pH regulates the polymerization of actin in the sea urchin egg cortex

The state of actin in the isolated cortex of the unfertilized sea urchin egg can be controlled by experimentally manipulating the pH of the isolation medium. Cortices isolated at the pH of the unfertilized egg (6.5--6.7) do not contain filamentous actin, while those isolated at the pH of the fertilized egg (7.3--7.5) develop large numbers of microvilli which contain bundles of actin filaments. Cortices that are isolated at pH 6.5 and then transferred to isolation medium buffered at pH 7.5 also develop actin filaments. However, the filaments are not arranged in bundles and microvilli do not form. Although the cortical granules in cortices isolated at pH 6.5 discharge at a free Ca++ concentration of approximately 10 micrometer, actin polymerization is not induced by increasing the Ca++ concentration of the isolation medium. These results suggest that the increase in cytoplasmic pH which occurs following fertilization induces the polymerization of actin in the egg cortex.     

A yellow crescent cytoskeletal domain in ascidian eggs and its role in early development

In this investigation, Triton X-100 extraction was utilized to examine the cytoskeleton of ascidian eggs and embryos. The cytoskeleton contained little carbohydrate or lipid and only about 20–25% of the total cellular protein and RNA. It was enriched in polypeptides of molecular weight (M_r) 54, 48, and 43 × 10³. The 43 × 10³M_r polypeptide was identified as actin based on its M_r, isoeletric point, and affinity for DNase I. Electron microscopy of the detergent-extracted eggs showed that they contained cytoskeletal domains corresponding to colored cytoplasmic regions of specific morphogenetic fate in the living egg. A yellow crescent cytoskeletal domain in the myoplasm was examined and shown to consist of a plasma membrane lamina (PML) and a deeper lattice of filaments which appeared to connect the yellow crescent pigment granules to the PML. The PML probably consists of integral membrane proteins stabilized by an underlying network of actin filaments since NBD-phallacidin stained this area of the egg cortex and the PML was extracted from the cytoskeleton by DNase I treatment. The yellow crescent cytoskeletal domain was found throughout the cortex of the unfertilized egg. During ooplasmic segregation it progressively receded into the vegetal hemisphere and was subsequently partitioned to the presumptive muscle and mesenchyme cells of the 32-cell embryo. It is suggested that contraction of the actin network in the yellow crescent cytoskeletal domain is the motive force for ooplasmic segregation. This structure may also serve as a framework for the positioning of morphogenetic determinants involved in muscle cell development.     

Development of cortical polarity in mouse eggs: involvement of the meiotic apparatus

Experiments were carried out to determine the origin of cortical polarity in mouse eggs and its possible relation to the meiotic apparatus. Cortices of mature eggs overlying the meiotic apparatus (microvillus-free area) were distinguished by an absence of microvilli and a thickened layer of actin. In contrast, the surfaces of immature oocytes were covered entirely with a dense population of microvilli and were subtended by a uniform layer of actin. When induced to undergo maturation, meiotic spindles formed in the center of immature oocytes and then moved peripherally. Coincident with the cortical localization of the meiotic spindle was the formation of a microvillus-free area, i.e., a loss of microvilli and a thickening of the actin layer associated with this region of the egg cortex. If immature oocytes were incubated in cytochalasin B, meiotic spindles formed; however, they failed to move peripherally and microvillus-free areas did not develop. Oocytes incubated in colchicine did not form meiotic spindles, although the chromosomes condensed and became localized to cortices where microvillus-free areas developed. Cytochalasin B-treated mature eggs maintained intact meiotic spindles and exhibited a disappearance of microvillus-free areas and a reduction in cortical actin. The chromosomes of mature eggs treated with colchicine remained associated with microvillus-free areas despite the disappearance of meiotic spindles. Occasionally, colchicine-treated eggs possessed more than one cortically located mass of chromosomes, each of which was associated with a microvillus-free area. These observations indicate that mechanisms involving the movement of the meiotic spindle to the oocyte cortex and development and maintenance of cortical polarity are cytochalasin B sensitive. Commensurate with the localization of meiotic chromosomes to the egg cortex is the reorganization of cortical actin and the formation of a microvillus-free area.     

