion of the short arm of an X chromosome was also a frequent event. The results provide the first direct evidence for the assignment of the mapping of this locus to the distal region of the short arm. Analysis of chromosome number distributions in the hybrids and segregants suggests that in selecting chromosomal segregants one may also select for hybrid lines with reduced chromosome stability.

Cell Line↗

PubMed2002

Resistance gene-analog polymorphism markers co-segregating with the YR5 gene for resistance to wheat stripe rust.

The Yr5 gene confers resistance to all races of the stripe rust pathogen ( Puccinia striiformis f. sp. tritici) of wheat in the United States. To develop molecular markers for Yr5, a BC(7):F(3) population was developed by backcrossing the Yr5 donor ' Triticum spelta album' (TSA) with the recurrent parent 'Avocet Susceptible' (AVS). Seedlings of the Yr5 near-isogenic lines (AVS/6* Yr5), AVS, TSA, and the BC(7):F(3) lines were tested with North American races of P. striiformis f. sp. tritici under controlled greenhouse conditions. The single gene was confirmed by a 1:2:1 segregation ratio for homozygous-resistant, heterozygous and homozygous-susceptible BC(7):F(3) lines. Genomic DNA was extracted from the parents (the Yr5 near-isogenic line and AVS) and 202 BC(7):F(3) lines. The resistance gene-analog polymorphism (RGAP) technique was used to identify molecular markers. The parents and the homozygous-resistant and homozygous-susceptible BC(7):F(3) bulks were used to identify putative RGAP markers for Yr5. Association of the markers with Yr5 was determined using segregation analysis with DNA from the individual BC(7):F(3) lines. Of 16 RGAP markers confirmed by segregation analysis with 109 BC(7):F(3) lines, and nine of the markers confirmed with an additional 93 BC(7):F(3) lines, three markers co-segregated with the resistance allele and three markers co-segregated with the susceptibility allele at the Yr5 locus. The other four markers were tightly linked to the locus. Analysis of a set of Chinese Spring nulli-tetrasomic lines with three markers that co-segregated with, or were linked to, the susceptibility allele confirmed that the Yr5 locus is on chromosome 2B. Of five RGAP markers that were cloned and sequenced, markers Xwgp-17 and Xwgp-18 that co-segregated with the Yr5 locus were co-dominant and had 98% homology with each other in both DNA and translated amino-acid sequences. The two markers had 97% homology with a resistance gene-like sequence from Aegilops ventricosa and had significant homology with many known plant resistance genes, resistance gene analogs and expressed sequence tags (ESTs) from wheat and other plant species. The markers Xwgp-17 and Xwgp-18 also had significant homology with the NB-ARC domain that is in several genes for plant resistance to diseases, nematode cell death and human apoptotic signaling. These markers should be useful to clone Yr5 and combine Yr5 with other genes for durable and superior resistance for the control of stripe rust.

Amino Acid Sequence↗

PubMed2003

Study of recombinant micro-organism populations characterized by their plasmid content per cell using a segregated model.

Numerous observations from recombinant systems have shown that properties such as the specific cell growth rate and the plasmid-free cell formation rate are related, not only to the average plasmid content per cell, but also to the plasmid distribution within a population. The plasmid distribution in recombinant cultures can have an effect on the culture productivity that cannot be modelled using average values of the overall culture. The prediction of the behaviour of a plasmid content distribution and its causes and effects can only be studied using segregated models. A segregated model that describes populations of recombinant cells characterized by their plasmid content distribution has been developed. This model includes critical causes of recombinant culture instability such as the plasmid partition mechanism at cell division, plasmid replication kinetics and the effect of the plasmid content on the specific growth rate. The segregated model allows investigation of the effect of each of these causes and that of the plasmid content distribution on the observable behaviour of a recombinant culture. The effect of two partitioning mechanisms (Gaussian distribution and binomial distribution) on culture stability was investigated. The Gaussian distribution is slightly more stable. A small plasmid replication rate constant results in a very unstable culture even after short periods of time. This instability is dramatically improved for a larger value of this constant, hence improving protein synthesis. For a very narrow initial plasmid distribution, a given plasmid replication rate and partitioning mechanism can become broad even after a relatively short period of time. In contrast, a very "broad" initial distribution gave rise to a "Gamma-like" distribution profile. If we compare the results obtained in the simulations of the segregated model with those of the non-segregated one (average model), the latter model predicts much more stable behaviour, thus these average models cannot predict culture instability with the same precision. When compared with the experimental results, the segregated model was able to predict the practical behaviour with accuracy even in a system with a high plasmid content per cell and a high rate of plasmid-free cell formation which could not be achieved with a non-segregated model.

Journal Article↗

PubMed1978

Particle segregation in chromaffin granule membranes by forced physical contact.