Cytochalasin B blocks sperm incorporation but allows activation of the sea urchin egg

Cytochalasin B (CB) (2 × 10⁻⁶ M) prevents the incorporation of sperm into the eggs of Lytechinus pictus and Strongylocentrotus purpuratus as judged by light and transmission electron microscopy (TEM). At lower concentrations of CB (2 × 10⁻⁷ M), sperm are successfully incorporated into the egg, but their migration in the area of the egg cortex is impaired. The site of action of CB on the sperm may be on the initial rotation of the sperm nucleus in the cortex; the subsequent migration is not affected by CB. Although sperm incorporation is prevented at the higher CB concentrations, the eggs become activated—as judged by cortical reaction, increased protein synthesis and increased respiration. These findings raise the concept that egg activation by sperm could result from some pre-fusion event and hence that sperm-egg fusion would not be a prerequisite for the triggering of development. An alternative hypothesis is that fusion occurs between the acrosome process membrane and egg membrane, but since CB has destroyed the integrity of the cortex actin, the fusion bridge is so weak that it cannot be maintained without some contractile or cytoskeletal support by the cortex. The sperm may activate the CB-treated egg in the same manner as pricking with a microelectrode sometimes does.     

The ultrastructure of the sea urchin egg cortex isolated before and after fertilization

The three-dimensional organization of cortices isolated from unfertilized and fertilized Strongylocentrotus purpuratus eggs has been examined by several techniques of light and electron microscopy. It has been found that when moderate shear forces are used, the isolated unfertilized egg cortex, in addition to cortical granules, contains acidic vesicles and an elaborate network of rough endoplasmic reticulum. This network provides a physical link between the cell surface and several kinds of cytoplasmic organelles (mitochondria, yolk granules, acidic vesicles) which are retained as part of the isolated cortex when gentle shear forces are applied. Furthermore a good visualization of actin in the cortex is provided: it is present as short filaments and mostly within the stubby microvilli of the egg. Finally, it has been noted that plaques exist on the inside face of the plasma membrane ready to assemble into typical clathrin coats that prefigure the burst of coated vesicle endocytosis that takes place after fertilization. The cortex isolated soon after fertilization is shown to contain coated pits and a scaffolding of filaments (mostly actin) in which many acidic vesicles are embedded.     

Assessment of the consumption of walleye pollock eggs by nekton and jellyfish in the northern Sea of Okhotsk in the spring

In the northern Sea of Okhotsk, nekton and jellyfish consumed as many as 831 × 10⁹ walleye pollock eggs per day in 2011. The nekton exerted the highest pressure, viz., 98.3% of the overall predation on pollock egg by aquatic animals. Of the entire quantity of consumed eggs, 55.9% were eaten by herring, 35.9% by walleye pollock, 6.5% by Sakhalin sole, and 1.7% by jellyfish. Among jellyfish, scyphomedusae Cyanea capillata and Chrysaora melonaster, as well as the hydromedusa Tima sachalinensis consumed the largest quantities of eggs. The total consumption of pollock egg by aquatic animals in 2011 was estimated at 42.4 × 10¹², or 11.4% of the entire quantity of eggs that were spawned by walleye pollock in the waters of the northern part of the sea. The total amount of pollock eggs that were eaten by herring and pollock together for 51 days in 2011 amounted to 38.9 × 10¹², which was 5.7 times as much as that in 2002. Thus, a significant growth of predation on pollock eggs by their main consumers, viz., herring and walleye pollock, was observed in 2011. This was caused by an increase in the populations of both species during the recent years and also by a higher concentration of pollock eggs.     

Actin cytoskeleton modulates calcium signaling during maturation of starfish oocytes

Before successful fertilization can occur, oocytes must undergo meiotic maturation. In starfish, this can be achieved in vitro by applying 1-methyladenine (1-MA). The immediate response to 1-MA is the fast Ca²⁺ release in the cell cortex. Here, we show that this Ca²⁺ wave always initiates in the vegetal hemisphere and propagates through the cortex, which is the space immediately under the plasma membrane. We have observed that alteration of the cortical actin cytoskeleton by latrunculin-A and jasplakinolide can potently affect the Ca²⁺ waves triggered by 1-MA. This indicates that the cortical actin cytoskeleton modulates Ca²⁺ release during meiotic maturation. The Ca²⁺ wave was inhibited by the classical antagonists of the InsP₃-linked Ca²⁺ signaling pathway, U73122 and heparin. To our surprise, however, these two inhibitors induced remarkable actin hyper-polymerization in the cell cortex, suggesting that their inhibitory effect on Ca²⁺ release may be attributed to the perturbation of the cortical actin cytoskeleton. In post-meiotic eggs, U73122 and jasplakinolide blocked the elevation of the vitelline layer by uncaged InsP₃, despite the massive release of Ca²⁺, implying that exocytosis of the cortical granules requires not only a Ca²⁺ rise, but also regulation of the cortical actin cytoskeleton. Our results suggest that the cortical actin cytoskeleton of starfish oocytes plays critical roles both in generating Ca²⁺ signals and in regulating cortical granule exocytosis.     