Bovine chromaffin granules were exposed to different isotonic non-ionic and ionic solutions (sucrose; Ca2+- and Mg2+-free phosphate-buffered saline; Tris-HCl + NaCl; Ca2+- and Mg2+-free phosphate-buffered saline + sucrose; Tris-HCl + sucrose) at pH 7 and then frozen either in suspension or as firm pellets. Freezing was performed without prefixation or antifreeze treatments either by 'standard' techniques (approx. 1 mm3 suspended or pelleted material on gold specimen supports dipped into liquid Freon) or with increased cooling rates by spraying suspensions into liquid propane ('spray-freezing'). Regardless of the freezing method, membrane-intercalated particles were always randomly distributed when chromaffin granules were frozen in suspension. In contrast, forced physical contact between granules produced by centrifugation (12000 X g, 25 min) provoked dispersal of membrane-intercalated particles, but only in the presence of ions. Sucrose or EDTA in an ionic environment had no inhibitory effect. The following conclusions are derived: (1) Even below the reported phase transition region particle clustering is possible. (2) Chromaffin granule membranes are not liable to thermotropic segregation of membrane-intercalated particles. (3) Although the low freezing rates of 'standard' freezing techniques produce large-scale segregation artefacts (by which suspended chromaffin granules are pushed together within the segregated solute) this does not result in intramembraneous particle segregation. (4) Forced physical contact produces a Ca2+-independent particle segregation, but only when repulsive electrostatic forces of membrane components are partially screened in an ionic environment. (5) This does not invalidate results obtained by others, showing Ca2+-mediated chromaffin granules agglomeration and segregation of membrane-intercalated particles, but it might indicate the occurrence of another, not directly Ca2+-dependent particle segregation mechanism in a prefusional stage of close membrane-to-membrane contact during exocytosis.

Animals↗

PubMed1998

Texture segregation, surface representation and figure-ground separation.

A widespread view is that most texture segregation can be accounted for by differences in the spatial frequency content of texture regions. Evidence from both psychophysical and physiological studies indicate, however, that beyond these early filtering stages, there are stages of 3-D boundary segmentation and surface representation that are used to segregate textures. Chromatic segregation of element-arrangement patterns--as studied by Beck and colleagues--cannot be completely explained by the filtering mechanisms previously employed to account for achromatic segregation. An element arrangement pattern is composed of two types of elements that are arranged differently in different image regions (e.g. vertically on top and diagonally on the bottom). FACADE theory mechanisms that have previously been used to explain data about 3-D vision and figure-ground separation are here used to simulate chromatic texture segregation data, including data with equiluminant elements on dark or light homogeneous backgrounds, or backgrounds composed of vertical and horizontal dark or light stripes, or horizontal notched stripes. These data include the fact that segregation of patterns composed of red and blue squares decreases with increasing luminance of the interspaces. Asymmetric segregation properties under 3-D viewing conditions with the equiluminant elements close or far are also simulated. Two key model properties are a spatial impenetrability property that inhibits boundary grouping across regions with non-collinear texture elements and a boundary-surface consistency property that uses feedback between boundary and surface representations to eliminate spurious boundary groupings and separate figures from their backgrounds.

Color Perception↗

PubMed1999

A comparison of the dynamics of simple (Fourier) and complex (non-Fourier) mechanisms in texture segregation.

Models of texture segregation frequently feature two processing mechanisms: simple, linear channels (1st-order, Fourier mechanisms) and complex channels (2nd-order, non-Fourier mechanisms). Using texture patterns designed to segregate primarily as a result of activity in one set of channels or the other, we employed the method of cued response to obtain speed-accuracy tradeoff (SAT) functions measuring the time course of texture segregation processing in simple and complex channels. Here, both simple-channel and complex-channel patterns are composed of Gabor-patch texture elements, thus equating input to simple channels and the first stage of complex channels. Subjects were required to identify the orientation of a rectangular texture-region embedded in a background field of a different texture. SAT functions were obtained by requiring subjects to respond within 200 ms after an auditory cue. We found that: (1) when segregation depended primarily on activity in simple channels, performance was faster and better than when it depended primarily on complex channels; (2) in contrast to a previous study (Sutter, A., & Graham, N. (1995). Investigating simple and complex mechanisms in texture segragation using the speed-accuracy tradeoff method. Vision Research, 35, 2825-2843), simple-channel (Fourier) patterns composed of two textured regions were just as easily segregated as simple-channel patterns in which one of the regions was blank instead of textured; (3) performance with complex-channel patterns composed of diagonally oriented Gabor-patches was considerably worse than performance with complex-channel patterns composed of vertically and/or horizontally oriented Gabor-patches; (4) among simple-channel patterns containing only one region of texture (background-only or rectangle-only), there were minimal differences in performance; and (5) as in previous experiments, there were large individual differences in the segregation of complex-channel (non-Fourier) patterns. All of the above results can be explained within the framework of the simple- and complex-channels model of texture segregation.

Adult↗

PubMed2003

Partial segregation of posterior crista and saccular fibers to the nodulus and uvula of the cerebellum in mice, and its development.