Spatial distribution of messenger RNA in the cytoskeletal framework of ascidian eggs

Maternal poly(A)⁺RNA, histone mRNA, and actin mRNA exhibit unique spatial distributions in the different ooplasmic regions of ascidian eggs. These RNAs also appear to migrate with their respective ooplasms during the episode of extensive cytoplasmic rearrangement that occurs after fertilization, suggesting they are associated with a structural framework. The role of the cytoskeletal framework (CF) in determining the spatial distribution of maternal mRNA was tested by subjecting Triton X-100 extracted (Styela plicata) eggs and early embryos to in situ hybridization with poly(U) and cloned DNA probes. Grain counts indicated that substantial proportions of the egg poly(A)⁺RNA, histone mRNA, and actin mRNA were present in the CF and that there was no alteration in the extent of mRNA-CF interactions during the period between fertilization and the two-cell stage. Analysis of grain distributions indicated that poly(A)⁺RNA, histone mRNA, and actin mRNA were concentrated in the same regions of detergent-extracted eggs as they are in intact eggs. The proportions and spatial distribution of these RNAs in the CF were not affected when the actin cytoskeleton was destabilized by cytochalasin B or DNAse I. The data suggest that maternal mRNA is associated with the CF, that this association is responsible for mRNA rearrangement during ooplasmic segregation, and that mRNA-CF interactions are not dependent on the integrity of the actin cytoskeleton.     

Ultrastructural Basis of the Tension Increase in Sea-Urchin Eggs Prior to Cytokinesis

Cortices of sea-urchin eggs were studied by electron microscopy to identify the structure responsible for the rise in tension at the egg surface prior to cleavage. During anaphase the tension increased and fine filaments of 70–90 Å in diameter appeared in the cell cortex forming a thin mesh-work beneath the cell membrane. The meshwork spread all around the egg cortex without reference to the mitotic axis and the number of filaments seemed to increase up to telophase. Immediately before appearance of the cleavage furrow, the meshwork in the anticipated furrow region became dense. As the furrow appeared the tension began to decrease and the meshwork disappeared. In the progressing furrow region fine filaments of the same size as that of the meshwork-filament were oriented in a bundle to form a contractile ring. Treatment with cytochalasin B suppressed both the tension increase and the formation of the filamentous meshwork. These results suggest that the component filament of the meshwork is an actin microfilament, and that the tension increase at anaphase is due to formation of a meshwork of actin microfilaments from which a contractile ring is subsequently derived at late telophase.     

Integrins on eggs: focal adhesion kinase is activated at fertilization,forms a complex with integrins,and is necessary for cortex formation and cell cycle initiation

We investigate the proposal that integrins and focal adhesion kinase (FAK) form a complex that has structural and signaling functions in eggs. FAK protein is present in eggs and is phosphorylated at fertilization. pY³⁹⁷FAK localizes to the membrane 30 min after fertilization, which correlates with the expression of βC integrins and egg cortex development. The βC integrin and pY³⁹⁷FAK coimmunoprecipitate from egg cortex lysates. PF573 228 and Y11, inhibitors of FAK, interfere with pronuclear fusion and reduce the abundance of pY³⁹⁷FAK and cortical actin without affecting microvillar actin. Cyclin E normally accumulates in the nucleus 15 min after fertilization, then returns to background levels. PF573 228– or Y11-treated eggs accumulate cyclin E in the nucleus; however, levels remain high. In addition, PF573 228 interferes with the accumulation of pERK1/2 in the nucleus and in eggs initiating mitosis. Injection of eggs with a fusion protein consisting of the focal adhesion–targeting domain of FAK fused to green fluorescent protein interferes with cortex formation and produces abnormal nuclei. These data indicate that an integrin–FAK adhesion complex forms at the egg surface that functions in formation of actin arrays in the egg cortex and provides signaling inputs for cell cycle initiation.     

Redistribution and Increase in Cortical Inositol 1,4,5-Trisphosphate Receptors after Meiotic Maturation of the Mouse Oocyte

Mouse oocytes develop sensitivity to inositol 1,4,5-trisphosphate (IP₃) during oocyte maturation. We recently reported that a change in the organization of the endoplasmic reticulum (ER) during oocyte maturation may contribute to this enhanced sensitivity (Mehlmannet al.,1995,Dev. Biol.170, 607–615). Here, we investigated whether there is an increase in the number of available IP₃receptors after maturation and whether there is a redistribution of IP₃receptors similar to the redistribution of the ER that occurs during maturation. Western blot analysis of the IP₃receptor in oocytes and eggs demonstrated a 1.8-fold increase in immunoreactive mass of the IP₃receptor following oocyte maturation. Microinjection of the function-blocking monoclonal antibody 18A10 inhibited IP₃-induced Ca²⁺release in a concentration-dependent manner in both eggs and oocytes. More antibody was required to inhibit Ca²⁺release to the same extent in eggs compared to oocytes when both were injected with the same concentration of IP₃, suggesting that eggs contain a greater number of functional IP₃receptors. Immunolocalization of the IP₃receptor revealed that receptors were present in large clusters, 1–2 μm in diameter, in the cortex of the mature egg except in a ring-shaped band of cortex adjacent to the meiotic spindle. In contrast, receptor clusters were located around the entire cortex of the immature oocyte and were much smaller (<1 μm); larger patches were sometimes seen, but they did not display the same spherical organization as those in eggs. These results suggest that the number of cortical IP₃receptors increases during mouse oocyte maturation and that this increase may contribute to enhanced Ca²⁺release at fertilization.     

Bee1, a Yeast Protein with Homology to Wiscott-Aldrich Syndrome Protein, Is Critical for the Assembly of Cortical Actin Cytoskeleton

Yeast protein, Bee1, exhibits sequence homology to Wiskott-Aldrich syndrome protein (WASP), a human protein that may link signaling pathways to the actin cytoskeleton. Mutations in WASP are the primary cause of Wiskott-Aldrich syndrome, characterized by immuno-deficiencies and defects in blood cell morphogenesis. This report describes the characterization of Bee1 protein function in budding yeast. Disruption of BEE1 causes a striking change in the organization of actin filaments, resulting in defects in budding and cytokinesis. Rather than assemble into cortically associated patches, actin filaments in the buds of Δbee1 cells form aberrant bundles that do not contain most of the cortical cytoskeletal components. It is significant that Δbee1 is the only mutation reported so far that abolishes cortical actin patches in the bud. Bee1 protein is localized to actin patches and interacts with Sla1p, a Src homology 3 domain–containing protein previously implicated in actin assembly and function. Thus, Bee1 protein may be a crucial component of a cytoskeletal complex that controls the assembly and organization of actin filaments at the cell cortex.     

Actin,more than just a housekeeping protein at the scene of fertilization

Since the first demonstration of sperm entry into the fertilized eggs of Mediterranean sea urchin Paracentrotus lividus by Hertwig (1876), enormous progress and insights have been made on this topic. However, the precise molecular mechanisms underlying fertilization are largely unknown. The two most dramatic changes taking place in the zygote immediately after fertilization are: (i) a sharp increase of intracellular Ca²⁺ that initiates at the sperm interaction site and traverses the egg cytoplasm as a wave, and (ii) the concomitant dynamic rearrangement of the actin cytoskeleton. Traditionally, this has been studied most extensively in the sea urchin eggs, but another echinoderm, starfish, whose eggs are much bigger and transparent, has facilitated experimental approaches using microinjection and fluorescent imaging methodologies. Thus in starfish, it has been shown that the sperm-induced Ca²⁺ increase in the fertilized egg can be recapitulated by several Ca²⁺-evoking second messengers, namely inositol 1,4,5-trisphosphate (InsP3), cyclic ADP-ribose (cADPr) and nicotinic acid adenine dinucleotide phosphate (NAADP), which may play distinct roles in the generation and propagation of the Ca²⁺ waves. Interestingly, it has also been found that the dynamic rearrangement of the actin cytoskeleton in the fertilized eggs plays pivotal roles in guiding monospermic sperm entry and in the fine modulation of the intracellular Ca²⁺ signaling. As it is well known that Ca²⁺ regulates the structure of the actin cytoskeleton, our finding that Ca²⁺ signaling can be reciprocally affected by the state of the actin cytoskeleton raises an intriguing possibility that actin and Ca²⁺ signaling may form a ‘positive feedback loop’ that accelerates the downstream events of fertilization. Perturbation of the cortical actin networks also inhibits cortical granules exocytosis. Polymerizing actin bundles also compose the ‘acrosome process,’ a tubular structure protruding from the head of fertilizing sperm. Hence, actin, which is one of the most strictly conserved proteins in eukaryotes, modulates almost all major aspects of fertilization.     