The projection of the posterior canal crista and saccular afferents to the cerebellum of embryonic and neonatal mice was investigated using carbocyanine dyes. Anterograde tracing from these two endorgans reveals a partial segregation of these two sets of afferents. The saccule projects predominantly to the uvula, with very minor input to the nodulus. The posterior canal projects mainly to the nodulus and, to a lesser extent, to the uvula. Retrograde tracing from the uvula and nodulus confirms this partial segregation for these two endorgans and extends it to other vestibular endorgans. Uvular injections result in many more labeled fibers in the gravistatic maculae than in the canals' cristae. In contrast, nodular injection reveals many more labeled fibers in the canal cristae than in the gravistatic maculae. This partial segregation may play a role in the information processing in these folia. Our developmental data suggest that the initial segregation at E17 coincides with the formation of the postero-lateral fissure. This embryonic segregation of the primary vestibular mossy fibers to the uvula and nodulus commences long before the maturity of their targets, the granule cells and unipolar brush cells. Thus, the segregation of the primary vestibular projection to the uvula and nodulus does not depend on cues related to the target cells. Rather, the segregation may reflect more global cerebellar patterning mechanisms involving guidance for the vestibular afferent fibers independent of the future target cells.

Aging↗

PubMed2003

Application of automated crystallography for transmission electron microscopy in the study of grain-boundary segregation.

Analytical Electron Microscopy (AEM) has brought significant progress in the study of grain-boundary segregation. Using X-ray energy-dispersive spectrometry (XEDS) in the AEM, elemental segregation information can be related to the crystallographic character of the same boundary via conventional Transmission Electron Microscope (TEM) diffraction techniques. While significant efforts have been made to improve XEDS analysis of sub-nanometer segregation layers, the methods for crystallographic characterization of grain boundaries have remained the same for several decades and labor-intensive processes. Recently, a method termed Automated Crystallography for TEM (ACT) was developed, which automates crystallographic characterization of grains under TEM observation. In the present work, we combine ACT and X-ray mapping via EDS in AEM for the study of Sb grain-boundary segregation in a rapidly solidified Cu-0.08 wt % Sb alloy. In contrast with previous reports, a large degree of anisotropy in Sb segregation level between different boundaries is found. ACT results suggest that one of the several grain boundaries observed with no detectable Sb segregation is very close to a Sigma 3 coincidence-site lattice structure. The reason for the observed anisotropy in the present alloy is discussed, based upon McLean's theory of segregation.

Journal Article↗

PubMed2002

The segregation of the Escherichia coli origin and terminus of replication.

Escherichia coli chromosome replication forks are tethered to the cell centre. Two opposing models describe how the chromosomes segregate. In the extrusion-capture model, newly replicated DNA is fed bi-directionally from the forks toward the cell poles, forming new chromosomes in each cell half. Starting with the origins, chromosomal regions segregate away from their sisters progressively as they are replicated. The termini segregate last. In the sister chromosome cohesion model, replication produces sister chromosomes that are paired along much of their length. The origins and most other chromosomal regions remain paired until late in the replication cycle, and all segregate together. We use a combination of microscopy and flow cytometry to determine the relationship of origin and terminus segregation to the cell cycle. Origin segregation frequently follows closely after initiation, in strong support of the extrusion-capture model. The spatial disposition of the origin and terminus sequences also fits this model. Terminus segregation occurs extremely late in the cell cycle as the daughter cells separate. As the septum begins to invaginate, the termini of the completed sister chromosomes are transiently held apart at the cell centre, on opposite sides of the cell. This may facilitate the resolution of topological linkages between the chromosomes.

Base Sequence↗

PubMed2000

On the mechanism of ooplasmic segregation in single-cell zebrafish embryos.

It has been previously shown that localized elevations of free cytosolic calcium are associated with a morphological contraction in the forming blastodisc and animal hemisphere cortex during ooplasmic segregation in zebrafish zygotes. It was subsequently proposed, in a hypothetical model, that these calcium transients might be linked to the contraction of a cortically located actin microfilament network as a potential driving force for segregation. Here, by labeling single-cell embryos during the major phase of segregation with rhodamine-phalloidin, direct evidence is presented to indicate that the surface contraction was generated by an actin-based cortical network. Furthermore, while zygotes incubated with colchicine underwent normal ooplasmic segregation, those incubated with cytochalasin B did not generate a constriction band or segregate to form a blastodisc. During segregation at the single-cell stage, ooplasm simultaneously moved in two directions: toward the blastodisc within the so-called axial streamers, and toward the vegetal pole in the peripheral ooplasm. The velocities of both axial and peripheral streaming movements are reported. By injection of a fluorescein isothiocyanate (FITC)-labeled 2000 kDa dextran into the peripheral ooplasm it was demonstrated that a portion of it feeds into the bases of the extending streamers, which helps to explain the lack of accumulation of ooplasm at the vegetal pole. These new data were incorporated into the original model to link the bipolar ooplasmic movements with the calcium-modulated, actin-mediated contraction of the animal hemisphere cortex as a means of establishing and driving ooplasmic segregation in zebrafish.

Actins↗