THE POLYMERIZATION OF ACTIN: ITS ROLE IN THE GENERATION OF THE ACROSOMAL PROCESS OF CERTAIN ECHINODERM SPERM

When Asterias or Thyone sperm come in contact with egg jelly, a long process which in Thyone measures up to 90 µm in length is formed from the acrosomal region. This process can be generated in less than 30 s. Within this process is a bundle of microfilaments. Water extracts prepared from acetone powders of Asterias sperm contain a protein which binds rabbit skeletal muscle myosin forming a complex whose viscosity is reduced by ATP. Within this extract is a protein with the same molecular weight as muscle actin. It can be purified either by collecting the pellet produced after the addition of Mg⁺⁺ or by reextracting an acetone powder of actomyosin prepared by the addition of highly purified muscle myosin to the extract. The sperm actin can be polymerized and by electron microscopy the polymer is indistinguishable from muscle F-actin. The sperm actin was shown to be localized in the microfilaments in the acrosomal processes by: (a) heavy meromyosin binding in situ, (b) sodium dodecyl sulfate (SDS) gel electrophoresis of the isolated acrosomal processes and a comparison to gels of flagella which contain no band corresponding to the molecular weight of actin, and (c) SDS gel electrophoresis of the extract from isolated acrosomal caps. Since the precursor for the microfilaments in the unreacted sperm appears amorphous, we suspected that the force for the generation of the acrosomal process is brought about by the polymerization of the sperm actin. This supposition was confirmed, for when unreacted sperm were lysed with the detergent Triton X-100 and the state of the actin in the sperm extract was analyzed by centrifugation, we determined that at least 80% of the actin in the unreacted sperm was in the monomeric state.     

A test for masked message: the template activity of messenger ribonucleoprotein particles isolated from sea urchine eggs

The hypothesis that the “masked message” of unfertilized eggs consists of nontranslatable mRNP particles was directly tested by in vitro translation of mRNPs in a system derived from wheat germ. Three classes of mRNPs were tested: particles prepared from sea urchin eggs in buffers containing 0.35 M K⁺, particles prepared from sea urchin eggs in 0.35 M Na⁺, and particles released with EDTA in 0.35 M K⁺ from polysomes of sea urchin embryos cultured in the presence of actinomycin D. The mRNA content of particles was monitored by determination of poly(A) content. The wheat germ system used is quantitatively stimulated by addition of mRNA derived from eggs or from any of the classes of mRNPs used. Particles prepared from eggs with Na⁺ or released from polysomes contain less protein than particles isolated from eggs in K⁺, and as expected these particles are fully translatable in vitro. Particles prepared from eggs in buffers containing 0.35 M K⁺ produce little or no stimulation in the in vitro system. That this lack of translation represents in vivo masking is indicated by several considerations: (1) The nontranslatable particles were prepared in 0.35 M K⁺ and 5 mM Mg²⁺, ion concentrations similar to those found in echinoderm eggs; (2) density and sedimentation rate characteristics of the particles are little changed by isolation; (3) RNA extracted from isolated particles is fully translatable; and (4) particles prepared from polysomes or under conditions which destabilize RNPs are translatable. These data support the masking hypothesis for the protein synthesis repression system of eggs.     

Cortical vesicle exocytosis in isolated cortices of sea urchin eggs: Description of a turbidometric assay and its utilization in studying effects of different media on discharge

Cortices of unfertilized sea urchin eggs can be isolated in suspension and will discharge the attached cortical vesicles (CVs) in response to calcium. We describe a simple turbidometric assay for monitoring the Ca²⁺-induced discharge of these vesicles and also compare the discharge of vesicles isolated in a high salt medium (primarily KCl) with a medium more closely simulating the internal milieu of the cell (primarily potassium gluconate and glycine). Discharge in response to calcium is similar in both media, requiring approximately 6 μM calcium for one-half maximal discharge. There are, however, significant differences in morphology and protein composition of the two types of preparations (more proteins present in the glycine cortices) and also in the rate of discharge of the vesicles in response to calcium (KCl cortices with $\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ 6 sec as opposed to 30 sec in the glycine cortices). The glycine cortices gradually lose their ability to respond to calcium but retention of calcium sensitivity is considerably aided by inclusion of ATP in the media; ATP has no apparent effect on discharge of the KCl cortices. The glycine cortices, as opposed to the KCl cortices, exhibited variation in calcium sensitivity during the breeding season and in the number of vesicles which would not break down in response to added calcium (referred to as refractory vesicles). The question of which type of cortex preparation most closely simulates the in vivo situation is discussed, and the view is presented that the glycine cortices most closely resemble the in vivo situation.     

Fenestrated endothelium of the adrenal gland: Freeze-fracture studies

Little is known of how adrenal hormones pass from the interstitial to the vascular space. We have begun to examine the adrenal endothelium as a barrier to hormone passage, by the freeze-fracturing technique. The endothelium of both cortex and medulla is fenestrated. Fractures from both regions show endothelial cells to be extremely thin in regions where fenestrations are abundant. En face fractures show fenestrae disposed in tracts; the fenestrae reaching a distribution of 35/μ². In both cortex and medulla there are areas of continuous endothelium which contain caveolae. Structures believed to represent fenestra diaphragms contain randomly disposed particles and occasional pits. We have not identified in replicas the central ring and pore described in thin-sectioned material (Elfvin, 1965). The main differences between freeze-fractured aspects of cortical and medullary endothelium are the greater abundance of caveolae in the medulla and the size of the fenestrae (fenestra rims in the medulla are 525–780 Å in diameter; in the cortex 570–1660 Å). These differences may reflect the different embryological origins of the medulla and cortex. While caveolae may participate in hormone transport, there is no evidence for this. In the medulla the caveolae are more numerous and may have a function not necessarily related to transport. Possibly, caveolae play a role in processing hormones and related substances. For example, ATP and specific proteins are released as well as epinephrine during exocytosis from chromaffin cells. Epinephrine enters the vascular space but ATP does not. ATPase enzymes are a common feature of caveolae of other endothelia and may occur as well in adrenal endothelium.     

Structural similarity between actin bundles from characean algal cells and sea urchin oocytes

The structure of actin bundles from internodal cells of Chara australis, an algal plant, was studied by electron microscopy of negatively stained specimens and optical diffraction. Gently prepared bundles revealed paracrystalline structures resembling the Mg²⁺-induced paracrystals of rabbit skeletal muscle actin (Hanson, 1968). In addition, the algal actin bundles sometimes had transverse striations at intervals of about 130 Å, as has been observed in actin bundles from sea urchin eggs (DeRosier et al., 1977; Spudich & Amos, 1979) and sea urchin coelomocytes (De Rosier & Edds, 1980; Otto & Bryan, 1981). This finding suggests that a common mechanism might be working in a variety of cells to organize actin filaments into functional bundles.     

Wave of cortical actin polymerization in the sea urchin egg

The distribution of actin filaments in the cortical layer of sea urchin eggs during fertilization has been investigated by light microscopy using fluorescently labeled phallotoxins. The cortical layer of both whole eggs and cortices isolated on a glass surface was examined. In cortices of unfertilized eggs, numerous fluorescent spots were seen, which may correspond to short actin filament cores in microvilli. After insemination, one of the sperm-attaching points on the egg surface first became strongly fluorescent. This fluorescence grew around the point of sperm penetration with the growth of the fertilization cone. Then, the cortical layer of the egg around the fertilization cone became strongly fluorescent and the fluorescence propagated in a wavelike manner over the entire cortex. The mechanism of the propagation of actin polymerization is discussed.     

Effects of Cytochalasin B on Muscle Cells in Tissue Culture

THE antibiotic cytochalasin B induces the formation of binucleated and multinucleated cells by preventing cytokinesis¹ and the cleavage furrow filaments in sea urchin eggs, 50-100 Å in diameter, disappear in the presence of this compound². It has a similar effect on the fine filaments found in embryonic pancreatic cells³ and inhibits cell migration¹. Many kinds of cells displaying amoeboid movements have been reported to have 50-100 Å filaments^4–6. Ishikawa et al. have demonstrated that 60–80 Å filaments in the cortex of various tissue cells bind heavy-meromyosin in a manner identical to that of actin filaments, which are also 60–80 Å in diameter⁷. Many investigators have referred to these thin filaments as actin or actin-like and assumed them to be responsible for, or associated with, cellular movements^8,9, contraction^10,11 and cytokinesis^12,13.